CVJA Volume 27 Issue 4 by Clinics Cardive Publishing

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CardioVascular Journal of Africa (official journal for PASCAR)

• Fine-tuning Heart Assist 5 devices with echocardiography • Sirtuin 1 in South African Indians with early-onset CAD • Management of acute MI patients at Nairobi Hospital • Acquired von Willebrand syndrome and stenosis • Remedy ® biodegradable peripheral stents • Coated or uncoated oxygenators during CPB surgery

Perindopril has proven outcomes in: • Coronary Artery Disease1 • Acute Myocardial Infarction2 • Stroke3 • End-stage Renal Failure4

• ECG findings in HIV patients in south-east Nigeria

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Pearinda 4. Each tablet contains 4 mg perindopril tert-butylamine. Reg. No.: RSA S3 A41/7.1.3/0649. NAM NS2 10/7.1.3/0476. Pearinda 8. Each tablet contains 8 mg perindopril tert-butylamine. Reg. No.: RSA S3 A41/7.1.3/0650. NAM NS2 10/7.1.3/0477. For full prescribing information, refer to the package insert approved by the Medicines Control Council, April 2009. Pearinda Plus 4. Each tablet contains 4 mg perindopril tertbutylamine and 1,25 mg indapamide. Reg. No.: RSA S3 A41/7.1.3/0633. NAM NS2 10/7.1.3/0611. For full prescribing information, refer to the package insert approved by the Medicines Control Council, April 2010. 1) The EUROPA study Investigators. “Efficacy of perindopril in reduction of cardiovascular events among patients with stable coronary artery disease: randomized, double-blind, placebo-controlled, multicentre trial (the EUROPA study)”. The Lancet 2003;362:782-788. 2) The PREAMI study Investigators. “Effects of angiotensin-converting enzyme inhibition with perindopril on left ventricular remodelling and clinical outcome. Results of the randomized perindopril and remodelling in elderly with acute myocardial infarction (PREAMI) study”. Arch Intern Med 2006;166:659-666. 3) PROGRESS Collaborative Group. “Randomised trial of a perindopril-based blood-pressurelowering regimen among 6105 individuals with previous stroke or transient ischaemic attack”. The Lancet 2001;358:1033-41. 4) Guerin AP, et al. “Impact of Aortic Stiffness Attenuation on Survival of Patients in End-Stage Renal Failure”. Circulation 2001;103;987-992. 5) Department of Health website http//www.doh.gov.za – Accessed on 29/03/2016. PAF291/04/2016.

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Cardiovascular Journal of Africa . Vol 27, No 4, July/August 2016

the affordable5 endurance ACE-inhibitor

• Management of arterial hypertension in Benin • Left ventricular assist device for advanced heart failure

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For further product information contact PHARMA DYNAMICS P O Box 30958 Tokai Cape Town 7966 Tel 021 707 7000 Fax 021 701 5898 Email info@pharmadynamics.co.za CUSTOMER CARE LINE 0860 PHARMA (742 762) www.pharmadynamics.co.za Fedaloc SR 30 mg. Each slow release tablet contains 30 mg Nifedipine. Reg. No.: RSA S3 A37/7.1/0302. NAM NS2 10/7.1/0033. Fedaloc SR 60 mg. Each slow release tablet contains 60 mg Nifedipine. Reg. No.: RSA S3 A37/7.1/0303. NAM NS2 10/7.1/0034. For full prescribing information, refer to the package insert approved by the Medicines Control Council, 25 November 2011. 1) IMS Data, December 2015; Units sold in the period 1 January 2015 – 31 December 2015. 2) Department of Health website http://www.hoh.gov.za – Accessed 10/03/2016. * Calculated cost for 30 tablets. FCF293/04/2016

ISSN 1995-1892 (print) ISSN 1680-0745 (online)

Vol 27, No 4, JULY/AUGUST 2016

CONTENTS

Cardiovascular Journal of Africa 207

www.cvja.co.za

From the Editor’s Desk P Commerford

Cardiovascular Topics 208

Fine-tuning management of the Heart Assist 5 left ventricular assist device with two- and three-dimensional echocardiography ZT Demirozu • N Arat • DS Kucukaksu

213 Sirtuin 1 rs1467568 and rs7895833 in South African Indians with early-onset coronary artery disease P Ramkaran • A Phulukdaree • S Khan • D Moodley • AA Chuturgoon 218 Assessment of the management of acute myocardial infarction patients and their outcomes at the Nairobi Hospital from January 2007 to June 2009 R Kimeu • C Kariuki 222 Acquired von Willebrand syndrome in children with aortic and pulmonary stenosis FK Binnetoğlu • K Babaoğlu • ŞG Filiz • E Zengin • G Altun • SÇ Kılıç • N Sarper 228

Correlation analysis between ApoM gene-promoter polymorphisms and coronary heart disease Y Zhang • L-Z Huang • Q-L Yang • Y Liu • X Zhou

238

Clinical features and patency rates of Remedy® biodegradable peripheral stents SK Tiryakioglu • O Tiryakioglu • O Karahan • S Demirtas • F Gokalp • K Erkoc • H Özkan • A Ozyazicioglu

242 Humoral immune response and coated or uncoated oxygenators during cardiopulmonary bypass surgery SO Karakisi • AG Kunt • Ş Bozok • İ Çankaya • M Kocakulak • U Muşabak • MF Sargon • Ş Ergene • G İlhan • H Karamustafa • N Tufekci • E Şener 246 High aortic pulse-wave velocity may be responsible for elevated red blood cell distribution width in overweight and obese people: a community-based, cross-sectional study IH Altiparmak • ME Erkus • A Kocarslan • H Sezen • O Gunebakmaz • Y Sezen • Z Kaya • A Yildiz • R Demirbag

INDEXED AT SCISEARCH (SCI), PUBMED, PUBMED CENTRAL AND SABINET

Editors

SUBJECT Editors

Editorial Board

Editor-in-Chief (South Africa) Prof Pat Commerford

Nuclear Medicine and Imaging DR MM SATHEKGE

prof PA Brink Experimental & Laboratory Cardiology

PROF A LOCHNER Biochemistry/Laboratory Science

PROF R DELPORT Chemical Pathology

PROF BM MAYOSI Chronic Rheumatic Heart Disease

Assistant Editor Prof JAMES KER (JUN) Regional Editor DR A Dzudie Regional Editor (Kenya) Dr F Bukachi Regional Editor (South Africa) PROF R DELPORT

Heart Failure Dr g visagie Paediatric dr s brown Paediatric Surgery Dr Darshan Reddy Renal Hypertension dr brian rayner Surgical dr f aziz Adult Surgery dr j rossouw Epidemiology and Preventionist dr ap kengne Pregnancy-associated Heart Disease Prof K Sliwa-hahnle

PROF MR ESSOP Haemodynamics, Heart Failure DR MT MPE Cardiomyopathy & Valvular Heart Disease DR OB FAMILONI Clinical Cardiology DR V GRIGOROV Invasive Cardiology & Heart Failure

International Advisory Board PROF DAVID CELEMAJER Australia (Clinical Cardiology) PROF KEITH COPELIN FERDINAND USA (General Cardiology) DR SAMUEL KINGUE Cameroon (General Cardiology)

PROF DP NAIDOO Echocardiography

DR GEORGE A MENSAH USA (General Cardiology)

PROF B RAYNER Hypertension/Society

PROF WILLIAM NELSON USA (Electrocardiology)

PROF MM SATHEKGE Nuclear Medicine/Society PROF J KER (SEN) Hypertension, Cardiomyopathy, PROF YK SEEDAT Cardiovascular Physiology Diabetes & Hypertension

DR ULRICH VON OPPEL Wales (Cardiovascular Surgery)

DR J LAWRENSON Paediatric Heart Disease

PROF ERNST VON SCHWARZ USA (Interventional Cardiology)

PROF H DU T THERON Invasive Cardiology

PROF PETER SCHWARTZ Italy (Dysrhythmias)

Vol 27, No 4, JULY/AUGUST 2016

CONTENTS

252 Electrocardiographic findings in a cross-sectional study of human immunodeficiency virus (HIV) patients in Enugu, south-east Nigeria BJC Onwubere • PO Njoku • EC Ejim • BC Anisiuba • SO Ike 258

Comparison of primary repair and patch plasty procedure on the P wave in adult atrial septal defect closure A Ucak • V Temizkan • M Ugur • AE Yedekci • O Uz • A Selcuk • AT Yilmaz

262

Carotid characteristics of black South Africans with five-year sustained hypertension M Maritz • CMT Fourie • JM van Rooyen • HW Huisman • AE Schutte

270

Frontline initiatives in early myocardial reperfusion with ST-elevation myocardial infarction

PUBLISHED ONLINE (Available on www.cvja.co.za and in Pubmed)

Cardio News

Cardiovascular Topics e1 Management of arterial hypertension in Cotonou city, Benin: general practitioners’ knowledge, attitudes and practice (Prise en charge des hypertendus dans la ville de Cotonou (Bénin) en 2011: connaissances attitudes et pratiques des médecins généralistes) LH Codjo • MD Houenassi • D Dokoui • SHM Dohou • A Wanvoegbe • A Agbodande • AC Attinsounon • A Alassani • S Ahoui • AC Dovonou • TA Adoukonou

Case Reports e7 A rare giant pericardial cyst mimicking a paracardiac mass H Akbayrak • S Yildirim • M Simsek • M Oc e10 Simultaneous presentation of giant aneurysms of the coronary sinus and superior vena cava Y Cheng • H Gao • Z Zheng • Y Mou e13 Successful continuous-flow left ventricular assist device implantation with adjuvant tricuspid valve repair for advanced heart failure C Lee • J Wei e16 An unusual condition during internal jugular vein catheterisation: vertebral artery catheterisation Ozge Korkmaz • Sabahattin Göksel • Burçak Söylemez • Kasım Durmuş • Ahmet Cemil İşbir • Öcal Berkan

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The Cardiovascular Journal of Africa, incorporating the Cardiovascular Journal of South Africa, is published 10 times a year, the publication date being the third week of the designated month.

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CARDIOVASCULAR JOURNAL OF AFRICA • Volume 27, No 4, July/August 2016

207

From the Editor’s Desk Acquired von Willebrand syndrome (AVWS) is a rare clinical condition characterised by prolonged bleeding time and decreased levels of factor VIII and von Willebrand factor. It has been reported to occur in patients with severe aortic stenosis and other cardiac conditions associated with high shear stress, such as para-valvar leaks after prosthetic valve replacement surgery. First reported decades ago, the mechanism was initially unclear but it has now become known. In this issue, Binnetoğlu and colleagues (page 222) report on a prospective series of children with aortic and pulmonary stenosis, and describe the frequency of occurrence and underlying pathophysiology. There is general agreement on the considerable variation in the prevalence of various cardiac diseases among the different ethnic groups in Africa and there may also be differences in the manner in which the diseases express themselves. There is less clarity about the contributions of intrinsic ‘genetic’ differences to that variability compared to acquired or environmental factors. Sirtuin 1 (SIRT1) has been identified as a candidate molecule affecting the epigenetic mechanisms of cardiovascular disease (CVD). Previous studies have shown that some SIRT1 singlenucleotide polymorphisms (SNPs) are associated with body mass index, diabetes, blood pressure, cholesterol metabolism and coronary artery calcification. An investigation conducted by Ramkaran and co-workers (page 213) in young South African Indians with coronary disease concluded that SNP variant alleles occurred more frequently in South African Indians than in black South Africans. The study is not large enough to definitively assess whether these variants may serve as risk factors that contribute to Indians developing early-onset CVD but the results are intriguing and warrant further investigation. Technological advances in many aspects of the management of CVD continue to amaze those of us who have been privileged to witness their evolution and development. Left ventricular assist devices (LVADs) and two- and three-dimensional echocardiography were the stuff of dreams when many of us were training but now are part of routine care in some parts of the world. Demirozu and co-workers (page 208) elegantly demonstrate how the use of advanced imaging techniques of echocardiography can be used to fine-tune the functioning of LVADs. Intravascular stenting was equally unthinkable only a few decades ago but has now revolutionised much of the management of CVD. The way in which a new generation of biodegradable stents promises to advance this area even further is described by Tiryakioglu et al. (page 238). Cardiologists and physicians trained in an era prior to the introduction of the technological advances mentioned above

Professor PJ Commerford

will be pleased to see that the old stalwart, electrocardiography (ECG) continues to be widely used. Some consider it does not receive the recognition it deserves as a cheap, non-invasive adjunct to the clinical examination. A cross-sectional study carried out on adults in Nigeria examined the ECGs of 100 HIV-infected patients on highly active anti-retroviral therapy (HAART), 100 HIV-infected HAART-naïve patients and 100 HIV-negative controls (Njoko et al., page 252). The clinical relevance of these findings and that of similar findings reported by others and discussed by the authors, remains unclear and requires long-term follow up studies accompanied by imaging, either by echocardiography or perhaps more helpfully, by cardiac magnetic resonance imaging. PJ Commerford Editor-in-Chief

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Cardiovascular Topics Fine-tuning management of the Heart Assist 5 left ventricular assist device with two- and three-dimensional echocardiography Zumrut Tuba Demirozu, Nurcan Arat, Deniz Suha Kucukaksu

Abstract Introduction: Left ventricular assist device (LVAD) implantation is a viable therapy for patients with severe end-stage heart failure, providing effective haemodynamic support and improved quality of life. The Heart Assist 5 (Micromed Cardiovascular Inc, Houston, TX) continuous-flow LVAD has been on the market in Europe since May 2009. Methods: We evaluated nine Heart Assist 5 LVAD patients with two- and three-dimensional transthoracic echocardiographic (TTE) and transoesophageal echocardiographic (TEE) parameters between December 2011 and December 2013. The pre-operative TTE LVAD evaluations included left ventricular (LV) function and structure, quantification of right ventricular (RV) function and tricuspid regurgitation (TR), assessment of aortic and mitral regurgitation, and presence of patent foramen ovale and intra-cardiac clots. Peri-operative TEE determined the inflow cannula and septum position, and assessed the de-airing process while weaning from cardiopulmonary bypass. Post-operative serial follow-up TTE showed the surgical results of LVAD implantation, determined the overall structure and function of the LV, RV and TR, and observed the inflow and outflow cannula position. Results: Nine patients who had undergone Heart Assist 5 LVAD implantation and had been followed up for more than 30 days were included in this study. Eight patients had ischaemic cardiomyopathy and one had adriamycin-induced cardiomyopathy. Pre-implantation data: the mean age of the patients was 52 ± 13 (34–64) years, mean body surface area (BSA) was 1.8 ± 0.2 (1.6–2.0) m2, mean cardiac index (CI) was 2.04 ± 0.4 (1.5–2.6) l/min/m2, mean cardiac output (CO) was 3.7 ± 0.7 (2.6–4.2) l/min, mean ejection fraction (EF) was 23 ±

Department of Heart Transplant and Mechanical Circulatory Support, Istanbul Bilim Medical Faculty, Sisli Florence Nightingale Hospital, Istanbul, Turkey Zumrut Tuba Demirozu, MD Deniz Suha Kucukaksu, MD

Department of Cardiovascular Surgery, Koc University Hospital, Istanbul, Turkey Zumrut Tuba Demirozu, MD, tdemirozu@yahoo.com

Department of Cardiology, Istanbul Bilim Medical Faculty, Sisli Florence Nightingale Hospital, Istanbul, Turkey Nurcan Arat, MD

5 (18–28)%, and right ventricular fractional area contraction (RVFAC) was 43 ± 9 (35–55)%. One patient had aortic valve replacement (AVR) during the LVAD implantation, and excess current alarms and increased power were suspected to be caused by a possible thrombus. Close follow up with TTE studies were carried out to clear the LV of thrombus formation, and the inflow cannula position was checked to maintain the septum in the midline, so preventing the suction cascade. Four patients were followed up for more than two years, and two were followed up for more than a year. Three patients died due to multi-organ failure. Follow-up speed-change TTE studies of six patients showed that the mean speed was 9 800 ± 600 (9 500–10 400) rpm, and mean CO was 4.7 ± 0.3 (4.3– 5.0) l/min during the three-month post-implant period. Conclusion: We believe that TTE can play a major role in managing LVAD patients to achieve optimal settings for each patient. A large series is mandatory for assessment of echocardiographic studies on Heart Assist 5 LVAD. Keywords: Heart Assist 5 LVAD, continuous-flow pumps, twoand three-dimensional echocardiography Submitted 11/2/15, accepted 14/11/15 Cardiovasc J Afr 2016; 27: 208–212

www.cvja.co.za

DOI: 10.5830/CVJA-2015-083

Left ventricular assist devices (LVADs) are the accepted treatment modality for advanced end-stage heart failure patients.1-5 LVADs have been reported to be effective therapy in supporting cardiovascular circulation and end-organ function for weeks and even years.1-3 Increased use of LVAD therapy for patients with advanced heart failure has revealed a new paradigm for this novel physiology. It has been reported that there is a knowledge gap on these new-generation, continuous-flow pumps and their effect on cardiac and end-organ physiology. There is a growing need for practising heart failure cardiologists and transplant surgeons who take care of these patients to evaluate the pumps and their effects in patients.1-7 Monitoring, tracking and management of these patients needs experience so that the pumps work more efficiently with minor side effects and complications. Echocardiography is the most important non-invasive imaging tool for LVAD assessment at pre-, peri- and post-

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Table 1. Demographic and pre-implant characteristics Parameters

Number of patients and mean ± SD

Gender Men

Women

Age, years

52 ± 13

Weight, kg

74 ± 12

Body surface area, m²

1.8 ± 0.2

Heart disease Ischaemic cardiomyopathy

Adriamycin-induced cardiomyopathy

Purpose of LVAD Bridge to transplantation

Diabetes mellitus

Hypertension

AICD-CRT-D implantation

Previous cardiac surgery CABG

Pre-implant support Intra-aortic balloon pump

Fig. 1. Heart Assist 5 LVAD implantation.

Fig. 2. Heart Attendant, the portable console.

23 ± 5

Left ventricular ejection fraction, % Cardiac index, l/min/m2

implant, and during long-term follow up.1-8 Serial, non-invasive echocardiographic studies would help management strategies of these patients and identify mechanical dysfunction timeously. The Heart Assist 5 LVAD (Micromed Cardiovascular Inc, Houston, TX) is small and compact, weighing 92 g and can easily be implanted above the diaphragm (Fig. 1). The Heart Assist 5 flow probe, an implantable ultrasonic flow, measures the blood flow quite accurately while blood is pumped through the outflow graft to the aorta. Changes in pump speed and electronic current provide important information about the volume of blood in the chambers and decompression of the heart, and thus minor and major side effects can easily be detected. Also remote monitoring of these patients helps with patient care, as the primary physicians are informed either by cellphone message or e-mail notification about marginal haemodynamic changes in LVAD parameters (Figs 2, 3). Although extensive data have been reported in the literature concerning echocardiographic evaluation of other continuousflow pumps, there are no data on the Heart Assist 5 LVAD and

Haemodynamic values 2.04 ± 0.4

Cardiac output, l/min

3.8 ± 0.7

PCWP, mmHg

30 ± 8

Coagulation profile Prothrombin time, sec

17.3 ± 7.5

International normalised ratio

1.58 ± 0.6

Partial thromboplastin time, sec

33.9 ± 2.8

Laboratory data 22 ± 10

BUN, mg/dl Creatinine, mg/dl

1.17 ± 0.3

Na, mEq/l

136 ± 2

Albumin, g/l

3.8 ± 0.5

Alkaline phosphatase, U/l

73 ± 25

SGOT, U/l

18 ± 10 26 ± 23

SGPT, U/l Total bilirubin, mg/dl

0.92 ± 0.5

SD = standard deviation; LVAD = left ventricular assist device; AICD = automatic implantable cardioverter defibrillator; CRT-D = cardiac resynchronisation therapy device; CABG = coronary artery bypass grafting; PCWP = pulmonary capillary wedge pressure; BUN = blood urea nitrogen; Na = sodium; SGOT = serum glutamic oxaloacetic transaminase; SGPT = serum glutamic pyruvic transaminase.

Fig. 3. Remote monitoring of Heart Assist 5.

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its long-term outcomes. The aim of our study was to describe this small, new-generation pump and the first implantation experience from Istanbul, Turkey.

Methods From December 2011 to December 2013, Heart Assist 5 LVADs were implanted in nine patients at our hospital. Eight patients had ischaemic cardiomyopathy and one had adriamycin-induced cardiomyopathy. The mean age of the patients was 53 ± 13 (34–64) years (Table 1). All patients’ pre-operative data (catheterisation, echocardiography, laboratory and infection parameters) and haemodynamic status had been assessed at the hospital’s medical review board meeting to evaluate their selection for LVAD implantation. All patients had signed informed consent and the study protocol was approved by the institutional review board. In December 2011, 55 serial two-dimensional transthoracic echocardiographic (TTE) examinations had been performed on these patients with the Vivid 3 (General Electric, Fairfield, Connecticut), and 16 serial three-dimensional TTE examinations had been done with the Philips IE33 xMATRIX (Royal Philips Electronics, Amsterdam, Netherlands). Nine transoesophageal echocardiograms (TEE) were done in the operating room under general anaesthesia during the LVAD implantation to evaluate inflow cannula and septum position, and to monitor the de-airing process while weaning from cardiopulmonary bypass. The specific protocol used for these echocardiographic studies included standard TTE parasternal, apical, subcostal and suprasternal notch views.1-13 All TTEs and TEEs were performed by certified cardiologists in different clinical settings. In hospital during the pre-implant period, they evaluated the intracardiac structure [patent foramen ovale (PFO), atrial septal defect (ASD)], chamber dimensions, left ventricular (LV) and right ventricular (RV) function, valvular structure and function, pericardial disease, volume status and abnormalities of the aorta. In hospital during the post-implant LVAD period, the aim was to visualise the septum and inflow cannula position, the intracardiac volume, outflow conduit, decompression of the left ventricle with rpm speed change and its effects on the aortic valve opening time. Doppler interrogation of the inlet cannula and outflow conduit flows were performed as described in the literature.10,12,13 Follow-up TTEs were done for clinical indications, including LV and RV ejection fraction (EF), RV fractional area contraction (RVFAC), unexplained change in haemodynamic status including minor dehydration, ventricular tachycardia attacks, suspicion of device malfunction, excess current alarms, and optimisation of LVAD speed. Echo-specific parameters and differences in image quality or artifact generation were not evaluated in our study. During all echocardiographic studies, the transplant surgeons were at the bedside or in the out-patient clinic, in communication with the heart failure cardiologist setting up the pump parameters, optimising the LVAD speed, and observing the aortic valve opening times and septum inflow cannula position to prevent the suction cascade. Images from 55 TTEs were retrospectively analysed by the heart failure cardiologist who had mainly performed all these echo studies. The inlet cannula/outflow conduit velocities were measured in m/s by spectral Doppler. The pulsality index of the pump, LV end-diastolic dimension (LVEDD), LV end-systolic

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dimension (LVESD), interventricular septum (IVS), posterior wall thickness, RVFAC, and tricuspid annular plane systolic excursion (TAPSE) were measured, and the functioning of the valves was routinely evaluated pre-implant and at the second week, and first, third, sixth, ninth and 12th months post-implant.

Statistical analysis All echocardiographic data were collected in parallel with the patients’ laboratory parameters (biochemical, blood count, coagulation profile) in an electronic database, and descriptive statistics were calculated using Microsoft Excel (Microsoft Corp, Redmond, Washington). Continuous variables were expressed as the mean value ± standard deviation.

Results As our Heart Assist 5 LVAD patient data were limited, we did not compare the patients’ outcome parameters pre- versus postimplant, knowing that with this small group, all analyses would be statistically non-significant. We therefore analysed these patients’ data as descriptive statistics. The titanium impeller of Heart Assist 5 LVAD is small, lightweight and located pericardially. The titanium housing enables direct visualisation of the impeller. The outflow and inflow cannulas were visualised in all studies. The inflow cannula and septum (left, right, neutral) position, outflow graft anastomosis to the aorta, aortic valve cusp status, and aortic valve opening times with speed changes were evaluated and visualised in all 55 echocardiographic studies (Figs 4–6). Pre-implant echocardiographic data of the six patients: mean EF was 23 ± 5 (18–28)%, mean LVEDD was 6.9 ± 0.6 (6.3–7.7) cm, mean LVESD was 5.8 ± 0.5 (5.1–6.4) cm, mean IVS was 0.9 ± 0.1 (0.9–1.1) cm, mean RVFAC was 43 ± 9 (35–55)%, and mean TAPSE was 17 ± 4 (13–23) mm. The 12-month post-implant echocardiographic data of the six patients were: mean EF was 19 ± 6 (10–25)%, mean LVEDD was 6.4 ± 0.4 (6.1–7.0) cm, mean LVESD was 5.6 ± 0.3 (5.2–6.0) cm, mean RVFAC was 35 ± 11(21–43), mean TAPSE was 13 ± 2 (11–16) mm, mean rpm was 9 800 ± 600 (9 500–10 400) rpm, mean pulsality index (PI) was 2.79 ± 1.7 (1.9–4.9) m/s, outflow cannula

Fig. 4. Outflow cannula velocity measurement with Doppler.

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Fig. 5. E valuation of aortic valve opening times with M mode TTE.

velocity was 0.9 ± 0.2 (0.8–1.2) m/s, and aortic valve opening was observed every one-to-one cycle or one-to-three cycle. Since eight patients had ischaemic cardiomyopathy and one had had coronary artery bypass grafting (CABG) 10 years earlier, remission of their end-stage heart failure was not expected. All our patients had had automated internal cardioverter defibrillator (AICD) and cardiac resynchronisation therapy (CRT-D) implantation prior to the LVAD implantation. One patient had a ventricular fibrillation (VF) event in the ward and had cardiopulmonary resuscitation. He had his AICDCRT-D implanted prior to the LVAD surgery. Two patients had experienced ventricular tachycardia (VT) at home due to dehydration during the summer, and their AICD had been interrupted during the attacks. There were a few hospitalisations because of minor complications. One patient complained of a coffee-grinding noise coming from his pump three months post-implant, and the remote monitoring system also documented frequent excess current alarms and increased power at different hours of the day. The patient was hospitalised for possible thrombus formation or device malfunction. The data were analysed at Micromed Cardiovascular Inc, Houston, Texas. The coagulation profile was in the normal range for a LVAD patient, his INR was 3.2, and his lactate dehydrogenase level was 606 U/l, which was gradually elevated up to 120 days after LVAD implantation. Serial echocardiographic studies were done weekly to rule out possible thrombus formation in the left ventricle, the orifice of the inflow cannula, outflow conduit and aortic valve cusps. The patient received heparin infusion with close monitoring of his coagulation profile. There was no absolute finding in the echo studies to confirm thrombus formation or determine the reason for the coffee-grinding noise, since we were not able to visualise the titanium impeller. We guessed that there may have been a blood clot there, which caused the increased power. The noise stopped after the fifth day of heparin infusion. A female patient had uncontrolled diabetes, she had gradually gained 7 kg by the sixth month post-operatively, and suffered from recurrent urinary tract infections. She was admitted with fever, tachycardia, post-diabetic ketoacidosis and documented urinary tract infection. Her echocardiographic study was done in a sub-optimal position due to her tachycardia. She received

Fig 6. Left ventricular inflow cannula visualisation with 3D TTE.

anti-arrhythmic and antibiotic regimens, and a few days later she was in sinus rhythm, her heart rate was 89–95 beats per minute, and her fever was gone. Her echo studies were repeated and LVAD optimisation was done. She was discharged from hospital with 9 500 rpm, her CO was 4.7 l/min and the pulsality index was 2.31 m/s. Four patients were followed up for more than two years, and two for more than a year. Three patients died due to multi-organ failure. One international patient had heart transplantation in the Republic of China. The longest supported patient was for more than three years, with a good quality of life.

Discussion All data for each patient had been collected in each portable console, Heart Attendant (Micromed Cardiovascular Inc, Houston, TX), so with the universal serial bus (usb) connection, all the data could be downloaded and could be easily analysed. Online connection of the Heart Attendant to the internet assists in patient care and provides 24 hours a day, seven days a week monitoring of the LVAD patient. This technology decreases hospital admissions for each alarm detection, as the clinicians can track the patient with the remote monitoring system and help him/her by phone. This technology differentiates Heart Assist 5 LVAD from other continuous-flow pumps. The implantable flow probe also helps to assess even minor dehydration problems, which can be solved without admission to hospital. We experienced low-flow alarms during the summer months and suggested our patients take in extra fluid during summer.15 Fine-tuning of the Heart Assist 5 LVAD with the implantable flow probe and portable console was convenient, and the LVAD speed change echo studies enabled easy optimisation of the device during in-hospital echocardiographic studies. There are reports of large single-centre echocardiographic studies of other continuous-flow and centrifugal pumps in the literature.8-13 These pumps have been approved by the United

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States Food and Drug Administration (FDA) and Conformitė Europėenne (CE), and have been on the market for a long time. Many centres have implanted these pumps in their end-stage heart failure patients all over the world. Long-term outcomes of these pumps have been published recently in the literature.1-13 Frazier et al. demonstrated the modified Micromed axial flow pump has an in vitro responsivity approaching that of the natural heart.16 Bluestein et al. reported that the thrombogenicity of Heart Assist 5 was 2.5-fold lower than the HeartMate II LVAD.17 Although our experience with Heart Assist 5 LVAD was not from a large series, compared with other continuous-flow and centrifugal pumps that we have implanted in our clinic, we tried to standardise pre-operative LVAD evaluations and post-implant follow up of LVAD patients in collaboration with the heart failure cardiologists. We believe that as a heart failure/transplant team, a systematic approach for the evaluation of these patients suffering from advanced heart failure is important. Close follow up of these patients will affect the success of LVAD surgery and long-term prognosis of the patients, which will lessen the side effects and enable rapid and accurate identification of mechanical and systemic malfunctions. Our group was the first to implant this version of Heart Assist 5 LVAD in Turkey. An earlier version of this pump was implanted in Turkey in April 2001.14 There were no data in the literature on the Heart Assist 5 LVAD using echocardiographic parameters and LVAD optimisation protocols during long-term follow up of the patients. In our clinical studies we were able to visualise the inflow cannula and outflow graft anastomoses to the aorta in most of the echocardiographic studies. The remote monitoring system and the implantable flow probe measurements were reliable, and tracking and monitoring of these patients outside of hospital was easy. Our study has limitations since there were so few patients with Heart Assist 5 LVAD support in our group. However, if other centres start implanting Heart Assist 5 LVAD in their end-stage heart failure patients, more data on these pumps and their longterm outcome will be available for clinicians in the future. This pump has had CE approval in Europe since May 2009, and the National Health Government solved the reimbursement issues of these continuous-flow pumps in 2012.

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Usefulness of two-dimensional echocardiographic parameters of the left side of the heart to predict right ventricular failure after left ventricular assist device implantation. Am J Cardiol 2012; 109(2): 246–251. 3.

Demirozu ZT, Etheridge WB, Radovancevic R, Frazier OH. Results of HeartMate II left ventricular assist device implantation on renal function in patients requiring post-implant renal replacement therapy. J Heart Lung Transplant 2011; 30: 182–187.

Noor MR, Bowles C, Banner NR. Relationship between pump speed and exercise capacity during HeartMate II left ventricular assist device support: influence of residual left ventricular function. Eur J Heart Fail 2012; 14(6): 613–620.

Raina A, Seetha Rammohan HR, Gertz ZM, Rame JE, Woo YJ, Kirkpatrick JN. Postoperative right ventricular failure after left ventricular assist device placement is predicted by preoperative echocardiographic structural, hemodynamic, and functional parameters. J Card Fail 2013; 19(1): 16–24.

Lund LH, Gabrielsen A, Tirén L, Hallberg A, El Karlsson K, Eriksson MJ. Derived and displayed power consumption, flow and pulsatility over a range of HeartMate II left ventricular assist device settings. Am Soc Artif Intern Organs J 2012; 58(3): 183–190.

Soleimani B, Haouzi A, Manoskey A, Stephenson ER, El-Banayosy A, Pae WE. Development of aortic insufficiency in patients supported with continuous flow left ventricular assist devices. Development of aortic insufficiency in patients supported with continuous flow left ventricular assist devices. Am Soc Artif Intern Organs J 2012; 58(4): 326–329.

Chapman CB, Allana S, Sweitzer NK, Kohmoto T, Murray M, Murray D, et al. Effects of HeartMate II left ventricular assist device as observed by serial echocardiography. Echocardiography 2013 Jan 11. doi: 10.1111/ echo.12100. [Epub ahead of print].

Mookadam F, Kendall CB, Wong RK, Kalya A, Warsame T, Arabia FA, et al. Left ventricular assist devices: physiologic assessment using echocardiography for management and optimization. Ultrasound Med Biol 2012; 38(2): 335–345.

10. Topilsky Y, Maltais S, Oh JK, Atchison FW, Perrault LP, Carrier M, et al. Focused review on transthoracic echocardiographic assessment of patients with continuous axial left ventricular assist devices. Cardiol Res Pract 2011; 187: 434. 11. Toda K, Fujita T, Domae K, Shimahara Y, Kobayashi J, Nakatani T. Late aortic insufficiency related to poor prognosis during left ventricular assist device support. Ann Thorac Surg 2011; 92(3): 929–934. 12. Topilsky Y, Hasin T, Oh JK, Borgeson DD, Boilson BA, Schirger JA, et al. Echocardiographic variables after left ventricular assist device

Conclusion

implantation associated with adverse outcome. Circ Cardiovasc Imaging 2011; 4: 648–661.

Our study was not designed for the appropriate combination of flow, power, pulsatily index and left ventricular unloading. It is impossible to establish or document cause–effect relationships while optimising LVAD parameters with so few patients. This study was an observational analysis of a new-generation pump. We believe that a concept should be studied in larger prospective series with different pumps using serial echocardiographic studies and comparing long-term outcomes of each pump.

13. Shah NR, Cevik C, Hernandez A, Gregoric ID, Frazier OH, Stainback

References

16. Frazier OH, Khalil HA, Benkowski RJ, Cohn WE. Optimization of

RF. Transthoracic echocardiography of the HeartWare left ventricular assist device. J Cardiac Fail 2012; 18: 745–748. 14. Kucukaksu DS, Sener E, Undar A, Noon GP, Tasdemir O. First Turkish experience with the Micromed DeBakey VAD. Texas Heart Inst J 2003; 30(2): 114–120. 15. Pektok E, Demirozu ZT, Arat N, Yildiz O, Oklu E, Eker D, et al. Remote monitoring of LVAD parameters after Heart Assist 5 implantation. Artif Organs 2013; 37(9): 820–825.

Ammar KA, Umdal MM, Kramer C, Sulamanjee N, Jan FM,

axial-pump pressure sensitivity for continuous-flow total artificial heart. J Heart Lung Transplant 2010; 29(6): 668–691.

Khandheria BK, et al. The ABCs of left ventricular assist device echo-

17. Chiu WC, Girdhar G, Xenos M, Alemu Y, Soares JS, Einou S,

cardiography: a systemic approach. Eur Heart J Cardiovasc Imaging

et al. Thromboresistance comparision of HeartMate II ventricular

2012; 13(11): 885–899.

assist device with the device thrombogenicity emulation optimized

Kato TS, Farr M, Schulze PC, Maurer M, Shahzad K, Iwata S, et al.

HeartAssist V VAD. J Biomech Eng 2014; 136(2): 021014.

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Sirtuin 1 rs1467568 and rs7895833 in South African Indians with early-onset coronary artery disease Prithiksha Ramkaran, Alisa Phulukdaree, Sajidah Khan, Devapregasan Moodley, Anil A Chuturgoon

Abstract Background: Sirtuin 1 (SIRT1), a class III histone deacetylase, has been identified as a candidate molecule affecting the epigenetic mechanisms of cardiovascular disease (CVD). Previous studies have shown that some SIRT1 single-nucleotide polymorphisms (SNPs) are associated with body mass index, diabetes, blood pressure, cholesterol metabolism and coronary artery calcification. We investigated two A>G SIRT1 SNPs, rs1467568 and rs7895833, in young South African (SA) Indians with coronary artery disease (CAD) and compared them to Indian and black controls. Methods: For rs1467568, a total of 287 subjects were recruited into this study (104 CAD patients, 99 age-, gender- and race-matched controls, and 84 age- and gender-matched black controls). For rs7895833, a total of 281 subjects were recruited into this study (100 CAD patients, 99 age-, gender- and race-matched controls, and 82 age- and gender-matched black controls). All patients were male, of Indian ethnicity, stable CAD confirmed on angiography, mean age 37.5 years; range 24–45. All subjects were genotyped using TaqMan SNP genotyping assays. Results: The variant allele for both SNPs was found at a higher frequency in the total Indian group compared to the total black population (rs1467568: 41 vs 18.5%, respectively, p < 0.0001, OR = 3.190, 95% CI: 2.058–40943; and rs7895833: 41 vs 22%, respectively, p < 0.0001, OR = 2.466, 95% CI: 1.620– 3.755). Indian controls presented with a higher frequency for both SNPs compared to black controls (rs1467568: 40 vs 18.5%, respectively, p < 0.0001, OR = 2.996, 95% CI: 1.850– 4.853; and rs7895833: 41 vs 22%, respectively, p < 0.0001, OR = 2.513, 95% CI: 1.578–4.004). No difference was seen in the distribution of both SNPs between CAD patients and either control group. We did not observe any association between the SNPs and clinical parameters in CAD patients and controls. Conclusion: Both SNP variant alleles occurred more frequently in SA Indians than in SA blacks. A larger study group and further analysis is required to assess whether these SIRT1 SNPs may serve as risk factors that contribute to Indians developing early-onset CAD.

Discipline of Medical Biochemistry and Chemical Pathology, University of KwaZulu-Natal, Durban, South Africa Prithiksha Ramkaran, BMedSc Hons Devapregasan Moodley, PhD Anil A Chuturgoon, PhD, chutur@ukzn.ac.za

Department of Physiology, School of Medicine, Faculty of Health Sciences, University of Pretoria, South Africa Alisa Phulukdaree, PhD

Department of Cardiology, Nelson R Mandela School of Medicine, University of KwaZulu-Natal, Durban, South Africa Sajidah Khan, MB ChB, FCP (SA), PhD

Keywords: sirtuin 1, rs1467568, rs7895833, single-nucleotide polymorphism, premature coronary artery disease, South African Indians Submitted 22/4/15, accepted 14/11/15 Cardiovasc J Afr 2016; 27: 213–217

www.cvja.co.za

DOI: 10.5830/CVJA-2015-085

Sirtuins are a class of NAD+-dependent proteins involved in a wide range of biological processes such as aging, transcription, apoptosis and inflammation.1 Sirtuin 1 (SIRT1) is located in the nucleus and cytoplasm, and plays an important role in epigenetic regulation by deacetylating a range of transcription factors to control downstream gene expression.2 The targets of SIRT1 include Forkhead box O (FOXO)1, (FOXO)3, peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1α), tumour suppressor p53, nuclear factorkappa B (NF-κB), Notch, hypoxia-inducible factor (HIF) 1α, liver X receptor (LXR), farnesoid X receptor (FXR) and sterol regulatory element-binding protein (SREBP)1c.3 Recent studies have demonstrated a protective role of SIRT1 in atherosclerosis, the underlying process of coronary artery disease (CAD).4 SIRT1 performs an anti-inflammatory function by downregulating the expression of several pro-inflammatory cytokines by interfering with the NF-κB signalling pathway. By deacetylating NF-κB, SIRT1 suppresses the expression of lectin-like oxidised low-density lipoprotein receptor-1 (Lox1), a scavenger receptor for oxidised low-density lipoproteins (oxLDL), therefore preventing foam cell formation.4 SIRT1 controls the activity of LXR, an important regulator of lipid homeostasis and inflammation.4 Activation of LXR results in expression of ATP-binding cassette (ABC) transporter ABCA1, which regulates the removal of cholesterol into high-density lipoproteins (HDL), a process known as reverse cholesterol transport (RCT). Dysfunctional RCT could lead to accumulation of cholesterol, thus stimulating foam cell production and the progression of atherosclerosis.4,5 Given the important role of SIRT1 in cardiovascular disease, research on genetic variation in the SIRT1 gene has become of interest. Genetic variations such as single-nucleotide polymorphisms (SNPs) in the SIRT1 gene have been associated with inflammation, body mass index, type 2 diabetes, blood pressure and dyslipidaemia, all of which are well-established risk factors for CAD.2,6-9 Coronary artery disease remains a leading cause of mortality worldwide, with an unusually high prevalence of earlyonset disease among the Indian population. South African (SA) Indians have a much higher prevalence of CAD compared to SA blacks.10 There are currently no studies on SIRT1 SNPs in SA Indians with CAD. We therefore investigated the SIRT1 A>G SNPs, rs1467568 and rs7895833 in young SA Indians with CAD and compared them to Indian and black controls.

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Methods A total of 287 subjects were recruited into the SIRT1 rs1467568 study (104 CAD patients, 99 age-, gender- and race-matched controls, and 84 age- and gender-matched black controls) following institutional ethical approval (BE067/14). The inclusion criteria for CAD patients were: Indian ancestry and unrelated adult males aged < 45 years, and stable CAD confirmed on angiography. The exclusion criteria for controls included an acute coronary syndrome/revascularisation procedure in the preceding three months, chronic renal or liver disease, malignancy and known inflammatory or infectious disease. Blood samples were obtained following an overnight fast. A full pathology report of clinical markers was assessed by routine laboratory testing at the Global Clinical and Viral Laboratory (Durban, South Africa), a South African national accreditation system (SANAS) certified laboratory. The following parameters were tested: haematology (Roche Sysmex 1800XT), chemistry (Beckman Coulter DXC600), endocrinology and high-sensitivity C-reactive protein (hsCRP) (Siemens Centaur XP) and serology (BD Biosciences FACS Calibur), as per international standards to obtain levels of total cholesterol, HDL-C, LDL-C, triglycerides, fasting glucose, two-hour glucose, fasting insulin, glycosylated haemoglobin, sodium, potassium, bicarbonate, chloride, urea, creatinine, glomerular filtration rate, CD4, CD8, CD45 and CD3 count. The physical measurements of weight, height, abdominal circumference, waist circumference and patient history were conducted by the cardiologist (Dr S Khan). Genomic DNA was extracted from the whole blood sample of each patient and control, according to the method described by Sambrook et al.11 Cells were transferred to 600-μl lysis buffer [0.5 % sodium dodecyl sulphate (SDS), 150 mM NaCl, 10 mM ethylenediaminetetra-acetic acid (EDTA), 10 mM Tris–HCl (pH 8.0)]. To this, RNase A (100 μg/ml; DNase-free) was added to the solution and incubated at 37°C for one hour. Proteinase K (200 μg/ml) was then added and incubated for three hours at 50°C. Protein contaminants were then precipitated by adding 5 mM 0.1% potassium acetate before centrifugation at 5 000 × g for 15 min. Supernatants containing genomic DNA were transferred to fresh tubes and extracted with 100% isopropanol on ice, and thereafter washed with 70% ethanol. DNA samples were dissolved in 10 mM Tris and 0.1 mM EDTA (pH 7.4, 4°C). DNA concentration was determined using the Nanodrop 2000 spectrophotometer, and all samples were standardised to a concentration of 10 ng/μl. Following the manufacturer’s protocol, TaqMan® SNP predesigned genotyping assay (Life Technologies, Cat #4351379) was used to genotype all subjects for both SNPs. The TaqMan® genotyping assay contains two primers for amplifying the sequence of interest and two TaqMan® minor groove-binding (MGB) probes for detecting alleles. The presence of two probe pairs in each reaction allows genotyping of the two possible variant alleles at the SNP site in a DNA target sequence. The genotyping assay determines the presence or absence of a SNP based on the change in fluorescence of the dyes associated with the probes. The TaqMan® MGB probes consist of targetspecific oligonucleotides with a reporter dye at the 5′ end of each probe: one VIC®-labelled probe to detect allele 1 sequence (A-allele in the case of rs1467568 and rs7895833) and one FAM™-labelled probe to detect allele 2 sequence (G-allele in the case of rs1467568 and rs7895833). A fluorescence signal for both dyes indicates heterozygous for allele 1–allele 2 (AG).

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A final reaction mixture consisted of 40 × TaqMan® predesigned genotyping assay, 2 × TaqMan® genotyping master mix, nuclease-free water, and a 10-ng genomic DNA template. The experiment was done using the Applied Biosystems® ViiA™ 7 Real-Time PCR system.

Statistical analysis The Hardy–Weinberg equilibrium was used to test for deviation of allele/genotype frequency. All other statistical analyses were performed with Graphpad prism software (version 5.0). Allele and genotype frequencies were calculated using the Fisher’s exact and chi-squared tests, respectively. The comparison of biochemical measures between the wild type and variant genotypes was done with a non-parametric t-test. Results are expressed as mean ± standard error. A p-value less than 0.05 was considered statistically significant.

Results SIRT1 rs1467568

The genotype distribution complied with the Hardy–Weinberg equilibrium in the CAD patients and Indian controls (chi-squared p = 0.233 and p = 0.941, respectively), but not in the black control group (chi-squared p < 0.05). No significant difference was observed in the distribution of the SIRT1 rs1467568 alleles between the CAD patients and Indian controls (41 vs 40% respectively, p = 0.9196, OR = 1.040, 95% CI: 0.6998–1.545). The Indian controls presented with a higher frequency of the variant allele compared to the black controls (40 vs 18.5%, respectively, p < 0.0001, OR = 2.996, 95% CI: 1.850–4.853). The variant allele was found at a higher frequency in the total Indian group compared to the total black population (41 vs 18.5%, respectively, p < 0.0001, OR = 3.057, 95% CI: 1.974–4.733) (Table 1).

SIRT1 rs7895833 The genotype distribution complied with the Hardy–Weinberg equilibrium in the CAD patients, Indian controls and black controls (chi-squared p = 0.970, p = 1.000 and p = 0.164, respectively). No significant difference was observed in the distribution of the SIRT1 rs7895833 alleles between CAD patients and Indian controls (40.5 vs 41%, respectively, p = 0.9188, OR = 0.9629, 95% CI: 0.6457–1.436). The Indian controls presented with a higher frequency of the variant allele compared to the black controls (41 vs 22% respectively, p < 0.0001, OR = 2.513, 95% CI: 1.578– 4.004). The variant allele was found at a higher frequency in the total Indian group compared to the total black population (41 vs 22% respectively, p < 0.0001, OR = 2.466, 95% CI: 1.620–3.755) (Table 1). Phulukdaree and co-workers reported biochemical measures of CAD patients and healthy controls in 2012.12 As expected, in our study, CAD patients presented with more conventional risk factors, such as higher body mass index (BMI), higher total and LDL cholesterol and triglyceride levels, and a higher prevalence of type 2 diabetes mellitus than the control groups. No association between the SIRT1 SNPs and biochemical measures were found in the CAD patients (Table 2), Indian controls (Table 3) and black controls (Table 4).

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Table 1. SIRT1 rs1467568 and rs7895833 genotype and allele frequencies in CAD patients and controls CAD patients (n = 104) n, (%)

SA Indian controls (n = 99) n, (%)

Total SA Indians (n = 203) n, (%)

SA black controls (n = 84) n, (%)

40 (38.46)

36 (36.36)

76 (37)

62 (73.81)

42 (40.38)

46 (46.46)

88 (43)

13 (15.48)

22 (21.15)

17 (17.17)

39 (19)

9 (10.71)

122 (59)

118 (60)

240 (59)

137 (81.5)

86 (41)

80 (40)

166 (41)

31 (18.5)

(n = 100)

(n = 99)

(n = 199)

(n = 82)

36 (36)

34 (34.34)

70 (35)

47 (57.32)

47 (47)

48 (48.48)

95 (48)

34 (41.46)

17 (17)

17 (17.17)

34 (17)

1 (1.22)

119 (59.5)

116 (59)

235 (59)

128 (78)

81 (40.5)

82 (41)

163 (41)

36 (22)

SIRT1 rs1467568 Genotypes

Alleles

SIRT1 rs7895833

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Genotypes

Alleles

Discussion Indian populations throughout the world show early-onset CAD, one to two decades earlier than other ethnic groups.13 South African Indians have the highest mortality rates due to CAD, while black South Africans have a very low prevalence of the disease.10 Increasing evidence has shown that SIRT1 is involved in CAD by regulating a number of key metabolic and physiological processes. SIRT1 serves as an anti-atherosclerotic factor

by mediating endothelial nitric oxide synthase (eNOS) and improving endothelial dysfunction, regulating inflammation, reversing cholesterol transport and reducing the risk of CAD. 14 Several SNPs have been identified in SIRT1, a candidate molecule involved in the epigenetic regulation of CAD. To date, there are only a few human genetic association studies regarding SIRT1 SNPs and CAD. Our study was the first investigation of SIRT1 rs1467568 and rs7895833 in SA Indian CAD patients. We observed that the variant alleles of both SIRT1 SNPs occurred more frequently in SA Indians compared to SA blacks. We did not observe any difference in allele frequencies between CAD patients and control groups. Previous studies have shown that some of the SIRT1 SNPs are associated with BMI and obesity, glucose tolerance and diabetes, blood pressure, cholesterol metabolism and coronary artery calcification, all of which contribute to the CAD phenotype.15-19 We examined the possible association between rs1467568 and rs7895833 in SIRT1 and BMI, and levels of total cholesterol, LDL, HDL, triglycerides, fasting glucose, fasting insulin, HbA1c, hsCRP, or IL-6 in CAD patients and control groups, but did not observe any association. The Rotterdam study investigated SIRT1 variation (assessed by three tagging SIRT1 SNPs: rs7895833, rs1467568 and rs497849) in relation to BMI and risk of obesity in 4 573 participants, including 413 individuals with prevalent and 378 with incident type 2 diabetes mellitus (T2DM).20 In homozygous carriers with prevalent T2DM, the SIRT1 haplotype 1 had 1.9 times (95% CI: 1.1–3.2) increased risk of CVD mortality compared to non-carriers.

Table 2. Characteristics of CAD patients according to the SIRT1 rs1467568 and SIRT1 rs7895833 genotypes SIRT1 rs1467568 genotype

SIRT1 rs7895833 genotype

Wild type (AA)

Variant (AG+GG)

p-value

Wild type (AA)

Variant (AG+GG)

p-value

27.52 ± 0.81

28.57 ± 0.55

28.02 ± 0.80

28.33 ± 0.59

Total cholesterol (mmol/l)

5.73 ± 0.32

5.17 ± 0.20

5.32 ± 0.24

5.46 ± 0.25

LDL (mmol/l)

3.70 ± 0.29

3.27 ± 0.21

3.41 ± 0.23

3.47 ± 0.24

HDL (mmol/l)

0.98 ± 0.04

0.89 ± 0.03

0.91 ± 0.04

0.93 ± 0.04

Triglycerides (mmol/l)

2.41 ± 0.28

2.37 ± 0.18

2.34 ± 0.24

2.38 ± 0.20

Fasting glucose (mmol/l)

6.48 ± 0.50

6.27 ± 0.33

6.18 ± 0.47

6.32 ± 0.34

Fasting insulin (μlU/ml)

16.97 ± 2.21

15.54 ± 1.12

14.17 ± 1.19

16.95 ± 1.61

HBA1c (%)

6.63 ± 0.33

6.61 ± 0.24

6.57 ± 0.34

6.60 ± 0.24

hsCRP (mg/l)

9.83 ± 2.58

6.97 ± 0.98

8.93 ± 2.42

7.78 ± 1.31

IL-6 (pg/ml)

2.80 ± 0.90

2.45 ± 0.59

2.41 ± 0.80

2.73 ± 0.68

BMI (kg/m2)

BMI = body mass index, LDL = low-density lipoprotein, HDL = high-density lipoprotein, HBA1c = glycated haemoglobin, hsCRP = high-sensitivity C-reactive protein, IL-6 = interleukin-6, ns = non-significant.

Table 3. Characteristics of Indian controls according to the SIRT1 rs1467568 and SIRT1 rs7895833 genotype SIRT1 rs1467568 genotype

SIRT1 rs7895833 genotype

Wild type (AA)

Variant (AG+GG)

p-value

Wild type (AA)

Variant (AG+GG)

p-value

25.88 ± 0.93

26.65 ± 0.69

25.14 ± 1.01

26.88 ± 0.66

Total cholesterol (mmol/l)

5.32 ± 0.16

5.54 ± 0.13

5.56 ± 0.19

5.41 ± 0.12

LDL (mmol/l)

3.47 ± 0.13

3.86 ± 0.12

3.88 ± 0.17

3.63 ± 0.11

HDL (mmol/l)

1.04 ± 0.07

0.91 ± 0.03

0.97 ± 0.07

0.95 ± 0.03

Triglycerides (mmol/l)

1.79 ± 0.22

1.92 ± 0.27

1.63 ± 0.17

2.00 ± 0.27

Fasting glucose (mmol/l)

5.59 ± 0.34

5.38 ± 0.16

5.27 ± 0.20

5.56 ± 0.22

Fasting insulin (μlU/ml)

15.91 ± 1.96

16.72 ± 1.66

13.36 ± 1.46

18.03 ± 1.75

HBA1c (%)

5.78 ± 0.21

5.65 ± 0.11

5.85 ± 0.16

5.63 ± 0.13

hsCRP (mg/l)

4.58 ± 0.60

7.95 ± 1.71

6.52 ± 1.41

6.87 ± 1.57

IL-6 (pg/ml)

2.16 ± 0.79

2.86 ± 0.63

2.83 ± 0.87

2.48 ± 0.60

BMI (kg/m2)

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Table 4. Characteristics of black controls according to the SIRT1 rs1467568 and SIRT1 rs7895833 genotype SIRT1 rs1467568 genotype

SIRT1 rs7895833 genotype Variant (AG+GG)

p-value

25.53 ± 0.54

27.13 ± 1.20

25.58 ± 0.63

26.57 ± 0.87

Total cholesterol (mmol/l)

4.12 ± 0.12

4.47 ± 0.23

4.30 ± 0.13

4.14 ± 0.18

LDL (mmol/l)

2.62 ± 0.10

3.02 ± 0.22

2.78 ± 0.12

2.71 ± 0.16

HDL (mmol/l)

1.05 ± 0.045

1.03 ± 0.088

1.08 ± 0.05

0.99 ± 0.06

Triglycerides (mmol/l)

0.99 ± 0.072

0.93 ± 0.15

0.98 ± 0.08

0.96 ± 0.11

Fasting glucose (mmol/l)

4.80 ± 0.084

4.90 ± 0.11

4.87 ± 0.11

4.77 ± 0.08

Fasting insulin (μlU/ml)

7.69 ± 0.67

12.11 ± 3.74

9.21 ± 1.73

8.59 ± 1.10

HBA1c (%)

5.83 ± 0.062

5.79 ± 0.082

5.87 ± 0.07

5.76 ± 0.077

hsCRP (mg/l)

6.64 ± 1.86

5.82 ± 1.23

7.81 ± 2.42

4.83 ± 0.84

Wild type (AA) BMI (kg/m2)

Variant (AG+GG)

p-value

Wild type (AA)

An intended replication study (Erasmus Rucphen family study) was carried out involving 2 347 participants. Both studies observed that the minor alleles of rs7895833 (G allele) and rs1467568 (A allele) were associated with lower BMI and a 13–18% decreased risk of obesity in two independent Dutch populations.17 In another study, the A allele of rs7895833 was associated with increased risk of obesity and hypertension in Japanese men.15 Recent studies investigated the association between SIRT1 SNPs (rs7895833, rs7069102, rs144124002 and rs2273773) and CAD in a Turkish population. While rs7069102, rs2273773 and rs144124002 were significantly associated with increased risk for CAD, they found no association between rs7895833 and CAD.21,22 Shimoyama et al. reported that SIRT1 rs7069102 and rs2273773 were associated with abnormal cholesterol metabolism and coronary artery calcification, respectively, in Japanese haemodialysis (HD) patients. The study also found that the A allele frequency of SIRT1 rs7895833 and G allele frequency of rs7069102 were significantly lower in HD patients compared to controls, suggesting an impact on survival.19 The allele frequencies of rs7895833 and rs1467568 show ethnic variation, and this is a possible reason for differing disease patterns among populations. The frequency of the rs7895833 A allele was relatively low (0.29) in Japanese compared to Dutch, Turkish and Caucasian subjects who had similar allele frequencies (0.80, 0.85 and 0.80, respectively).15,17,23 The A allele of rs1467568 (reported as the protective allele) showed marked difference in frequency between European (0.25) and Japanese (0.84) subjects.23

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Horio Y, Hayashi T, Kuno A, Kunomoto R. Cellular and molecular effects of sirtuins in health and disease. Clin Sci 2011; 121(5): 191–203.

Houtkooper RH, Pirinen E, Auwerx J. Sirtuins as regulators of metabolism and healthspan. Nat Rev Mol Cell Biol 2012; 13(4): 225–238.

10. Norman RBD, Pieterse SMD, Groenwald P. Revised burden of disease estimates for the comparative risk factor assessment, South Africa 2000. Cape Town: Medical Research Council, 2006. 11. Sambrook J, Russell DW. Rapid Isolation of Mammalian DNA in Molecular Cloning: A Laboratory Manual. 3rd edn. New York: Cold Spring Harbor Laboratory Press, 2001. 12. Phulukdaree A, Khan S, Moodley D, Chuturgoon AA. GST polymorphisms and early-onset coronary artery disease in young South African Indians. S Afr Med J 2012; 102(7): 627–630. 13. Sharma MGM. Premature coronary artery disease in Indians and its associated risk factors. Vasc Health Risk Mgmt 2005; 1(3): 217–225. 14. Ma L, Li Y. SIRT1: Role in cardiovascular biology. Clin Chim Acta

Conclusion Both SNP variant alleles occurred more frequently in SA Indians than in SA blacks, but no difference was found between CAD patients and controls. This study is limited by sample size and a larger study may be required to fully assess the functional significance of these polymorphisms.

2015; 440: 8–15. 15. Shimoyama Y, Suzuki K, Hamazima N, Niwa T. Sirtuin 1 gene polymorphisms are associated with body fat and blood pressure in Japanese. Translat Res 2011; 157(6): 339–347. 16. Peeters A, Beeckers S, Verrijken A, Mertens I, Roevens P, Peeters PJ, et al. Association of SIRT1 gene variation with visceral obesity. Human Genet 2008; 124(4): 431–436. 17. Zillikens MC, van Meurs JB, Rivadeneira F, Amin N, Hofman A,

P Ramkaran thanks the National Research Foundation (NRF) for a scholarship and UKZN (College of Health Sciences) for funding this study.

Oostra BA, et al. SIRT1 Genetic variation is related to BMI and risk of obesity. Diabetes 2009; 58(12): 2828–2834. 18. Botden IPG, Zillikens MC, de Rooij SR, Langendonk JG, Danser AHJ,

References

Sibrands EJG, et al. Variants in the SIRT1 gene may affect diabetes risk

35(2): 424–426.

Preyat N, Leo O. Sirtuin deacylases: a molecular link between metabolism and immunity. J Leukocyte Biol 2013; 93(5): 669–680.

in interaction with prenatal exposure to famine. Diabetes Care 2012; 19. Shimoyama Y, Mitsuda Y, Tsuruta Y, Suzuki K, Hamzima N, Niwa

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T. SIRTUIN 1 gene polymorphisms are associated with cholesterol metabolism and coronary artery calcification in Japanese hemodialysis patients. J Renal Nutr 2012; 22(1): 114–119. 20. Zillikens MC, van Meurs JB, Sijbrands EJ, Rivadeneira F, Dehghan A, van Leeuwen JP, et al. SIRT1 genetic variation and mortality in type 2 diabetes: interaction with smoking and dietary niacin. Free Rad Biol Med 2009; 46(6): 836–841. 21. İzmirli M, Goktekin O, Bacaksiz A, Uysal O, Kilic U. The effect of the SIRT1 2827 A>G polymorphism, resveratrol, exercise, age and occupa-

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tion in Turkish population with cardiovascular disease. Anatolian J Cardiol 2014; 14. 22. Kilic U, Gok O, Bacaksiz A, Izmirli M, Elibol-Can B, Uysal O. SIRT1 Gene polymorphisms affect the protein expression in cardiovascular diseases. PLoS One 2014; 9(2): e90428. 23. Maeda S, Imamura M, Kurashige M, Araki S, Suzuki D, Babazono T, et al. Association between single nucleotide polymorphisms within genes encoding sirtuin families and diabetic nephropathy in Japanese subjects with type 2 diabetes. Clin Exp Nephrol 2011; 15(3): 381–390.

Should the findings of the TASTE and TOTAL trials change clinical practice? The TOTAL and TASTE trials were undertaken to evaluate whether mechanical thrombus aspiration should routinely accompany primary percutaneous coronary intervention (PCI) for STEMI. Evaluating the evidence prior to the two trials, Dr David Kettles from East London, South Africa, observed that it’s been known for a long time that thrombus is the enemy of good cath lab outcomes. ‘In STEMI, epicardial flow does not equal reperfusion. Distal embolisation is often a problem and angioplasty and stenting can contribute to this. Fifteen per cent of patients undergoing primary PCI have visible distal emboli, and myocardial perfusion after primary PCI is the strongest predictor of mortality.’ He was speaking at AfricaPCR 2016. Aspiration is one of multiple approaches to deal with distal embolisation of thrombus and atherosclerotic debris. Many studies have suggested that manual thrombus aspiration improves ST-segment resolution and various surrogate markers. ‘It’s pathophysiologically plausible and relatively simple. It makes PCI easier and may have short- and long-term benefits. But what about the risk of complications? While the literature tends to downplay these, there is very possibly a higher stroke risk in real-world settings. It’s possible the trials prior to TASTE and TOTAL underestimated the risks. So we needed these randomised, controlled trials powered for mortality.’ Reviewing the two trials, Dr Hellmuth Weich, from Cape Town, South Africa, observed that TASTE was a multicentre, randomised, controlled trial evaluating all-cause mortality at 30 days in patients undergoing primary or rescue PCI for STEMI, either with or without thrombus aspiration. Patients were randomised after angioplasty. There was virtually no difference in outcomes between the two arms at 30 days (2.8 vs 3.0%), with comparable findings at one year. ‘While it was a good trial, it may have been underpowered. There was very low mortality overall and there might possibly have been a selection bias, given that patients were randomised post angioplasty.’ TOTAL was a bigger trial than TASTE, and subjects were randomised prior to angioplasty. Its primary endpoint was

a composite of cardiovascular death, recurrent myocardial infarction, class IV heart failure and cardiogenic shock at six months. Once again, there was no significant difference in outcomes between the two arms of the trial. Of concern, however, was an increase in stroke in the aspirated arm that continued up to six months after the procedure. Dr Weich noted in conclusion that there is therefore no evidence to suggest that manual thrombus aspiration be undertaken routinely. While the trials were interesting, it should also be kept in mind that the patient populations were not the same as those in Africa. In the discussions that followed, the feeling was that the trials don’t tell interventional cardiologists not to undertake aspiration, just not to do it routinely. Patient selection is therefore an important concern, as is technique, in order to prevent stroke. ‘Pay careful attention to the guiding catheter and start aspiration 2 cm proximal to the lesion’, said Dr Kettles. ‘Employ multiple slow-passage techniques – at least two or three passes. Withdraw the aspirate catheter under aspiration and aspirate the guide thereafter.’ Another important determinant is the size and extent of the thrombus. Summing up the key learnings, Dr William Wijns, chairman of PCR, made the following points: • There is no evidence of benefit of systematic mechanical aspiration during PCI based on primary or secondary efficacy measures. • There’s a possible safety signal in the form of increased stroke rates; this is a hypothesis-generating finding. • Operators may still choose thrombus aspiration in individual cases in order to facilitate procedural technique, bearing in mind that it will have no benefit in respect of endpoints such as mortality. ‘There is an open door for its selective use, for example in cases of significant thrombus burden. Unmet needs remain, and we require better tools to remove thrombus efficaciously while protecting the myocardium. We also need to bear in mind that TOTAL and TASTE are not fully relevant to Africa, where the dominant treatment of STEMI is pharmaco-invasive and most patients are late presenters.’ Source: AfricaPCR 2016

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Assessment of the management of acute myocardial infarction patients and their outcomes at the Nairobi Hospital from January 2007 to June 2009 Redemptar Kimeu, Charles Kariuki

Abstract Introduction: The demographics, clinical characteristics and management of patients presenting at the Nairobi Hospital with acute myocardial infarction have not been documented in the past. There is a paucity of studies on this subject in this region. Methods: A retrospective, hospital-based study was carried out, examining data of patients presenting at Nairobi Hospital with acute myocardial infarction between January 2007 and June 2009. The data collected were patient demographics, coronary artery disease (CAD) risk factors, clinical presentation, GRACE score risk stratification, coronary anatomical findings on angiography, interventions and outcomes during hospitalisation. Results: Sixty-four patients were recruited (mean age 56.7 years). The CAD risk-factor profile included systemic hypertension in 71.9% of patients, age over 55 or 65 years in men and women, respectively in 42.2%, 35.9% of subjects were smokers, low high-density lipoprotein cholesterol levels in 25%, diabetes mellitus in 25%, family history of premature coronary artery disease in 8%, prior acute coronary syndrome in 18.8%, ST-segment elevation myocardial infarction (STEMI) in 60.9% and non-ST-segment elevation myocardial infarction (NSTEMI) in 39.1% of patients. In the STEMI arm, 79.5% of patients underwent thrombolysis, 17.9% had rescue percutaneous coronary intervention (PCI) and 2.6% had no reperfusion therapy. Medical management was carried out in 29% of the patients, 19.1% had a coronary artery bypass graft and 40.4% had PCI. The mean duration of hospitalisation was 6.69 days. The in-hospital mortality rate was 9.4% and mean in-hospital probability of death according to the GRACE risk score was 16.05%. Discharge medication was a β-blocker in 84.5% of patients, an ACE inhibitor or angiotensin receptor blocker in 48.3%, low-dose aspirin in 96.6%, clopidogrel in 96.6% and statins in 93.1%. Conclusion: The risk-factor assessment in our population, albeit small, was in keeping with the traditional risk factors for coronary artery disease. There is, however, room for improvement in reconciling the gap between actual and recommended patient care. Keywords: acute myocardial infarction, CAD risk factors, outcomes of acute myocardial infarction, Nairobi Hospital, Kenya

Nairobi Cardiovascular Clinic, Nairobi, Kenya Redemptar Kimeu, MD, drkimeulekasi@talanasc.co.ke

Nairobi Cardiovascular Clinic, Nairobi, Kenya Charles Kariuki, MD, FACC, FRCP

Submitted 27/8/13, accepted 7/12/15 Cardiovasc J Afr 2016; 27: 218–221

www.cvja.co.za

DOI: 10.5830/CVJA-2015-091

The demographic and clinical characteristics of patients presenting with acute myocardial infarction at the Nairobi Hospital have not been documented in the past. Abnormal lipid levels, smoking, hypertension, diabetes mellitus, abdominal obesity, psychosocial factors, low consumption of fruit and vegetables, alcohol abuse, and no regular physical activity account for most of the risk factors for myocardial infarction worldwide in both genders and all ages in all regions.1 The riskfactor distribution for coronary artery disease (CAD) in our sub-population may be similar to those in the Western world.1,2 Control of these risk factors is key to the prevention of and reduction in the incidence of CAD.3 Over the past 20 years, there has been considerable progress with improved outcomes in the treatment of acute coronary syndromes (ACS). These include the establishment of coronary care units and the development of antiplatelet therapy, refinement of anticoagulation strategies and introduction of fibrinolytic therapies. Percutaneous coronary intervention (PCI) has become the intervention of choice in the acute setting in ST-segment elevation myocardial infarction (STEMI). Early invasive intervention in non-ST-segment elevation myocardial infarction (NSTEMI) is also advocated.4-8 Randomised clinical trials in STEMI patients have shown that efficient triaging and early reperfusion therapy decreases mortality rates. Early thrombolysis is effective in improving outcomes in acute STEMI. Although timely performance of primary PCI is more effective in the restoration of patency, and for lower re-occlusion rates, improved residual left ventricular function and better clinical outcomes, this benefit diminishes with any delays.5,9 The initial strategy in NSTEMI is to alleviate ischaemia and symptoms by using anti-ischaemic agents, antiplatelets, anticoagulants, IIb/IIIa inhibitors, to monitor the patient with serial ECGs, and carry out repeat measurements of markers of myocardial necrosis. The invasive coronary approach has been shown to reduce mortality rates in NSTEMI patients.10 Research evidence has documented reduced mortality rates at 30 days with the use of β-blockers, ACE inhibitors, antiplatelet therapy and statins, smoking cessation, and timely reperfusion therapy in acute myocardial infarction.11,12 The GRACE risk model has been validated to establish the in-hospital mortality risk in patients with STEMI and NSTEMI. In this model, the risk factors predicting early mortality include age over 70 years, prior myocardial infarction, Killip class at

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admission, anterior myocardial infarction, and the combination of hypotension and tachycardia. The GRACE risk model assists in risk profiling and hence prioritising care.13,14 The Nairobi Hospital has a well-equipped emergency department and intensive care unit (ICU), and a modern cardiac catheterisation laboratory with well-trained staff. At the Nairobi Hospital, fibrinolytic therapy is the treatment of choice for STEMI. Primary PCI is increasingly used however it is not yet available timeously and consistently at all hours, due to various logistical factors. This was a retrospective study of the Nairobi Hospital ICU and high-dependence unit (HDU) in-patient records to describe the demographics, risk factors, clinical characteristics, management and outcomes of patients diagnosed with acute myocardial infarction admitted to the Nairobi Hospital ICU and HDU from January 2007 to June 2009.

Methods The hospital ethics committee gave consent for the study.The in-patient patients’ records were retrieved and a cardiologist verified the diagnosis of STEMI and NSTEMI, after which the pre-specified data variables were retrieved and filled into a pre-designed study pro forma. Patients who presented with a diagnosis of acute myocardial infarction and were over 18 years were included. STEMI was defined according to the European Society of Cardiology guidelines, as patients with acute chest pain and persistent (> 20 minutes) ST-segment elevation at the J-point in two contiguous leads with the cut-off points: ≥ 0.2 mV in men or ≥ 0.15 mV in women in leads V2–V3 and/or ≥ 0.1 mV in other leads. For the purposes of reperfusion strategy, true posterior myocardial infarction was considered in patients with ST-depression in the anterior leads or new prominent R waves on the same leads.15,16 NSTEMI was defined as patients with chest pain but without persistent ST elevation, new horizontal or down-sloping ST depression ≥ 0.05 mV in two contiguous leads; and/or T inversion ≥ 0.1 mV, flat T waves and pseudo-normalisation of T waves in two contiguous leads with prominent R wave or R/S ratio or no ECG changes at presentation. The confirmation of acute myocardial infarction was made based on elevated cardiac biomarkers.15,16 Significant atheromatous lesions were defined as a left main stem lesion of > 50% stenosis and lesions involving the left anterior descending, left circumflex and right coronary arteries > 70% stenosis. We analysed the demographic data, mode of transport to the accident and emergency department, first medical contact, time of arrival after onset of chest pain, door-to-first ECG time, and ECG-to-cardiologist time. The GRACE risk score of each patient on admission was documented as per the parameters at admission. The risk factors for coronary artery disease analysed included age > 55 years in men and > 65 years in women, male gender, current smoking, low high-density lipoprotein cholesterol (HDLC) level < 40 mg/dl (1.04 mmol/l), systemic hypertension (blood pressure > 140/90 or on antihypertensives), diabetes and family history of premature coronary artery disease. The initial medication given at the accident and emergency department, the type of myocardial infarction and the reperfusion treatment for STEMI were noted. The door-to-

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needle time and fibrinolytic agent used were both recorded. The coronary anatomy and TIMI grade for coronary blood flow were documented as per the cardiologist’s notes and confirmed by an interventional cardiologist who studied the digital images of the coronary angiogram. The in-hospital complications, duration of hospitalisation, in-hospital deaths and discharge medications were noted.

Statistical analysis The extracted data were entered into the Statistical Package for Social ScienceTM (SPSS) version 13.0 for Windows (SPSS, Chicago, IL, USA) statistical software to check for errors and perform the requisite statistical tests. Data were analysed using the same software. Descriptive analysis was performed to characterise the number and type of patient outcomes. To obtain insight into the social demographic factors of the patients, frequency tables were used with accompanying percentages. Bivariate comparisons of continuous symmetric characteristics, such as duration of time between onset of chest pain/ symptoms and arrival at hospital, door-to-first ECG time and ECG-to-cardiologist time were performed using the Student’s t-test and Mann–Whitney test for non-symmetric characteristics for patients with independent variables (in-hospital outcome). Fisher’s exact test and the chi-squared test, as appropriate, were used for comparison of categorical characteristics, such as gender, coronary artery anatomy and in-hospital complications with patients’ in-hospital outcomes. Correlations between variables were tested using the Pearson’s correlation co-efficient. Prevalence of risk factors was calculated with accompanying 95% confidence intervals. Statistical significance was defined as a two tailed p-value ≤ 0.05.

Results Sixty-four patients fulfilled the criteria for STEMI and NSTEMI from January 2007 to June 2009 and 87.5% were male. The mean age was 56.7 years. Of the study population, 60.9% were from the community, while 26.6% were referrals from other health facilities; 28.1% were brought by ambulance but mode of transport was not documented in 67.2% of the cases. Five per cent of the patients were already hospitalised and 89.1% had their first medical contact in hospital. Among the patients, 17.2% arrived within one hour of onset of chest pain, whereas 40.6% arrived at the emergency department more than 12 hours after the onset of chest pain. The presence of coronary artery risk factors in this population was as follows: systemic hypertension was found in 71.9% of patients, 42.2% were over 55 or 65 years in men and women, respectively, 35.9% were cigarette smokers, 25% had a low HDL-C level, 25% had diabetes mellitus, 8% had a documented family history of premature coronary artery disease, 18.8% had a previous history of acute coronary syndrome and 9.4% had chronic kidney disease. The documented door-to-ECG time was less than 10 minutes in only 10.9% of patients and the ECG-to-cardiologist time was less than 30 minutes in 36.5%. Both aspirin and clopidogrel were received by 96.9% of patients on arrival at the emergency department. A loading dose of aspirin was given in 53.2% of patients, whereas 62.9% received a loading dose of clopidogrel;

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89.1% received enoxaparin and only 9.4% received unfractionated heparin; 67.2% had transthoracic echocardiography during their hospital stay, of whom 76.9% had a left ventricular ejection fraction (LVEF) > 40%. For type of acute myocardial infarction, 60.9% had a diagnosis consistent with STEMI and 39.1% had NSTEMI. In the NSTEMI arm, 44% of the patients received IIb/IIIa inhibitors, mainly eptifibatide infusion, and 68% had a coronary angiogram done before hospital discharge. The coronary anatomy was consistent with significant atheromatous lesions in 79% of the patients. Of those who had no coronary angiography, 60% had non-invasive testing for myocardial ischaemia before discharge. In the STEMI arm, 79.5% of patients received thrombolysis, 17.9% underwent rescue PCI and 2.6% did not receive reperfusion therapy. None of the patients underwent primary PCI. The door-to-needle time was less than 120 minutes in 45.2% of the thrombolysis patients and 38.7% had no documentation of door-to-needle time. The thrombolytic agent was tenecteplace in 80.6% and steptokinase in 6.5% of patients. Only 51.6% of the thrombolysis patients had an ECG done at 90 minutes post thrombolysis, of whom 56.3% had achieved reperfusion. Of the patients who did not achieve reperfusion, 66.7% underwent rescue PCI. The coronary anatomy was consistent with significant atheromatous lesions in 82.1% of the patients who underwent angiographic studies. Of the patients who underwent coronary angiography, 29% were managed medically, 19.1% were referred for coronary artery bypass grafting (CABG), and 40.4% had PCI with 84.2% receiving drug-eluting stents. Of the patients who underwent PCI, 87.5% achieved TIMI flow of grade 2 to 3. Cardiogenic shock occurred in 17.2% of patients, new atrial fibrillation in 6.3% and cardiac arrest in 3.1%. Sustained ventricular tachycardia or ventricular fibrillation occurred in 5.3%, atrioventricular block in 4.7% and acute kidney injury (creatinine > 200 μmol/l) in 6.3%. The mean duration of hospitalisation was 6.69 days. In-hospital mortality rate was 9.4%. The mean in-hospital probability of death according to the GRACE risk score was 16.05%. Upon discharge from hospital, 84.5% were prescribed β-blockers, 48.3% were on ACE inhibitors or angiotensin receptor blockers (ARBs), 96.6% were on low-dose aspirin, 96.6% on clopidogrel and 93.1% on statins.

Discussion Cardiovascular risk factors have been determined from several landmark studies, mostly performed in the Western world. The African forum has recently embarked on some local studies. The patient population in our study was similar in age and gender to that of other African studies on traditional cardiovascular risk factors in patients with confirmed coronary artery disease.2,17 The most prevalent risk factor in our study was systemic hypertension, which reflects the findings in previous studies. In the overall INTERHEART study, 39% of the patient population had systemic hypertension, whereas in the INTERHEART Africa arm, it was reported in 42.3% of cases.1,2,17,18 Emergency medical systems were not in place in our local setting at the time we carried out this study, therefore first

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medical contact was at the emergency department. Most patients arrived after three hours from onset of chest pain. The initial triage at the emergency department was not optimal with regard to door-to-ECG, ECG-to-cardiologist and door-to-thrombolysis times going beyond the recommended door-to-needle time of 30 minutes. The hospital is currently looking at a system that will allow for immediate interactive communication with the concerned cardiologist to advise on the management of these patients. The thrombolytic agent used most frequently was tenecteplace. Almost all patients received antiplatelet and antithrombin co-therapy as per the current guidelines. The gold standard of acute management of ST-elevation myocardial infarction is primary PCI, with thrombolytic therapy being effective when given early. Thrombolysis was the standard approach in this study. Most patients in our study had an echocardiogram performed during their hospital course. It is generally indicated that echo assessment of cardiac anatomy and function be done in the first 24 to 48 hours after acute myocardial infarction.4 Our data showed that the in-hospital mortality rate of patients with a diagnosis of acute myocardial infarction was 9.4%. Despite the small size of the study population, this is comparable to data derived from European studies. The mean probability of in-hospital death in our study according to the GRACE risk score was 16.05%. Secondary preventive medication was prescribed for most patients as per the standard recommendations. However ACE inhibitors or ARBs were prescribed in less than 50% of patients.

Conclusion The risk-factor assessment in our population of patients, albeit small, was in keeping with traditional risk factors for coronary artery disease found in other studies. The risk for acute myocardial infarction was found to increase with higher income and educational level in our black African population, in contrast with findings in other African groups. With advances in the field of cardiology, the local emergency medical system will improve and timely invasive management of patients presenting with acute myocardial infarction will be available. Currently, there is room for improvement in reconciling the gap between actual and recommended patient care. There is also a need to develop local management protocols for patients with acute myocardial infarction, based on local specialist experience and the available facilities. The Nairobi Hospital is committed to putting in place facilities to allow for primary PCI and early thrombolysis.

References 1.

Yusuf S, Hawken S. Effect of potentially modifiable risk factors associated with myocardial infarction in 52 countries (the INTERHEART study): case-control study. Lancet 2004; 364: 937–952.

Kamotho C, Ogola E, Joshi E. Cardiovascular risk factor profile of black Africans undergoing coronary angiography. East Afr Med 2004; 81: 82–86.

Furberg C, Wright J, Davis B, ALLHAT Collaborative Research Group. Major outcomes in high risk hypertensive patients randomised to angiotensin-converting enzyme inhibitors or calcium channel blocker versus diuretic. The Antihypertensive and Lipid Lowering Treatment

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elevated acute coronary syndromes. Eur Heart J 2007; 31: 1462–1536.

to Prevent Heart Attack Trial (ALLHAT). J Am Med Assoc 2002; 288: 2981–2997. 4.

11. Cannon C, Braunwald E, McCabe C, Rader DJ, Rouleau JL, Belder R, et al. Intensive versus moderate lipid lowering with statins after acute

Antman E, Hand M, Armstrong W, Armstrong. P, Bates E, Green

coronary syndromes. N Eng J Med 2004; 350: 1495–1504.

L, et al. 2007 focused update of the ACC/AHA 2004 guidelines for the management of patients with ST-elevation myocardial infarction:

12. Kim A, Michael J, Omar H. A validated prediction model for all forms

a report of the American College of Cardiology/American Heart

of acute coronary syndrome: estimating the risk of 6-month post-

Association Task Force on practice guidelines. Circulation 2008; 117:

discharge death in an international registry. J Am Med Assoc 2004; 291: 2727–2733.

296–329. 5.

Brahmajee K, Elizabeth H, Harlan K. Time to treatment in primary

13. Granger B, Goldberg J, Dabbous O, Pieper KS, Eagle KA, Cannon CP,

percutaneous coronary intervention. N Engl J Med 2007; 357: 1631–

et al. Predictors of hospital mortality in the Global Registry of Acute Coronary Events. Arch Inter Med 2003; 163: 2345.

1638. 6.

Goodman S, Cohen M, Bigonzi F, Gurfinkel E, Radley D, Le Iouer V,

14. Babaev A, Frederick P, Pasta D, Every N, Sichrovsky T, Hochman JS.

et al. A comparison of low molecular weight heparin with unfraction-

Trends in management and outcomes of patients with acute myocardial

ated heparin for unstable coronary artery disease. N Eng J Med 1997;

infarction complicated by cardiogenic shock. J Am Med Assoc 2005; 294: 448–454.

337: 447–452. 7.

Ferguson J, Calif R, Antman E, SYNERGY Trial Investigators.

15. O’Connor G, Quinton HB, Traven ND, Ramunno LD, Dodds

Enoxaparin versus unfractionated heparin in high risk patients with

TA, Marciniak TA, Wennberg JE. Geographic variation in the treatment

non-ST-segment elevation acute coronary syndromes managed with

of acute myocardial infarction: the Cooperative Cardiovascular Project. J Am Med Assoc 1999; 281: 627–633.

an intended early invasive strategy: Primary results of the SYNERGY randomized trial. J Am Med Assoc 2004; 292: 45–54. 8.

16. Bassand P, Hamm CW, Ardissino D, Boersma E, Budaj A, Fernández-

Sabatine M, Cannon C, Gibson M, López-Sendón JL, Montalescot

Avilés F, et al. Guidelines for the diagnosis and treatment of non

G, Theroux P, et al. Addition of clopidogrel to aspirin and fibrinolytic

ST-segment elevated acute coronary syndromes. Eur Heart J 2007; 28: 15.

therapy for myocardial infarction with ST-segment elevation. N Eng J Med 2005; 352: 1179–1189. 9.

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17. Thygesen K, Alpert J, White H, on behalf of the Joint ESC/ACCF/

Werf F, Bax J, Betriu A, Blomstrom-Lundqvist C, Crea F, Falk V, et

AHA/WHF task force for the redefinition of myocardial infarction.

al. Management of acute myocardial infarction in patients presenting

Universal definition of myocardial infarction. Eur Heart J 2007; 28:

with persistent ST-segment elevation. Eur Heart J 2005; 29: 2909–2945.

2525–2538.

10. Bassand P, Hamm C, Ardissino D, Task Force for the management

18. Steyn K, Sliwa K, Hawken S, Commerford P, Onen C; INTERHEART

of acute coronary syndromes (ACS) in patients presenting without

Investigators in Africa. Risk factors associated with myocardial infarc-

persistent ST-segment elevation of the European Society of Cardiology

tion in Africa the INTERHEART Africa study. Circulation 2005; 112:

(ESC). Guidelines for the diagnosis and treatment of non ST-segment

3554–3561.

During prolonged low-intensity exercise, caffeine alters blood glucose levels The effects of caffeine versus maltodextrin during exercise were observed in patients with type 2 diabetes. Researchers examined the effects on blood pressure (BP), heart rate (HR) and blood glucose (BG) levels associated with the intake of caffeine in comparison to maltodextrin (CHO) during prolonged periods of low-intensity exercise in patients with type 2 diabetes. Researchers conducted a pilot study on eight individuals with type 2 diabetes who were aged 55 ± 10 years. The participants either received 1 g/kg of CHO or 1.5 mg/kg of caffeine before undergoing exercise. They then exercised for 40 minutes, executed at 40% HR reserve, and recovered for 10 minutes. Their BP and exertion, assessed by the Borg scale, were checked every two minute, and their BG levels were checked every 10 minutes. The ANOVA test was used for statistical analysis, and a p-value < 0.05 indicated statistically significant results.

Neither of the treatments produced significant changes in BP and HR. However, 1.5 mg/kg caffeine significantly reduced BG levels by 75 mg/dl (65% CI; p < 0.05) as opposed to 1 g/kg maltodextrin, which produced no significant change in BG levels during the 40-minute period of exercise.

References 1.

Da Silva LA, De Freitas L, Medeiros TE, Osiecki R, Garcia Michel R, Snak AL, et al. Caffeine modifies blood glucose availability during prolonged low-intensity exercise in individuals with type-2 diabetes. Colomb Med (Cali); 2014; 45 (2):72-76. eCollection 2014 Apr. PubMed PMID: 25100892.

http://www.diabetesincontrol.com/index.php?option=com_content &view=article&id=16750&catid=1&Itemid=17.

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Acquired von Willebrand syndrome in children with aortic and pulmonary stenosis Fatih Köksal Binnetoğlu, Kadir Babaoğlu, Şayegan Güven Filiz, Emine Zengin, Gürkan Altun, Suar Çakı Kılıç, Nazan Sarper

Abstract Introduction: This prospective study was planned to investigate the frequency and relationship of acquired von Willebrand syndrome (AVWS) with aortic and pulmonary stenosis in patients. Methods: A total of 84 children, ranging from two to 18 years of age, were enrolled in this study. Of these, 28 had isolated aortic stenosis, 32 had isolated pulmonary stenosis and 24 were healthy. Children with aortic and pulmonary stenosis associated with other congenital heart diseases were excluded. Children with hypothyroidism, renal or liver disease, malignancy or autoimmune disease were also excluded. Wholeblood count, blood group, factor VIII level, prothrombin time (PT), activated partial thromboplastin time (aPTT), von Willebrand factor antigen (VWF:Ag), ristocetin co-factor (VWF:RCo), and bleeding time using a platelet-function analyser (PFA-100) were performed in all patients. All of the children in the study underwent a detailed physical examination and echocardiographic evaluation. Results: A history of bleeding was positive in 18% of the aortic stenosis group, 9% of the pulmonary stenosis group, and 4% of the control group. Seven of 60 (12%) patients had laboratory findings that implied a diagnosis of AVWS, and two of these (28%) had a history of bleeding. The frequency of AVWS was 14% in patients with aortic stenosis and 9% in those with pulmonary stenosis.

Department of Paediatric Cardiology, Çanakkale Onsekiz Mart University, Çanakkale, Turkey Fatih Köksal Binnetoğlu, MD, koksaldr@yahoo.com

Department of Paediatric Cardiology, Kocaeli University Medical Faculty, Kocaeli, Turkey Kadir Babaoğlu, MD

Department of Paediatrics, Karabük State Hospital, Karabük, Turkey Şayegan Güven Filiz, MD

Department of Paediatric Haematology, Kocaeli University Medical Faculty, Kocaeli, Turkey Emine Zengin, MD Nazan Sarper, MD

Department of Paediatric Cardiology, Kocaeli Derince Education and Research Hospital, Kocaeli Turkey

Conclusion: AVWS is not rare in stenotic obstructive cardiac diseases. A detailed history of bleeding should be taken from patients with valvular disease. Even if the history is negative, whole blood count, PT and aPTT should be performed. If necessary, PFA-100 closure time and further tests should be planned for the diagnosis of AVWS. Keywords: aortic stenosis, children, pulmonary stenosis, von Willebrand Submitted 21/3/15, accepted 7/12/15 Cardiovasc J Afr 2016; 27: 222–227

www.cvja.co.za

DOI: 10.5830/CVJA-2015-093

Acquired von Willebrand syndrome (AVWS) is a rare clinical condition characterised by prolonged bleeding time and decreased levels of factor VIII and von Willebrand factor, as in congenital von Willebrand disease (VWD). It was first defined by Simone et al. in a seven-year-old boy with systemic lupus erythematosus.1 The actual prevalence of AVWS is uncertain. From 1968 to 1999, a total of 266 cases were reported. The International Society of Thrombosis and Haemostasis reported a retrospective analysis of 186 cases in 2000.2 AVWS is usually associated with underlying diseases such as lymphoproliferative disorders (48%), cardiovascular diseases (21%), myeloproliferative disorders (15%), other neoplasms (5%) and autoimmune diseases (2%).2 In rare cases, AVWS is also associated with hypothyroidism, uraemia and certain drugs, such as valproic acid and ciprofloxacin. A few studies have reported on the association between AVWS and aortic stenosis in adults, but there are not enough studies reporting on the relationship between AVWS and right ventricular outflow obstruction. In patients with aortic stenosis, there is a loss of high-molecular-weight multimers, which are active in coagulation, due to increased shear stress. This situation can cause bleeding problems after menstruation, circumcision, dental extraction and congenital heart disease surgery. No other coagulation tests are routinely used in people with heart disease, because AVWS has been identified in only a small number of patients. Therefore, it is of importance to identify AVWS in children with congenital heart diseases.1,3-5 In this study, we aimed to evaluate AVWS in children with aortic and pulmonary stenosis, and their clinical consequences.

Gürkan Altun, MD

Department of Paediatric Haematology, Cumhuriyet University Medical Faculty, Sivas, Turkey Suar Çakı Kılıç, MD

Methods A total of 84 children, ranging from two to 18 years of age, were enrolled in this study. Of these, 28 had isolated aortic

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stenosis, 32 had isolated pulmonary stenosis and 24 were healthy. Children with aortic and pulmonary stenosis associated with other congenital heart diseases were excluded. Children with hypothyroidism, renal or liver disease, malignancy or autoimmune disease were also excluded. A standardised screening questionnaire was used to evaluate each patient’s bleeding symptoms. Written informed consent was obtained from the parents of each patient, and the local ethics committee approved the study. Drugs and foods that could affect the coagulation tests were stopped one week before the evaluation. All the children in the study underwent a detailed physical examination and echocardiographic evaluation. Transthoracic echocardiography was performed with a Vivid 7 (GE Vingmed, Horten, Norway) echocardiograph. M-modes of two-dimensional images were obtained from the parasternal long-axis views. Interventricular septal wall thickness, left ventricular posterior wall thickness and left ventricular internal diameters were measured in all the children. Cardiac chamber sizes and left ventricular systolic and diastolic function were assessed in accordance with the guidelines of the American Society of Echocardiography.6 The mean and peak transvalvular pressure gradients were calculated using the modified Bernoulli equation. Patients with aortic stenosis were classified according to peak pressure gradient as insignificant (< 25 mmHg), mild (25–50 mmHg), moderate (50–74 mmHg) or severe (> 75 mmHg).7 Patients with pulmonary stenosis were classified according to peak gradient as mild (< 36 mmHg), moderate (36–64 mmHg) and severe (> 64 mmHg).8,9 The following blood collection and laboratory assays were performed: whole blood count, blood group, factor VIII level, prothrombin time (PT), activated partial thromboplastin time (aPTT), von Willebrand factor antigen (VWF:Ag), ristocetin co-factor (VWF:RCo), bleeding time using a platelet-function analyser (PFA-100; Dade Behring, Marburg, Germany) and platelet aggregation using a lumi-aggregometer. The PFA-100 is a high-shear system for in vitro testing of platelet function that simulates primary haemostasis after injury to a small vessel by determining the closure time of adenosine Table 1. Abbreviations and interpretations in the study Test name

Abbreviation

Interpretation

Von Willebrand factor antigen

VWF:Ag

Measurement of the quantity of VWF monomers but no information given about its functional ability

Ristocetin co-factor assay

VWF:Rco

Measurement of the ability of VWF to agglutinate formalin-fixed platelets in presence of ristocetin

Ristocetin-to-VWF antigen ratio

VWF:Rco/ VWF:Ag

Parameter of the capacity of available VWF to bind platelets

Collagen-binding capacity

VWF:CB

Measurement of the ability of high molecular weight VWF multimers to bind to sub-endothelial collagen

Collagen-binding capacity-to-VWF antigen ratio

VWF:CB/ VWF:Ag

Measurement of the biological capacity of available VWF for binding to collagen.

Platelet-functional analyser (PFA-100) closure time (collagen and epinephrine) or (collagen and ADP)

PFA-100 CEPI Screening test for primary haemostaor PFA-100 sis. Evaluates platelet disorders and CADP functions.

Ristocetin-induced platelet aggregation

RIPA

Measurement of the ability of various agonists to platelets to aggravate in vitro activation and platelet-to-platelet activation

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diphosphate (ADP) cartridges. It is a highly sensitive way to screen patients for von Willebrand factor defect.10,11 Plasma VWF:Ag levels were evaluated by immunoturbidometry. VWF:Ag and VWF:RCo levels were standardised according to the blood group of each patient. Platelet aggregation tests were studied in 14 patients in whom the PFA-100 ADP collagen closure time was prolonged by low levels of VWF:Ag and VWF:RCo. A diagnosis of AVWS was established by the following: (1) acquired history of bleeding; (2) low values of VWF:RCo and collagen-binding capacity (VWF:CB); and (3) VWF:RCo/ VWF:Ag and VWF:CB/VWF:Ag ratios less than 0.7 in cases of borderline or normal values of VWF:RCo and VWF:CB, and no curves in the platelet-aggregation test with ristocetin (RIPA).2 The gold standard for the detection of structural abnormalities of VWF is multimer analysis using electrophoretic separation and immunostaining. However this labour-intensive and time-consuming assay is not available in many laboratatories. Interpretation of the von Willebrand profile analysis is summarised in Table 1.

Statistical analysis SPSS software version 13.0 (SPSS Inc, Chicago, IL) was used for analysis. All results were expressed as mean ± SD. Assessment of significance between the groups was evaluated with the chi-squared and Mann–Whitney U-test and one-way ANOVA. Correlations between variables were assessed with Pearson’s rank-correlation test. A p-value < 0.05 was considered significant.

Results The study group consisted of 28 patients with aortic stenosis (23 males, five females), 32 patients with pulmonary stenosis (18 males, 14 females), and 24 healthy children (14 males, 10 females). The mean ages were 8.09 ± 3.73 years in the aortic stenosis group, 5.72 ± 3.79 years in the pulmonary stenosis group, and 9.12 ± 4.70 years in the control group. A history of bleeding was positive in five patients with aortic stenosis, three with pulmonary stenosis and in one healthy child (Table 2). The mean follow-up time in patients with aortic stenosis was 4.19 ± 1.91 years, and 3.28 ± 1.98 years in patients with pulmonary stenosis (Table 3). Echocardiographic findings: the distribution of patients grouped by degree of stenosis is provided in Table 4. Haematological parameters: although the PT was in the normal range in patients with aortic and pulmonary stenosis, their PT values were found to be significantly shorter than that of the control group (p = 0.03 and 0.006, respectively). There was no significant difference between the PTs of the aortic and pulmonary stenosis groups (p = 0.52). The aPTT of the patients Table 2. Episodes of bleeding of study group Bleeding type

Patients (n)

Diagnosis

Bleeding after circumcision

1 AS, 1 PS

Epistaxis

1 healthy, 2 AS

Bleeding after minor trauma

1 AS, 1 PS

Bleeding after dental extraction

Postoperative bleeding

AS: aortic stenosis, PS: pulmonary stenosis.

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Aortic stenosis (n = 28) Gender (M/F)

Pulmonary stenosis (n = 32)

Table 6. The comparison of haematological parameters of patients with aortic stenosis according to degree of stenosis

Control (n = 24)

p-value

23/5

18/14

14/10

0.07

Age (year)

8.09 ± 3.73

5.72 ± 3.79

9.12 ± 4.70

0.007

Follow-up period (year)

4.19 ± 1.91

3.28 ± 1.98

–

0.07

10.71 ± 2.34

< 0.001

Aortic PPG (mmHg)

46.84 ± 17.63* 13.24 ± 2.78

Aortic MPG (mmHg)

24.43 ± 10.59

Pulmonary PPG (mmHg)

12.7 ± 1.49

Pulmonary MPG (mmHg)

–

0.004

47.15 ± 11.70** 11.26 ± 2.18 24.95 ± 7.65

< 0.001

–

0.003

M: male, F: female, MPG: mean pressure gradient, PPG: peak pressure gradient. *p < 0.001 aortic stenosis versus pulmonary stenosis and control. **p < 0.001 pulmonary stenosis versus aortic stenosis and control. Table 4. The distribution of patients according to degree of stenosis Group

Mild

Aortic stenosis, n (%) Pulmonary stenosis, n (%) Total, n (%)

19 (67.9)

Moderate

Severe

Total

6 (21.4)

3 (10.7)

28 (46.7)

3 (9.4)

25 (78.1)

4 (12.5)

32 (53.3)

22 (36.7)

31 (51.7)

7 (11.7)

60 (100)

with pulmonary stenosis was significantly higher than that of the control group (p = 0.019) (Table 5). The VWF:Ag and VWF:RCo levels were not statistically different among the patients with aortic stenosis according to degree of stenosis (Table 6). PFA-100 collagen epinephrine closure time was significantly higher in patients with severe aortic stenosis than in those with mild or moderate aortic stenosis (p = 0.003 and p = 0.01, respectively). PFA-100 collagen epinephrine closure times were positively correlated with degree of aortic stenosis (p = 0.03) (Fig. 1). The platelet count of patients with mild pulmonary stenosis was significantly lower than that of patients with severe pulmonary stenosis (p = 0.04). The PT of the patients with severe pulmonary stenosis was significantly higher than that of those with moderate pulmonary stenosis (p = 0.003). The aPTT of the patients with mild pulmonary stenosis was significantly higher than that of those with moderate and severe pulmonary Table 5. Comparison of haematological parameters of the groups Haematological parameters Platelet count (cells/mm³)

Aortic stenosis (n = 28)

Pulmonary stenosis (n = 32)

Control (n = 24)

336.570 ± 58.053 356.440 ± 103.244 346.920 ± 58.670

PT (s)

13.46 ± 0.64

13.34 ± 0.68

aPTT (s)

29.55 ± 2.06

p-value 0.62

13.87 ± 0.75a

0.017

30.49 ± 3.48

28.82 ± 1.50

0.059

Factor VIII (%) 123.57 ± 42.92

118.69 ± 47.71

130.79 ± 37.35

0.58

100.86 ± 31.97

98.44 ± 31.34

97.42 ± 21.49

0.90

94.75 ± 29.36

97.28 ± 34.28

89.00 ± 19.04

0.56

PFA-100 CADP (s)

116.86 ± 45.14

117.34 ± 46.75

–

0.96

PFA-100 CEPI (s)

178.64 ± 56.19

168.22 ± 55.44

–

0.47

0.95 ± 0.21

1.01 ± 0.35

0.91 ± 0.12

0.36

VWF:Ag (%) VWF:RCo (%)

VWF:RCo/ VWF:Ag

PT: prothrombin time; aPTT: activated partial thromboplastin time; PFA-100 CADP: platelet-function analyser-100 adenosine diphosphate collagen closure time; PFA-100 CEPI: platelet-function analyser-100 collagen epinephrine closure time; VWF:Ag: von Willebrand factor antigen; VWF:RCo: ristocetin co-factor. PFA-100 ADP closure time was carried out in only patients with aortic and pulmonary stenosis (n = 60), but the other tests were done in all children (n = 84). a p = 0.03, aortic stenosis versus control group and p = 0.006, pulmonary stenosis versus control group. b p = 0.019, pulmonary stenosis versus control group.

Mild aortic Moderate aortic stenosis (n = 19) stenosis (n = 6) (mean ± SD) (mean ± SD) Platelets (cells/mm³)

Severe aortic stenosis (n = 3) (mean ± SD)

329.680 ± 57.915 352.000 ± 60.332 349.330 ± 69.292

p-value 0.67

PT (s)

13.579 ± 0.642

13.183 ± 0.752

13.267 ± 0.208

0.37

aPTT (s)

30.005 ± 1.658

28.300 ± 3.039

29.167 ± 1.656

0.20

Factor VIII (%)

120.21 ± 39.76

132.67 ± 63.96

126.67 ± 6.02

0.82

VWF:Ag (%)

101.32 ± 35.17

95.17 ± 27.27

109.33 ± 24.50

0.82

95.11 ± 29.37

98.00 ± 29.67

86.00 ± 39.05

0.85

PFA-100 CADP (s)

110.74 ± 39.79

119.00 ± 32.33

151.33 ± 92.13

0.36

PFA-100 CEPI (s)

166.37 ± 52.05

173.50 ± 34.51

266.67 ± 46.14c

0.01

0.95 ± 0.19

1.03 ± 0.27

0.76 ± 0.20

0.21

VWF:RCo (%)

VWF:RCo/ vVWF:Ag

stenosis (p = 0.002 and p = 0.009, respectively) (Table 7). PFA-100 collagen epinephrine and PFA-100 ADP closure times were not different among the patients with pulmonary stenosis (p = 0.93 and p = 0.18). VWF:Ag levels were normal in all patients with aortic stenosis and in the control group. However, it was low in three patients with pulmonary stenosis (two female, one male) and all were asymptomatic. The family histories of these three patients were negative for bleeding. VWF:RCo was lower in one patient who had a deficiency of VWF:Ag. Of the patients with low VWF:Ag, PFA-100 ADP collagen closure time was prolonged in two patients, and PFA-100 collagen epinephrine closure time was prolonged in all of them. PFA-100 ADP collagen closure time was prolonged in three patients with aortic stenosis and PFA-100 collagen epinephrine closure time was prolonged in four patients with aortic stenosis. Only two of four patients with prolonged PFA-100 collagen epinephrine closure time had a positive bleeding history (Table 8). PFA-100 ADP collagen closure times and PFA-100 collagen epinephrine closure times were performed only in patients with aortic and pulmonary stenosis. PFA-100 ADP collagen closure time was normal in 40 (66.7%) patients and prolonged 300 PFA-100 CEPI (sec)

Table 3. Demographic features of the study groups

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250 200 150 100 50 10.00

20.00

30.00

40.00

50.00

60.00

Mean pressure gradient (mmHg)

Fig. 1. Correlation between PFA-100 collagen epinephrine closure time and mean pressure gradient in patients with aortic stenosis.

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Table 7. The comparison of haematological parameters of patients with pulmonary stenosis according to degree of stenosis Moderate pulmoMild pulmonary nary stenosis Severe pulmonary stenosis (n = 3) (n = 25) stenosis (n = 4) (mean ± SD) (mean ± SD) (mean ± SD) p-value 308.330 ± 52.596 345.800 ± 97.395 459.000 ± 123.018

Platelet (cells/mm³)

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Table 9. Abnormal haematological parameters between the study groups Aortic stenosis Pulmonary stenosis (n = 28) (n = 32)

p-value

Low VWF:Ag, n (%)

3 (100)

0.08

Low VWF:RCo, n (%)

3 (75)

1 (25)

0.16

0.01

Prolonged PFA-100 CADP, n (%)

11 (55)

9 (45)

0.36

Prolonged PFA-100 CEPI, n (%)

19 (56)

15 (44)

0.10

1 (50)

0.66

PT (s)

13.667 ± 0.152

13.204 ± 0.649e

14.000 ± 0.812

0.05

Low VWF:RCo/VWF:Ag, n (%)

aPTT (s)

36.267 ± 7.691f

29.920 ± 2.186

29.775 ± 3.313

0.007

Factor VIII (%)

93.33 ± 21.07

116.80 ± 44.65

149.50 ± 73.07

0.28

VWF:Ag (%)

91.67 ± 28.00

95.88 ± 28.72

119.50 ± 48.37

0.35

VWF:Ag: von Willebrand factor antigen; VWF:RCo: ristocetin co-factor; PFA-100 CADP: platelet-function analyser-100 adenosine diphosphate collagen closure time; PFA-100 CEPI: platelet-function analyser-100 collagen epinephrine closure time.

VWF:RCo (%)

115.00 ± 22.71

93.88 ± 33.86

105.25 ± 46.21

0.54

PFA-100 CADP (s)

163.00 ± 98.73

110.72 ± 39.85

124.50 ± 29.49

0.18

PFA-100 CEPI (s)

157.00 ± 37.16

169.64 ± 54.78

167.75 ± 81.80

0.93

1.33 ± 0.57

0.99 ± 0.33

0.87 ± 0.14

0.20

VWF:RCo/ VWF:Ag

PT: prothrombin time; aPTT: activated partial thromboplastin time; PFA-100 CADP: platelet-function analyser-100 adenosine diphosphate collagen closure time; PFA-100 CEPI: platelet-function analyser-100 collagen epinephrine closure time; VWFAg: von Willebrand factor antigen; VWF:RCo: ristocetin co-factor. d p = 0.04, severe versus mild pulmonary stenosis. e p = 0.003, severe versus moderate pulmonary stenosis. f p = 0.002, mild versus moderate pulmonary stenosis, p = 0.009, mild versus severe pulmonary stenosis.

in 20 (33.3%). Of the 20 patients, 11 had aortic stenosis (six had mild stenosis, three had moderate stenosis and two had severe stenosis), and nine had pulmonary stenosis (one mild, six moderate and two severe). PFA-100 collagen epinephrine closure time was prolonged in 34 (56.6%) patients. Of these, 19 had aortic stenosis (12 mild, four moderate, three severe) and 15 had pulmonary stenosis (one mild, 12 moderate, two severe). A total of 16 of 60 patients who underwent PFA-100 ADP collagen closure time and PFA-100 collagen epinephrine closure time evaluation had a positive bleeding story (26.6%) (Tables 9, 10).

Discussion VWD is classified into three main types. Type 1 VWD is the most common form of disorder and is characterised by a mildto-moderate decrease in the plasma levels of VWF. Plasma VWF from these individuals has a normal structure. Plasma levels of ristocetin co-factor and factor VIII tend to be proportionately decreased. Type 2A VWD is associated with the absence of large highmolecular-weight (HMW) multimers from the plasma and

Table 10. Relationship between VWF components and PFA-100 CEPI and CADP PFA-100 CADP

PFA-100 CEPI

Normal Prolonged Normal Prolonged (n = 40) (n = 20) p-value (n = 26) (n = 34) p-value Low VWF:Ag

0.20

–

0.12

Low VWF:RCo

0.06

–

0.07

Positive bleeding history

0.06

0.26

platelets, which results in impaired ability to mediate platelet adhesion to the endothelium. This type is common in patients with cardiac stenotic diseases, especially in aortic stenosis, because of mechanical destruction of the HMW multimers of VWF under high shear stress. Type 2B VWD is a rare form of VWD characterised by partial loss of plasma HMW multimers and increased responsiveness when exposed ex vivo to the antibiotic ristocetin. In Type 3 VWD, plasma levels of factor VIII and VWF are virtually undetectable.12 Studies to investigate the prevalence of VWD in patients with heart disease are mainly carried out in adults. Although there are few reports in children, the relationship between congenital heart defects and VWD has been elucidated. In a study by Arslan et al., the prevalence of VWD was reported as 12.2% in 49 children with congenital heart disease.13 We found a similar ratio in our study (11.6%). In a study by Gill et al.,14 abnormal von Willebrand factor multimers was detected in all patients with congenital heart disease, but only six of them had low VWF:Ag levels. In this study, multimer analysis was repeated on five patients after surgical correction, and four patients had a normal multimeric

Table 8. The clinical and laboratory features of patients with abnormal platelet aggregation tests and low VWF:RCo/VWF:Ag Age

Diagnosis

Gender

Symptoms

Blood group

VWF: Ag (%)

VWF: Rco (%)

PFA-100 CADP

PFA-100 CEPI

VWF:RCo/ VWF:Ag

PPG (mmHg)

MPG (mmHg)

B RH+

55 (↓)

99 (N)

100 (N)

179 (↑)

1.8 (N)

AB RH+

52 (↓)

46 (↓)

135 (↑)

263 (↑)

0.88 (N)

AB RH+

64 (↓)

131 (N)

277 (↑)

196 (↑)

2 (N)

Yes

O RH+

56 (N)

49 (↓)

115 (N)

212 (↑)

0.87 (N)

18 21

A RH+

62 (N)

49 (↓)

210 (↑)

182 (↑)

0.79 (N)

O RH+

58 (N)

46 (↓)

155 (↑)

185 (↑)

0.79 (N)

Yes

O RH+

75 (N)

53 (N)

137 (↑)

217 (↑)

0.7 (N)

AS: aortic stenosis; F: female; M: male; MPG: mean pressure gradient; N: normal; PS: pulmonary stenosis; PFA-100 CADP: platelet-function analyser-100 adenosine diphosphate collagen closure time; PFA-100 CEPI: platelet-function analyser-100 collagen epinephrine closure time; VWF:Ag: von Willebrand factor antigen; VWF:RCo: ristocetin co-factor; PPG: peak pressure gradient. *This patient’s platelet aggregation curve induced by ristocetin was horizontal.

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structure. Rauch et al. reported four cases of VWD in 12 patients with patent ductus arteriosus, and six months after closure of the defect, the abnormal levels of von Willebrand factor had returned to within normal limits.15 The prevalence of bleeding episodes in patients with congenital heart disease is controversial. Several studies reported that at least 20% of patients with aortic stenosis have a history of bleeding.16-18 In the study by Gill et al.,14 66% of patients with ventricular septal defect and 50% with aortic stenosis had bleeding episodes; however, patients with atrial septal defect did not have bleeding episodes. Most of these episodes were excessive bleeding after injury or epistaxis, and easy bruising. In our study, two of seven patients (28.5%) with a diagnosis of AVWS had episodes of bleeding. These two patients had aortic stenosis, and PFA-100 collagen epinephrine closure time was prolonged in both. Although Froom et al.19 reported a relationship between frequent epistaxis episodes and low VWF:Ag, Rauch et al.15 did not find any association between bleeding episodes and the von Willebrand deficiency in patients with patent ductus arteriosus. We also could not find any relationship between low VWF:Ag levels and bleeding. In our study, it was noteworthy that 33% of patients with pulmonary stenosis with prolonged PFA-100 ADP collagen closure times had bleeding episodes. The PFA-100 closure time has been suggested to be useful in screening for disorders of primary haemostasis, including VWD.9 In a study by Vincentelli et al.,17 92% of patients with severe aortic stenosis and 50% of patients with moderate aortic stenosis had prolonged PFA-100 ADP collagen closure times. However, in our study, 67% of the patients with severe aortic stenosis and 50% of those with moderate aortic stenosis had prolonged PFA-100 ADP collagen closure times. In our study, PFA-100 collagen epinephrine closure time was prolonged in all seven patients with impaired haematological parameters matching AVWS. Only one of seven patients had a bleeding history, which suggests that absence of a history of bleeding episodes is not enough to exclude AVWS. For this reason, patients with heart disease should be screened with PFA-100 closure time before any intervention or surgery, even in the absence of a history of bleeding. Yoshida et al.16 reported a positive correlation between VWF:Ag levels and effective valvular area in patients with aortic stenosis. However, we were unable to show any correlation between degree of valvular stenosis and VWF:Ag and VWF:RCo levels. Routine coagulation tests and thrombo-elastometric measurements are not enough to exclude a diagnosis of AVWS. Laboratory investigations for AVWS include patient’s history of bleeding (lifelong mucocutaneous, postoperative bleeding) and a family history of bleeding, screening procedures [e.g. platelet count, PTT, concentration of factor VIII (FVIII:C), PFA-100], and confirming tests [e.g. VWF:Ag, VWF:RCo, VWF collagenbinding activity (VWF:CB), RIPA, and analysis of VWF multimers by gel electrophoresis].20 Most of these tests are time consuming and available only in specific centres. In some studies, the VWF:Ag levels were normal, while in others, they were low.13, 15,17,20 In our study, VWF:Ag levels were normal in all the healthy children and in the patients with aortic stenosis, but they were low in 5% of patients with pulmonary stenosis. The VWFR:RCo level was low in one of three patients

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with low VWF:Ag levels. In a study by Gill et al.,14 VWF:RCo levels were low in seven of 12 patients with acyanotic congenital heart disease. Only 6% of our patients had low VWF:Ag levels. Early diagnosis of AVWS is difficult, due to a lack of sensitivity of the tests used. Tiede et al.5 found that the sensitivity of PFA-100 was 80% for the diagnosis of AVWS. In our study, the sensitivity of VWF:Ag (23%) and VWF:RCo/VWF:Ag < 0.7 (26%) was too low to rule out this disease. We agree with Tiede et al.5 They suggested that a substantial number of patients present with normal or abnormal test results. The analysis of VWF multimers should always be part of the diagnostic workup.5 There were some study limitations; first, the low number of patients in each group, and second, for financial reasons, we could not perform VWF multimer analysis, which is the gold-standard test for the diagnosis of acquired VWD. Finally, patients in the control group did not undergo PFA-100 closure time tests.

Conclusion We suggest that a history of bleeding must be evaluated carefully in patients with stenotic obstructive heart disease, due to the increased risk of AVWS. Furthermore, even in the absence of bleeding episodes, PFA-100 closure time should be evaluated routinely before any intervention or surgery. If any abnormality is detected, VWF:Ag, VWF:RCo and platelet aggregation tests should be performed. Although the gold-standard test for AVWS is detection of structural abnormalities of VWF, it is a time-consuming assay and not available in most centres.

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10. Mammen EF, Alshameeri RS, Comp PC. Preliminary data from a field trial of the PFA-100 system. Semin Thromb Hemost 1995; 21(Suppl 2): 113–121. 11. Fressinaud E, Veyradier A, Truchaud F, et al. Screening for von Willebrand disease with a new analyzer using high shear stress: a study of 60 cases. Blood 1998; 91: 1325–1331. 12. Karayalçın, G. Von Willebrand’s disease. In: Lanzkowsky P (ed). Manual of Pediatric Hematology and Oncology, 3rd edn. San Diego: Academic Press, 2000: 310–315. 13. Arslan MT, Ozyurek R, Kavaklı K, Levent E, Ulger Z, Gurses D. Frequency of acquired von Willebrand’s disease in children with congenital heart disease. Acta Cardiologica 2007; 62: 403–408. 14. Gill JC, Wilson AD, Endres-Brooks J, Montgomery RR. Loss of the largest von Willebrand factor multimers from the plasma of patients

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with congenital cardiac defects. Blood 1986; 67: 758–761. 15. Rauch R, Budde U, Koch A, Girisch M, Hofbeck M. Acquired von Willebrand syndrome in children with patent ductus arteriosus. Heart 2002; 88: 87–88. 16. Yoshida K, Tobe S, Kawata M, Yamaguchi M. Acquired and reversible von Willebrand disease with high shear stres aortic valve stenosis. Ann Thorac Surg 2006; 81: 490–494. 17. Vincentelli A, Susen S, Le Tourneau T, Six I, Fabre O, Juthier F. Acquired von Willebrand syndrome in aortic stenosis. N Engl J Med 2003; 349: 343–349. 18. Carrasco E, Lopez R, Rattalino M, Lema G, Pereira J, Canessa R. Aortic stenosis and acquired von Willebrand disease: lack of association. J Cardiothorac Vasc Anesth 2011; 25: 615–618. 19. Froom P, Marqulis T, Grenadier E, Palant A, David M. Von Willebrand factor and mitral valve prolapse. Thrombosis Haemostasis 1988; 60: 230–231. 20. Pareti FI, Lattuada A, Bressi C, Zanobini M, Sala A, Steffan A. Proteolysis of von Willebrand factor and shear stres-induced platelet aggregation in patients with aortic valve stenosis. Circulation 2000; 102: 1290–1295.

The role of interventions in secondary hypertension Interventional therapies can benefit patients with secondary hypertension significantly in respect of mortality and quality of life. Notable causes of secondary hypertension that are amenable to intervention are coarctation of the aorta and renal artery stenosis (RAS). Five percent of hypertension derives from secondary causes, Prof Ikechi Okpechi from the University of Cape Town told delegates at AfricaPCR 2016. ‘We need to look for the common clinical clues suggestive of secondary hypertension and investigate accordingly. Work-up requires a clear strategy. A patient history needs to be taken and a physical examination performed. Twenty-four-hour ambulatory blood pressure monitoring is necessary to rule out primary hypertension. Only screen where there is a clinical suspicion and start with simple tests.’

Coarctation of the aorta This is an important cause of secondary hypertension and one that is often missed, according to Johannesburg cardiologist, Dr Jeff Harrisberg. ‘The clinical clue is weak or absent femoral pulses. The primary treatments are surgery and/or catheter interventions.’ Indications for coarctation stenting include long-segment coarctation and associated hypoplasia. Covered stents are preferred and balloon angioplasty should be avoided, as it is associated with an uncontrolled response. Possible complications to be aware of include: • dissection, aneurysm and rupture • femoral artery damage • stent migration • cerebrovascular accident or peripheral embolism.

Nice-to-haves when undertaking coarctation stenting include a biplane cath lab, surgical and ICU back up, general anaesthesia, percutaneous femoral artery closure devices and CT angiography. ‘But all of these are not always available in real-world settings. Minimum requirements are a singleplane cath lab with accurate measuring capabilities, the full range of one brand of bare or covered stents, a variety of appropriately sized balloons and large, long sheaths (12–16 Fr)’, concluded Dr Harrisberg.

RAS The goal of renal angioplasty and/or stenting for RAS is to provide renal parenchymal protection while controlling blood pressure, which can be challenging, and preventing cardiovascular events. ‘RAS is a relatively uncommon condition, with an incidence of less than 2%’, said Dr Yemi Johnson, from Lagos, Nigeria. Ultrasound can be a useful investigative tool in good hands, but the healthcare practitioner needs to be trained to look for RAS. ‘Most instances of RAS are unproblematic, so we need clear indications to intervene. These include resistant hypertension, drug intolerance and ischaemic nephropathy.’ Dr Johnson summed up as follows. ‘Renal stenting is a simple procedure if done properly, certainly simpler than coronary artery stenting. It has a high success rate and, while serious complications are possible, these can be prevented with the correct technique.’ Source: AfricaPCR 2016

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Correlation analysis between ApoM gene-promoter polymorphisms and coronary heart disease Yao Zhang, Li-Zhu Huang, Qing-Ling Yang, Yan Liu, Xin Zhou

Abstract Objectives: Apolipoprotein M (ApoM), a 25-kDa plasma protein belonging to the lipocalin protein family, is predominantly associated with high-density lipoprotein cholesterol (HDL-C). Studies have suggested ApoM to be important for the formation of pre-β-HDL and to increase cholesterol efflux from macrophage foam cells. The aim of this study was to explore the association of single-nucleotide polymorphisms (SNPs) in the ApoM promoter with coronary atherosclerotic disease (CAD), and the contribution of latent factors. Methods: ApoM was measured in samples from two separate case–control studies, of whom 88 patients developed CAD and 88 were controls. Whole-blood samples from subjects were genotyped by PCR-restriction fragment length polymorphism (PCR-RFLP). Luciferase activities were measured for HepG2 cells with two SNPs, rs805296 (T-778C) and rs940494 (T-855C), and after interfering with or overexpressing the predicted transcription factors. The ability of the SNPs to combine with nucleoproteins was analysed by electophoretic mobility shift assay (EMSA). Results: Mean plasma ApoM concentrations in the CAD and non-CAD groups were 9.58 ± 4.30 and 12.22 ± 6.59 μg/ ml, respectively. Correlation studies of ApoM concentrations with several analytes showed a marked positive correlation with HDL-C, fasting plasma glucose and triglyceride levels. The CC genotype showed lower luciferase activities compared to the TC and TT genotypes. The ApoM-855 mutant-type could bind to the AP-2α. Interference and overexpression of AP-2 increased and decreased luciferase activities of the wild and mutant types to different degrees. Conclusion: ApoM may be a biomarker of CAD. ApoM855 T→C substitution provides binding sites for AP-2α and reduces ApoM transcription activity.

Department of Biochemistry and Molecular Biology, Wan Nan Medical College, Anhui 241000, China Yao Zhang, MD, zhangyao@ahedu.gov.cn

Clinical Testing and Diagnosis, Experimental Centre of Bengbu Medical College, Anhui 233000, China Li-Zhu Huang, MD Yan Liu, MD

Department of Biochemistry and Molecular Biology, Beng Bu Medical College, Anhui 233000, China Qing-Ling Yang, MD

Centre for Gene Diagnosis, Zhongnan Hospital, Wuhan University, Wuhan, Peoples’ Republic of China Xin Zhou, PhD

Keywords: coronary heart disease, ApoM, SNP, luciferase activity Submitted 21/3/14, accepted 8/1/16 Cardiovasc J Afr 2016; 27: 228–237

www.cvja.co.za

DOI: 10.5830/CVJA-2016-001

Apolipoprotein M (ApoM) is a novel lipocalin superfamily protein.1,2 Although also found in low-density lipoprotein (LDL), very low-density lipoprotein and chylomicrons, ApoM is primarily found in high-density lipoprotein (HDL) where it binds to sphingosine-1-phosphate (S1P) anchors.3 Recent studies have suggested that ApoM may affect HDL metabolism, increasing the formation of pre-β-HDL particles.4 ApoM has been shown to protect LDL against Cu++-induced oxidation,5 and to contribute to the anti-inflammatory function of HDL.6 Small circulating HDLs are involved in reverse cholesterol transport, and ApoM may affect this process by regulating pre-β-HDL.7 The binding of ApoM to S1P in HDL particles may also have an antioxidant role. By affecting the immune and anti-inflammatory functions of HDL,8,9 ApoM may reduce atherosclerosis-related inflammation, preventing the onset and development of atherosclerosis. We used the online prediction software, TRANSFAC, to predict transcription factor (TF) binding sites for the normal and mutated ApoM-855 and ApoM-778 sites, and found ApoM T-855C provided binding sites for activating protein 2 (AP-2). Activating protein-2α (AP-2α) was one of the first identified and studied TFs.10 The AP-2 gene, encoding a 437-amino acid protein of ~52 kDa, regulates the transcription of various genes regulating embryonic development, cell growth and differentiation.11,12 Vertebrates possess five subtypes of AP-2, α, β, γ, δ and ε.13,14 The AP-2 protein has been shown to regulate atherosclerosisassociated genes, including matrix metalloproteinase-2, vascular endothelial growth factor, ApoE, tryptase and adiponectin ATP-binding cassette transporter AI (ABCAI). In addition, AP-2α plays a role in atherosclerosis. It may mediate foam cell formation in mouse and human atherosclerotic lesions.15 AP-2α was found in ApoE-/- mouse lesions within the artery wall, but was not detected in mouse arteries without atherosclerotic lesions. Similarly, AP-2α was observed in the human atherosclerotic aortic wall, mainly within the atherosclerotic plaque. Recent studies involving the genetics of ApoM have led to major breakthroughs in metabolic and disease characteristics. In particular, associations have been found between diabetes and polymorphisms in the promoter region of the ApoM gene.16,17 To explore the association of ApoM gene polymorphisms with coronary heart disease (CHD) in a Chinese Han population, we performed a population-based case–control study. We examined whether ApoM promoter polymorphisms could lead to changes in TF binding, and therefore, changes in promoter activity.

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Methods All relevant ethical and clinical approvals were obtained for this study. A total of 88 patients with coronary atherosclerotic disease (CAD) (63 males; mean age 60.80 ± 9.27 years) and 88 unrelated control individuals (53 males; mean age 58.18 ± 10.43 years) were retrospectively enrolled from among in-patients at the Anhui Cancer Hospital of Bengbu Medical College, Bengbu City, Anhui Province, China. All participants were of Han Chinese descent. The criterion for inclusion in the CAD group was ≥ 50% stenosis in at least one major segment of the coronary arteries, determined by coronary artery angiography. Individuals in the control group had negative coronary artery angiography results (used to rule out CAD). A history of conventional risk factors for CAD or hypercholesterolaemia (total cholesterol ≥ 5.7 mmol/l) was obtained from the medical records. Exclusion criteria for both groups were familial hypercholesterolaemia, diabetes mellitus, cancer, renal disease and any other chronic illness. For lipid analysis, whole blood samples were drawn from all participants in the morning after a 12-hour fast. Fasting plasma glucose (FPG), triglyceride (TG), HDL and LDL cholesterol (HDL-C and LDL-C), and total cholesterol (TC) levels were determined for each subject using an automated chemistry analyser (AU2000; Olympus Promarketing, Tokyo, Japan). For ApoM analysis, 5-ml blood samples were collected in EDTA (as anticoagulant) after an overnight fast. Samples were centrifuged at 3 000 rpm for 10 minutes at room temperature. Separated sera were stored at –40°C. The serum ApoM level was quantified with ELISA, using horseradish peroxidase and the anti-ApoM antibodies 1E2 and 8F12 (Hunan Far Tai Biotechnology Co, Ltd). To minimise errors, all samples were used in the same reaction system, each test was repeated twice, and the average value was used in the final analysis.

Genotyping of ApoM T-778C and T-855C polymorphisms Genomic DNA was extracted from peripheral blood using a salting-out protocol. Single-nucleotide polymorphisms (SNPs) selected for this study are recorded in the public dbSNP database (http://www.ncbi.nlm.nih.gov/SNP/). Two SNPs, rs9404941 (T-855C) and rs805296 (T-778C), representing at least 5% of the minor allele frequency in the promoter region of the ApoM gene in the Han Chinese, were genotyped with the PCR-RFLP method. PCR primers for the ApoM T-855C and T-778C SNPs were designed based on GeneBank sequences (Table 1). PCR products were digested in a total volume of 20 μl containing 1 μg of PCR product, 5 U of restriction endonuclease HaeIII (-855) or RsaI (-778) (Fermentas, USA), 2 μg acetylated bovine serum albumin (BSA), and 2 μl restriction enzyme buffer. ApoM polymorphisms were detected with 2.5% agarose gel (Promega, USA) electrophoresis and visualised with ethidium bromide staining. In the T-778C polymorphism of the ApoM gene, the T→C substitution creates an RsaI restriction site. The PCR product with the C allele was digested into two fragments (272 and 164 bp), whereas the T allele was not cut by RsaI. Because T-855C contains GGC(C/T) and HaeIII recognises the GGCC sequence, digestion of TT homozygotes with HaeIII produced 496-,

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Table 1. Primer, probe and oligonucleotide sequences used in this study Primer/probe/oligonucleotide

Sequence

ApoM T-855C and T-778C SNPs 5′-GGTACCGTCTTTGCTAAGGGCTTTATGTGCATTA-3′ (forward) 5′-AAGCTTTGTTGGTGTCAGGCAGAATGTGTCCAA-3′ (reverse) Luciferase reporter plasmids

5′-AAGCTTCTCCTACTCGGGAATCAT-3′ (forward) 5′ -GGTACCTCCAGAGCCTCCACCATA-3′ (reverse)

Wild-type T1 probe synthesis

5′ -TCGACATCCCAGGCTCAAGCAATCCT-3′

Wild-type T2 probe synthesis

5′-AGGATTGCTTGAGCCTGGGATGTCGA-3′

Mutant-type C1 probe synthesis

5′-TCGACATCCCAGGCCCAAGCAATCCT-3′

Mutant-type C2 probe synthesis

5′-AGGATTGCTTGGGCCTGGGATGTCGA-3′ Sense 5′-UUCUCCGAACGUGUCACGUTT-3′

Negative control

Anti-sense 5′-ACGUGACACGUUCGGAGAATT-3′

Interference fragment 1

Sense 5′-CCAGAUCAAACUGUAAUUATT-3′ Anti-sense 5′-UAAUUACAAGUUUGAUCUGGTT-3′

Interference fragment 2

Sense 5′-GGAAGAUCUUUAAGAGAAATT-3′ Anti-sense 5′-UUUCUCUUAAAGAUCUUCCTT-3′

Interference fragment 3

Sense 5′-CCUGCUCAUCACUAGUATT-3′ Anti-sense 5′-UACUAGUGAUGUGAGCAGGTT-3′

AP-2α forward primer

5′-CTGGGCACTGTAGGTCAATCT-3′

AP-2α reverse primer

5′-CCTCCTCGATGGCGTGAG-3′

GAPDH forward primer

5¢-CAAGGTCATCCATGACAACTTTG-3¢

GAPDH reverse primer

5¢-GTCCACCACCCTGTTGCTGTAG-3¢

352- and 84-bp fragments, CC homozygotes gave 84-, 157- and 195-bp products, and CT heterozygotes produced 84-, 157-, 195and 436-bp fragments (Fig. 1C). To confirm detection of the T-778C polymorphism of the ApoM gene by PCR-RFLP, the PCR products were also sequenced (Shengong Bio Company, Shanghai, China; Fig. 1C).

Construction of luciferase reporter plasmids Four reporter plasmids, encompassing −1334 to +227 bp of the human ApoM promoter, were constructed. Plasmids containing the regions −855T to −778T, −855T to –778C, −855C to −778T and −855C to −778C were named PGL3-TT, PGL3-TC, PGL3CT and PGL3-CC, respectively (Fig. 2). PCR primers containing a KpnI- and HindIII-inserted forward and reverse primer, respectively, used in the construction of these plasmids, are shown in Table 1. PCR products were first ligated into the PMD®8-T vector (Takara, Dalian, China) and then subcloned into the pGL3-basic vector (Promega). All constructs used in this study were sequenced to confirm their authenticity (Fig. 2).

Cell culture, transfection and reporter gene assays The HepG2 hepatoma cell line was propagated in Dulbecco’s modified Eagle’s medium (DMEM; Hyclone) with 10% foetal calf serum (FCS) and maintained in 5% CO2 at 37°C. HepG2 cells were seeded in 24-well plates at 5 × 104 HepG2 cells/ well, respectively. After 24 hours, the cells were transfected with pGL3-basic (a promoterless control), pGL3-promoter (a promoter control), and two pGL3-basic constructed plasmids. The pRL-SV40 plasmid (Promega) was co-transfected as a normalising control. To prevent transcription of ApoM by endogenous liver cell extracts, recombinant vector transfection groups were set up with and without liver cell extracts. After six hours, the AP-2α siRNA and AP-2α siRNA negative control fragments (Gima) were co-transfected into the two pGL3-basic constructed plasmid groups. The AP-2α expression vector (GeneChem)

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1000 bp 7000 bp 5000 bp 3000 bp 2000 bp

1561 bp

1500 bp

489 bp

436 bp

353 bp

352 bp 272 bp

242 bp 195 bp

1000 bp

190 bp

164 bp

147 bp 110 bp

157 bp

89 bp

84 bp

Fig. 1. P CR-RFLP analysis results for the ApoM T-855/-778C locus. A. Sequencing map for PCR products of the two alleles of T-855C and T-778C. B. ApoM gene promoter pGL3 reconstruction vector (6379 bp) with restriction enzyme digestion. Lane M: ladder of molecular size markers; lane 1: vectors without restriction enzyme digestion; and lane 2: vectors with restriction enzyme digestion. C. Electrophoresis results of SNPs of ApoM proximal promoter and products separated on a 3% agarose gel and stained with ethidium bromide. Lane M: marker; lane 1: missed cleavages; lane 2: -T855C/C; lane 3: -T855C/TC; lane 4: T-778C/CC; lane 5: T-778C/TC.

groups were transfected into cells in the same way. Transfections were performed in triplicate and repeated three times. After 48 hours of incubation, the cells were collected and analysed for luciferase activity with the Dual-Luciferase Reporter assay system (Promega).

Nuclear extract preparation HepG2 cells were propagated in DMEM with 10% FCS under 5% CO2 at 37°C. Cells were cultured to a density of 1 × 107 cells/ ml, according to the manufacturer’s instructions. HepG2 cells were collected in a 15-ml centrifuge tube and centrifuged at 500 g for five minutes at 4°C. The supernatant was discarded, and the cells were washed three times with 2 ml of pre-cooled phosphate-buffered saline (PBS). For every 20 μl of cell sedimentation, 200 μl of reagent A were added. The mixture was vortexed for five seconds, which caused the cell sedimentation to disperse completely, and placed in an ice bath for 15 minutes. Then 10 μl of cell plasma protein extraction reagent B was added to the solution. The mixture was vortexed for five seconds, placed in an ice bath for one minute, vortexed

again for five seconds, and centrifuged at 16 000 g for five minutes at 4°C. The supernatant was removed and 50 μl of phenylmethylsulfonyl fluoride (PMSF) nuclear protein extraction reagent was added. The mixture was vortexed for 30 seconds, placed on ice for two minutes, and vortexed again for 30 seconds. This cycle was repeated for a total of 30 minutes. After a final centrifugation at 16 000 g for 10 minutes at 4°C, the nuclear extract was drawn into a pre-cooled 1.5-ml centrifuge tube.

Electrophoretic mobility shift analysis (EMSA) Sequences used for T1 and T1 wild-type and C1 and C2 mutanttype probe synthesis (Table 1) were labelled and unlabelled with biotin, respectively. Nuclear cell extracts were incubated with biotin-labelled double-stranded (ds) oligonucleotide probes containing the wild-type or mutant AP-2α binding sites in the ApoM promoter (wild-type T1 and mutant-type C1 probes in Table 1). Competition analysis was performed using the mutant probe with the AP-2α site. Supershifts were performed with antibodies against AP-2α and Sp1 (Abcam).

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Fig. 2. p GL3 recombination vector sequencing. PGL3-TT means the regions −855T to −778T; PGL3-TC means the regions −855T to –778C; PGL3-CT means the regions −855C to −778T; PGL3-CC means the regions −855C to −778C.

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Small interference (siRNA) transfection and efficiency Three pairs of ds siRNA oligonucleotides were obtained from Gima Biotechnology Company, China (Table 1). Five groups were designated: a blank control group, a negative control group and three siRNA groups. Cells were seeded in six-well plates at 1 × 105 cells/well, and then 1 μl of siRNA and 2 μl of siRNAMate were added to each well, corresponding to a density of 70 to 80% at the time of transfection. All of the steps were strictly performed according to the manufacturer’s specifications. Cells were harvested at 48 hours. The Sp1 siRNA and AP-2α expression vectors were transfected into cells in the same way. Six hours after fluorescein amidite (FAM)-labelled siRNA transfection, fluorescence was observed under a fluorescence microscope. The transfection efficiency was determined as: number of fluorescent cells ___________________      × 100%. efficiency =      total number of cells

Semiquantitative RT-PCR Total RNA was extracted from the cell clones using TRIzol reagent (Invitrogen). The cDNAs were reverse-transcribed from total RNA. The primers used are shown in Table 1. The sizes of the PCR products for AP-2α and GAPDH were 385 bp and 496 bp, respectively. PCR products were checked by agarose gel electrophoresis. The abundance of each mRNA was detected and normalised to that of GAPDH mRNA.

Western blotting analysis Cells in all groups were collected after 72 hours, and the total protein was extracted with RIPA lysis buffer (Beyotime Institute of Biotechnology). The protein was measured with the BCA protein assay and diluted with cell lysate to an equal concentration in each group (40 μg protein/group). A 10% SDS-PAGE analysis was performed. The proteins were transferred to a PVDF membrane (250 mA, 2 h), blocked with 5% BSA in PBS containing Tween-20 (PBST), and incubated with a 1:500 dilution of anti-AP-2α overnight at 4°C.

Table 2. Clinical data for CAD and control groups CAD Index

p-value

ACS group

SAP group

Age (years)

60.80 ± 9.27

61.00 ± 8.68

62.96 ± 11.22

0.400

58.18 ± 10.43

0.081

Male/female

63/25

20/11

25/22

0.091

53/35

0.112

Number (cases)

Control

p-value

All CAD

TC (mmol/l)

4.47 ± 1.41

5.17 ± 1.34

4.34 ± 1.44

0.010

4.33 ± 0.49

0.299

TG (mmol/l)

1.97 ± 1.28

2.20 ± 1.27

2.09 ± 1.32

0.698

1.02 ± 0.37

0.000#

HDL-C (mmol/l)

1.05 ± 0.25

1.05 ± 0.21

1.04 ± 0.27

0.951

1.32 ± 0.21

0.000#

LDL-C (mmol/l)

2.70 ± 1.23

3.01 ± 1.48

2.61 ± 1.05

0.140

2.53 ± 0.41

0.202

FPG (mmol/l)

6.40 ± 2.40

8.36 ± 4.11

11.58 ± 3.88

0.000*

4.86 ± 0.45

0.000#

ApoM (μg/ml)

9.58 ± 4.30

12.22 ± 6.59

0.037

Gensini score

6.55 ± 2.74

6.32 ± 2.20

0.670

80.48 ± 72.46

45.96 ± 51.00

0.017*

The membrane was washed with TBST and incubated with a peroxidase-conjugated secondary antibody (1:1 000) for two hours. Specific antibody binding was detected using a chemiluminescence detection system, according to the manufacturer’s recommendations. Net intensities of the bands on the Western blots were quantified using Tanon GIS software. After development, the membrane was stripped and re-probed with antibody against β-actin (1:1 000) to confirm equal sample loading.

Statistical analysis Unless otherwise noted, results are reported as mean ± standard deviations (SD). General characteristics in the two groups and serum ApoM levels for different genotypes were statistically evaluated using the unpaired Student’s t-test (Prism software, version 4; GraphPad Inc, La Jolla, CA, USA). Significance was established at a p-value < 0.05.

Results Association between risk factors for CAD and ApoM levels in CAD patients The general characteristics of age- and gender-matched non-CAD and CAD patients are shown in Table 2. The mean ages of the non-CAD and CAD patients were 60.80 ± 9.27 and 58.18 ± 10.43, respectively (p > 0.05). The CAD patients had higher TG (1.97 ± 1.28 mmol/l) and FPG levels (6.40 ± 2.40 mmol/l), and lower HDL-C levels (1.05 ± 0.25 mmol/l) than the non-CAD patients (all p = 0.000). According to their clinical symptoms and the American College of Cardiology (ACC)/American Heart Association (AHA) diagnostic guidelines, patients in the CAD group were divided into two subgroups: acute coronary syndrome (ACS group, n = 31) and stable angina pectoris (SAP group, n = 57). The Gensini score was calculated for the CAD group and both CAD subgroups, according to CHD severity (Table 2). The ACS group had a higher average Gensini score (80.48 ± 72.46, p = 0.017), higher TC level, and lower ApoM level than the SAP group (all p > 0.05). Analysis of the serum ApoM lipid levels of the patients showed that the serum level of ApoM was positively correlated with HDL-C and negatively correlated with LDL-C and TG levels (p < 0.05). The correlation with TC was not significant (Table 3). We used the multiple linear stepwise regression method to describe the relationship between serum ApoM and HDL-C and LDL-C levels. Linear dependencies and other related indicators

Table 3. Multiple linear regression analysis of serum ApoM Partial regression coefficient

Constant

3.592

2.081

HDL-C

9.767

1.316

0.573

7.421 0.000*

FPG

–0.539

0.138

–0.301

–3.908 0.000*

–0.464

0.217

–0.138

–2.136 0.036*

1 branch

Index

2 branch

Data are means ± SD. *Indicates statistical significance (p < 0.05) compared with ACS and SAP group. # Indicates statistical significance (p < 0.05) compared to the control group with Dunnett’s test. Because the Gensini scores were skewed distributions, Napierian logarithmic transformation was applied for normalisation. CAD, coronary artery disease; ACS, acute coronary syndrome; SAP, stable angina pectoris; TC, total cholesterol; TG, triglyceride; HDL-C and LDL-C, high- and low-density lipoprotein cholesterol, respectively; FPG, fasting plasma glucose; ApoM, apolipoprotein M.

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Standard regression coefficient

t-value p-value 1.726 0.088

*Statistical significance (p < 0.05). HDL-C, high-density lipoprotein cholesterol; FPG, fasting plasma glucose; TG, triglycerides.

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were used as dependent variables (ŷ). ApoM was used as the independent variable (x). When HDL-C, FPG and TG were used as three indicators in the regression equation (x1, x2 and x3, respectively), the resulting general multiple regression equation was ŷ = 3.592 + 9.767x1 – 0.539x2 – 0.464x3 with the regression coefficient test, with p = 0.000 and a coefficient of determination of 0.651. The standardised regression equation was ŷ = 0.573x1 – 0.301x2 – 0.138x3. The closest relationship was between serum HDL-C and ApoM levels (Table 3).

Association of polymorphisms in the ApoM proximal promoter region with CAD DNA sequencing of the polymorphic regions in the proximal promoter of the ApoM gene revealed that SNPs T-778C and T-855C of the ApoM gene were valid in the Han Chinese. The SNPs were accurately detected by PCR-RFLP (Fig. 1). Table 4 shows the plasma lipid and ApoM levels in each group according to the SNP status. In the non-CAD group, we found no significant difference in lipid plasma levels between the TT and the TC+CC groups. In the CAD group, the TC, TG and ApoM levels were significantly different between the TT and TC/CC groups. The ApoM plasma levels in the TT and TC+CC groups were 10.35 ± 4.41 and 6.46 ± 4.06 μg/ml, respectively in T-778C, and 10.17 ± 5.68 and 6.07 ± 4.70 μg/ml, respectively in T-855C (both p < 0.05; Table 4).

PCR amplification of the human ApoM gene promoter region From the above results, that patients with the TC/CC genotype showed lower plasma ApoM levels compared to those with the TT genotype, and that ApoM levels were lower in CAD compared to non-CAD patients, we inferred that the ApoM promoter variation may alter the promoter activity. To verify whether the −778T→C and −855T→C variation affected the ApoM promoter activity, we applied a reporter gene assay to detect the luciferase expression with transfection of the specific allele(s) of the polymorphism. We designed PCR primers to amplify a 1562-bp slice of the promoter region of ApoM. Four reporter plasmids, containing

regions −855T to −778T, −855T to –778C, −855C to −778T and −855C to −778C, were named PGL3-TT, PGL3-TC, PGL3CT and PGL3-CC, respectively. The luciferase activities of the PGL3-TC/CC promoters were lower than those of the PGL3-TT promoter (PGL-TT, 1.67 ± 0.14; PGL-TC, 1.28 ± 0.11; PGL-TC, 0.77 ± 0.21; PGL-CC, 0.25 ± 0.10, p = 0.001; Fig. 3)

Transcription factor prediction The TRANSFAC online prediction software was used to predict the combined TF binding sites upstream of the ApoM gene (Fig. 4A, B). The binding sites were identified on the ApoM promoter for some TFs, including Sp1 and AP-1. The AP-2 binding sites were located at -479 to -488 bp, -349 to -358 bp and -104 to -113 bp on the ApoM promoter. TRANSFAC was used to predict TF binding sites for the normal and mutated ApoM-855 and ApoM778 sites (Fig. 4C–F). ApoM T-855C provided binding sites for AP-2. ApoM-778T but not ApoM-778C had binding sites for hepatocyte nuclear factor-3 (HNF-3), CCAAT enhancer binding protein (C/EBP) and TATA box-binding protein (TBP). We also used the TESS online software to predict TF binding sites of the normal and mutated ApoM-855 (Fig. 4G, H). The results showed that ApoM T-855C provided binding sites for AP-2.

EMSA results Small molecular weight chains have faster mobility in EMSA, whereas the electrophoretic velocity will vary with probe and protein binding. The velocity of the electrophoretic band indicates the presence or absence of binding. We observed a hysteresis band when the C probe but not the T probe was incubated with nuclear protein (Fig. 5A). The competitive inhibition test showed that the hysteresis band of the C probe incubated with nuclear protein could be suppressed by unlabelled C probe, but not by unlabelled T probe (Fig. 5A). The supershift test showed that incubating the C probe with the AP-2α antibody produced a more lagging band (Fig. 5B), whereas the Sp1 antibody did not have this effect (Fig. 5C). The antibody activity of Sp1 was verified by immunohistochemistry analyses (Fig. 6A).

Control

CT+CC

p-value

CT+CC

p-value

TC (mmol/l)

4.61 ± 1.37

4.77 ± 1.86

0.030#

4.33 ± 0.49

4.31 ± 0.52

0.900

TG (mmol/l)

1.93 ± 1.01

3.05 ± 2.02

0.002#

1.03 ± 0.82

0.82 ± 0.32

0.157

LDL-C (mmol/l)

2.78 ± 1.18

2.60 ± 1.46

0.610

2.53 ± 0.41

2.60 ± 0.44

0.655

HDL-C (mmol/l)

1.05 ± 0.23

1.01 ± 0.34

0.557

1.32 ± 0.22

1.35 ± 0.19

0.706

10.35 ± 4.41

6.46 ± 4.06

0.009#

13.22 ± 9.18

3.57 ± 3.86

0.007#

TC (mmol/l)

4.43 ± 1.39

5.30 ± 1.49

0.017#

4.34 ± 0.49

4.23 ± 0.54

0.542

TG (mmol/l)

1.95 ± 1.32

2.70 ± 1.04

0.020#

1.02 ± 0.38

0.97 ± 0.24

0.711

LDL-C (mmol/l)

2.66 ± 1.11

3.03 ± 1.52

0.237

2.54 ± 0.40

2.46 ± 0.50

0.604

HDL-C (mmol/l)

1.07 ± 0.26

0.97 ± 0.20

0.123

1.32 ± 0.21

1.32 ± 0.26

0.990

10.17 ± 5.68

6.07 ± 4.70

0.009#

12.87 ± 9.40

8.30 ± 6.45

0.184

Lipid parameter rs805296

ApoM (μg/ml) Rs9404941

ApoM (μg/ml)

Data are means ± SD. #Statistical significance (p < 0.05) for TC/CC genotype group vs TT group with Dunnett’s test. TC, total cholesterol; TG, triglyceride; LDL-C, HDL-C, low- and high-density lipoprotein cholesterol, respectively; ApoM, apolipoprotein M.

ApoM promoter activity

Table 4. Lipid profiles according to genotype CAD

233

50 45 40 35 30 25 20 15 10 5 0

Influence of SNPs T-778C and T-855C on ApoM prometer activity with liver cell extracts

PGL3basic

PGL3control group

PGL3TT PGL3TC PGL3CT PGL3CC

Fig. 3. Relative luciferase activity of the ApoM promoter for each transfected group. Data are means ± SD. All groups were significantly different (p < 0.05 by Dunnett’s test) compared to the PGL3-TT group.

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C D

F G

Fig. 4. P rediction of transcription factor binding in the ApoM promoter region. A. Transcription factor binding prediction for ApoM. B. AP-2α potential binding sites of ApoM start promoter sequence. Mutation site analysis for ApoM-855 and ApoM-778 by TRANSFAC (C–F), and by TESS (G–H). Binding sites with wild-type alleles of ApoM-855 (C), and ApoM-778 (D), and binding sites with mutant-type alleles of ApoM-855 (E), and ApoM-778 (F) by TRANSFAC. Binding sites with wild-type (C), and mutant-type alleles (D) of ApoM-855 by TESS.

Luciferase activity detection with AP-2α overexpression or interference

(Fig. 7C–F), and examined changes in the luciferase activity of the -855 site (Fig. 8). With AP-2α interference, the luciferase activity in cells expressing the wild-type allele (1.63) was less elevated compared to cells expressing the mutant-type allele (2.99). The luciferase activities of treated cells were increased compared to the

To examine the role of the TF AP-2α in mediating ApoM promoter activity, we treated HepG2 cells expressing the mutanttype or wild-type allele of the ApoM-855 site with siRNAs against AP-2α (Fig. 7A, B) or with an AP-2α expression vector

B 1

C AP-2α

DNA–protein complex DNA–protein complex

mutant probe + nuclear protein + Sp1 antibody

mutant probe + nuclear protein

free probe free probe

Fig. 5. E xperimental results for transcription factor binding in the ApoM promoter region. A. EMSA. Lane 1: biotin-labelled T probe + nuclear protein (NE); lane 2: biotin-labelled C probe; lane 3: biotin-labelled C probe + NE; lane 4: biotin-labelled C probe + NE + unlabelled C probe; and lane 5: biotin-labelled C probe + NE + unlabelled T probe. B. AP-2α supershift. Biotin-labelled C probe + NE + AP-2α antibody. C. Sp1 supershift. Biotin-labelled C probe + NE + Sp1 antibody.

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6 Luciferase activity

5 4

3 2 1 0

Fig. 6. C onfirmation of EMSA supershift results. To determine whether unbound Sp1 existed with the Sp1 antibody, we conducted cell immunohistochemistry experiments. Cells expressing Sp1 were dyed brown–yellow (B), and negative control cells were stained blue (A). The results indicated that the extracted nucleoprotein contained Sp1, and there was no problem in terms of the Sp1 antibody quality. These results further confirmed the EMSA supershift results.

untreated control cells (p < 0.05). With AP-2α overexpression, the activity levels of cells expressing the wild-type allele were more reduced than those of cells expressing the mutant-type (0.25 vs 0.38). The luciferase activity levels of AP-2α overexpressing cells were lower than those of the untreated A

TT untreated

CT AP-2α siRNA

GV141-AP-2α

Fig. 8. Relative luciferase activity of the ApoM promoter in each transfected group. For TT groups: untreated, 1.84 ± 0.12; TT + AP-2α, 4.83 ± 0.08; TT + GV141-AP-2α, 1.38 ± 0.06; For CT groups: untreated, 0.80 ± 0.08; CT + AP-2α, 3.19 ± 0.07; CT + GV141-AP-2α, 0.51 ± 0.05; PGL3-basic was used as a negative control and PGL3-control as a positive control. #p < 0.05 and p < 0.05 vs -855 wild-type untreated group and ##p < 0.05 and p < 0.05 vs -855 mutant untreated group by paired t-test.

cells (p < 0.05). Taken together, these results suggest that AP-2α was a negative regulatory factor for ApoM expression. E

Mark

600 bp 500 bp 400 bp 300 bp

AP-2AGAPDH

GAPDH AP-2α

B 1

0.74 0.72 0.70 0.68 0.66 0.64 0.62 0.60 0.58 0.56

AP-2α

D M

600 bp 500 bp 400 bp 300 bp

1 GAPDH AP-2α

F AP-2AGAPDH

β-actin

2 AP-2α β-actin

1.02 1.00 0.98 0.96 0.94 0.92 0.90 0.88 0.86 0.84 0.82

empty plasmid

AP-2α expression plasmid

empty plasmid

AP-2α expression plasmid

Fig. 7. E fficiency of interference in siRNA fragments. After we determined the transfection efficiency of the fluorescent cells to be > 70%, we transfected HepG2 cells with siRNAs against AP-2α fragments and tested the ability of these fragments to interfere with AP-2α mRNA and protein production (A, B). Interference efficiency was calculated using the Tanon GIS image analysis system. The interference efficiencies of fragments 1, 2, and 3 on the AP-2α mRNA levels were 86.06, 83.68 and 92.43%, respectively, and on the AP-2α protein levels were 61.15, 58.42 and 79.59%, respectively. Because the interference efficiency of fragment 3 was the highest, we selected this fragment for use in the subsequent experiments. Interference and overexpression of AP-2α in HepG2 cells. RT-PCR (A) and Western blot analyses (B) of AP-2α mRNA and protein expression levels, respectively, in HepG2 cells treated with three interference fragments for 72 hours. AP-2α overexpression in HepG2 cells resulted in elevated AP-2α RNA (C), and protein expression levels (D), compared to controls. AP-2α mRNA vs GAPDH mRNA: 0.63 ± 0.01 vs 0.71 ± 0.02 (E); AP-2α protein vs β-actin protein: 0.89 ± 0.03 vs 0.99 ± 0.01 (F), both p < 0.05 by paired t-test.

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Discussion The results of this study confirm the previously identified18,19 association between CHD and SNPs T-855C and T-778C in the promoter region of the ApoM gene. Luciferase activity associated with the -855 T→C substitution was significantly less than that of the promoter with -855 TT. With software predictive analysis we found the possible reason for this finding was that the -855 T→C substitution permitted the TFs AP-2α and Sp1 to bind to the promoter. When HepG2 cells were transfected with the ApoM promoter containing the -855 T→C substitution, AP-2α combined with the ApoM-855 area, thereby decreasing promoter activity. These findings confirm that changes in the activation of the ApoM promoter region may induce variations in the ApoM plasma concentration. Our results suggest that C alleles at the ApoM promoter -855 and -778 were associated with increased CHD risk. In our population-based case–control study, we enrolled 88 CHD patients (63 males, mean age: 60.80 ± 9.27 years) and 88 unrelated individuals (53 males, mean age: 58.18 ± 10.43 years) as a control group. The CHD group was divided into ACS and SAP groups, and the plasma levels of TG, TC, HDL-C, FPG and LDL-C were evaluated. Genomic DNA from whole blood of these subjects was subjected to PCR amplification and restriction enzyme digestion to determine genotype with regard to the ApoM T-855C and T-778C polymorphisms. CHD patients had higher TG (1.97 ± 1.28 mmol/l; p = 0.000) and FPG levels (6.40 ± 2.40 mmol/l; p = 0.000), and lower HDL-C levels (1.05 ± 0.25 mmol/l; p = 0.000) than non-CHD patients. The allelic frequencies were in Hardy–Weinberg equilibrium. After adjustment for age, gender and serum glucose level, multiple logistic regression analysis showed that, compared to the wild-type TT genotype of the two SNPs, carriers of the C allele had an increased risk of CHD, with an odds ratio (OR) of 1.819, 95% confidence interval (CI) of 1.142–2.898, and p = 0.012 (T-855C: OR = 3.206, 95% CI = 1.139–2.204, p = 0.037; T-778C: OR = 3.290, 95% CI = 1.487–7.280, p = 0.004). Luciferase activities of the promoter constructs with CC were significantly lower than those of the constructs with TC and TT. To detect whether different alleles of the ApoM proximal promoter region may affect the expression of target genes, thereby affecting the metabolism of ApoM, we constructed different genotypes of the promoter reporter gene to examine how mutations in the ApoM proximal promoter would affect promoter activity. The presence of a C allele at -855 or -778 bp of the ApoM promoter region may lead to lower ApoM levels and allow prediction of disease severity in the patient. However, due to the small number of cases analysed, more clinical data are needed for this conclusion to be confirmed. We investigated whether the DNA sequence of ApoM from -844 to -869 bp was involved in transcriptional regulation of the ApoM gene. Using EMSA experiments, we showed that the mutant allele (-855C) could bind with nuclear proteins, whereas the wild-type allele (-855T) could not. Competitive inhibition experiments showed that the combination was due to specific binding by the TF AP-2α. To explore the role of AP-2α in ApoM promoter activity, we examined the luciferase activities of the wild-type and mutant-type alleles after interference of AP-2α. Whereas AP-2α interference increased the luciferase activities of the treated cells,

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the wild-type was elevated to a lesser extent than the mutant-type. There were other AP-2α binding sites in addition to the -855 site. These results suggest that AP-2α may be a negative regulatory factor of ApoM. The increased luciferase activity of the mutant type with Apo-2α interference compared to the wild-type may indicate that the mutant had more binding sites for AP-2α, or that the mutated -855 site can bind with AP-2α. Multiple epidemiological studies have shown that serum HDL levels are negatively correlated with the risk of early CHD.20 Generally, clinical CAD is divided into two major types, ACS and SAP. Patients with ACS had significantly lower ApoM levels, probably due to the fact that ApoM is a major apolipoprotein of HDL. It has been confirmed that ApoM is required for pre-βHDL formation and cholesterol efflux to HDL, and that it protects against atherosclerosis.4 ApoM increased formation of pre-β-HDL particles and had a profoundly protective effect on atherosclerotic lesion formation in hypercholesterolaemic Ldlr-/mice.4 Atherosclerotic lesion areas in aortic roots and the thoracic aorta were reduced in Ldlr-/- mice infected with Ad-ApoM.4 The unstable lesion (also vulnerable plaque, the formation being mainly due to dyslipidaemia) is the basic pathological aetiological factor of ACS. Therefore the presence of an enlarged unstable lesion may be because the decreased serum ApoM level prohibited the formation of sufficient amounts of mature, functional HDL to promote the mobilisation of cellular cholesterol in vivo.21 The serum glucose level was different between CHD patients and normal controls, and serum ApoM and serum glucose levels were negatively correlated (both p < 0.05). Very low ApoM levels increase the risk of atherosclerosis.22 Therefore the serum ApoM level may be a valuable marker for identifying high-risk groups. TFs are the most important regulators of protein expression by genes and the most important factors to influence ApoM expression. Gene promoter regulation may underlie the low ApoM levels in CHD patients. Recent studies have shown that several TFs participate in the regulation of ApoM expression, such as HNF-1α, liver receptor homolog-1 (LRH-1), forkhead box A2 (Foxa2),23,24 liver X-activated receptor (LXR),25,26 leptin,27 interleukin-1 (IL-1),28 transforming growth factor (TGF) and epidermal growth factor (EGF).9 Our study has some limitations. Although our results confirm findings on the effect of the T-778C polymorphism on CHD, our analysis of the association of the ApoM promoter region SNPs with CHD was limited to a single locus. Such single-locus associations may be different in different populations. We found that the ApoM plasma concentration was decreased in CHD patients, the rs805296 and rs9404941 SNPs were associated with CHD occurrence and severity, and the rs9404941 SNP was associated with plasma TC and TG changes. However, the small sample size of this study limits its statistical power, and the results should be replicated in studies with larger sample sizes to avoid false positives. Expression of the ApoM protein and its relationship with diseases need to be further studied and discussed.

Conclusion ApoM may be a biomarker of CAD. ApoM-855 T→C substitution provides binding sites for AP-2α and reduces ApoM transcription activity.

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This work was supported by the Natural Science Foundation of Anhui (No

237

Gene 2003; 321: 93–102.

090413111, 1508085MH149) and the Wuhu municipal scientific research

15. Lu G. The role of AP-2alpha in atherosclerosis and HeLa apopto-

projects (No 2013 cxy04). The experimental research work was completed at

sis. Hunan: Biochemistry and molecular biology of Hunan Normal

the Clinical Testing and Diagnosis Experimental Centre of Bengbu Medical College and the key laboratory of bioactive macromolecules of Wangnan Medical College in Anhui province.

University Teaching and Research Section, 2008: 70–74. 16. Zhou JW, Tsui SK, Ng MC, Geng H, Li SK, So WY, et al. Apolipoprotein M gene (APOM) polymorphism modifies metabolic and disease traits in type 2 diabetes. PLoS One 2011; 6: e17324.

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Xu N, Dahlback B. A novel human apolipoprotein (apoM). J Biol Chem 1999; 274: 31286–31290.

variant of apolipoprotein M increases susceptibility to coronary artery

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Christoffersen C, Obinata H, Kumaraswamy SB, Galvani S, Ahnström

546–551.

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19. Jiao GQ, Yuan ZX, Xue YS, Yang CJ, Lu CB, Lu ZQ, et al. A prospec-

provided by HDL-associated apolipoprotein M. Proc Natl Acad Sci U

tive evaluation of apolipoprotein M gene T-778C polymorphism in

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relation to coronary artery disease in Han Chinese. Clin Biol Chem 2007;

Wolfrum C, Poy MN, Stoffel M. Apolipoprotein M is required for

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Christoffersen C, Nielsen LB, Axler O, Andersson A, Johnsen AH, M-containing lipoproteins. J Lipid Res 2006; 47: 1833–1843. Christoffersen C, Nielsen LB. Apolipoprotein M – a new biomarker in sepsis. Crit Care 2012; 16: 126.

density lipoprotein as a protective factor against coronary heart disease. The Framingham Study. Am J Med 1977; 62: 707–714. 21. Elsøe S, Christoffersen C, Luchoomun J, Turner S, Nielsen LB. Apolipoprotein M promotes mobilization of cellular cholesterol in vivo. Biochim Biophys Acta 2013; 1831: 1287–1292. 22. Faber K, Hvidberg V, Moestrup SK, Dahlbäck B, Nielsen LB. Megalin is a receptor for apolipoprotein M, and kidney-specific megalin-defi-

lipoprotein metabolism and atherosclerosis. Curr Opin Lipidol 2006; 17:

ciency confers urinary excretion of apolipoprotein M. Mol Endocrinol 2006; 20: 212–218.

Feingold KR, Shigenaga JK, Chui LG, Moser A, Khovidhunkit W,

23. Lee CS, Friedman JR, Fulmer JT, Kaestner KH. The initiation of liver

Grunfeld C. Infection and inﬂammation decrease apolipoprotein M

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20. Gordon T, Castelli WP, Hjortland MC, Kannel WB, Dawber TR. High

Bjrn D, Nielsen LB. Apolipoprotein M: a novel player in high density 291–295.

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density lipoprotein metabolism and atherosclerosis. J Curr Opin Lipidol

Dahlbäck B. Isolation and characterization of human apolipoprotein

confer the susceptibility to the development of type 1 diabetes. Clin Biol 18. Xu WW, Zhang Y, Tang YB, Xu YL, Zhu HZ, Ferro A, et al. A genetic

against atherosclerosis. Nature Med 2005; 11: 418–422.

Apolipoprotein M promoter polymorphisms alter promoter activity and

Dahlback B, Nielsen LB. Apolipoprotein M – a novel player in high-

pre beta-HDL formation and cholesterol efflux to HDL and protects 5.

17. Wu X, Niu N, Brismar K, Zhu X, Wang X, Efendic S, et al.

435: 944–947.

Xu N, Hurtig M, Zhang XY, Ye Q, Nilsson-Ehle P. Transforming growth

24. Wolfrum C, Howell JJ, Ndungo E, Stoffel M. Foxa2 activity increases

factor-beta downregulates apolipoprotein M in HepG2 cells. Biochim

plasma high density lipoprotein levels by regulating apolipoprotein M. J

Biophys Acta 2004; 1683: 33–37. 10. Han M, Zhong YL. Transcription factor AP-2 and its Roles in Cancer. Acta Laser Biol Sinica 2004; 13: 462–464. 11. Huang QC, Cao JH. The role of activating protein-2 transcription factors in breast cancer. Basic Clinical Med 2012; 32: 841–843. 12. Hilger-Eversheim K, Moser M, Schorle H, Buettner R. Regulatory roles of AP-2 transcription factors in vertebrate development, apoptosis and cell cycle control. Gene 2000; 260: 1–12. 13. Zhao F, Satoda M, Licht JD, Hayashizaki Y, Gelb BD. Cloning and characterization of a novel mouse AP-2 transcription factor, AP-2delta, with unique DNA binding and transactivation properties. J Biol Chem 2001; 276: 40755–40760. 14. Tummala R, Romano RA, Fuchs E, Sinha S. Molecular cloning and characterization of AP-2 epsilon, a fifth member of the AP-2 family.

Biol Chem 2008; 283: 16940–16949. 25. Zhang X, Zhu Z, Luo G, Zheng L, Nilsson-Ehle P, Xu N. Liver X receptor agonist downregulates hepatic apoM expression in vivo and in vitro. Biochem Biophys Res Commun 2008; 371: 114–117. 26. Calayir E, Becker TM, Kratzer A, Ebner B, Panzenböck U, Stefujl J, et al. LXR-agonists regulate ApoM expression differentially in liver and intestine. Curr Pharm Biotech 2008; 9: 516–521. 27. Xu N, Nilsson-Ehle P, Hurtig M, Ahrén B. Both leptin and leptinreceptor are essential for apolipoprotein M expression in vivo. Biochem Biophys Res Commun 2004; 321: 916–921. 28. Wolfrum C, Asilmaz E, Luca E, Friedman JM, Stoffel M. Foxa2 regulates lipid metabolism and ketogenesis in the liver during fasting and in diabetes. Nature 2004; 432: 102710–102732.

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Clinical features and patency rates of Remedy® biodegradable peripheral stents Selma Kenar Tiryakioglu, Osman Tiryakioglu, Oguz Karahan, Sinan Demirtas, Fatih Gokalp, Kamuran Erkoc, Hakan Özkan, Ahmet Ozyazicioglu

Objective: The aim of this study was to investigate the mid-term results of Remedy® biodegradable stents, which have recently come into use for lower-extremity arterial occlusive disease. Methods: Sixty-five patients, who underwent surgical intervention in various cardiovascular surgery clinics throughout Turkey, were included in the study. The total number of stents used was 92. The mean age of the patients was 64.11 ± 24.13 years (20–82), and 16 (24.6%) were female. The mean number of stents per patient was 1.42, and 70.7% of the lesions were TASC type A. Patients were followed for a mean of 32 months. Sixty-five patients underwent a control examination using either digital subtraction angiography or colour Doppler ultrasonography. In-stent restenosis was defined as ≥ 50% stenosis in the stent area in asymptomatic patients. The procedure was repeated if the degree of stenosis was ≥ 70%. Results: During the follow-up period, restenosis (≥ 50% stenosis) was observed in seven patients (10.7%). The patency rate after secondary intervention was 100%, and there was no loss of limbs in any patient. Restenosis was observed in six patients with superficial femoral artery stents, and in one patient with a popliteal arterial stent. Conclusion: Our experience shows that Remedy® biodegradable peripheral stents were safe and effective in our cohort of patients, with acceptable patency rates.

Submitted 17/9/15, accepted 8/1/16 Published online 26/1/16 Cardiovasc J Afr 2016; 27: 238–241

www.cvja.co.za

DOI: 10.5830/CVJA-2016-002

The clinical use of stents for the treatment of peripheral artery disease is favoured for both long-term primary and secondary patencies, as well as shorter hospitalisation periods.1 The major stent types include bare-metal, nitinol, drug-eluting (for example, Sirolimus), ePTFE + nitinol (Viabahn) and biodegradable stents.2 Biodegradable stents contain the biologically degradable PLLA (poly-L-lactic acid) polymer. A few years after their implantation, biodegradable stents dissolve into water and carbon dioxide, and are absorbed by the arterial tissue.3-5 The development of restenosis in the stented segment does not limit the use of other procedures, and it is suitable for re-ballooning.6 In addition, when compared to metal stents, PLLA stents are more suitable as a platform for drug-releasing stents.7,8 This study investigated the mid-term results of Remedy® biodegradable stents.

Methods Keywords: biodegradable stent, peripheral vascular disease, clinical outcomes, patency rates, restenosis Department of Cardiology, Bursa State Hospital, Bursa, Turkey Selma Kenar Tiryakioglu, MD

Department of Cardiovascular Surgery, Bahçeşehir University Medical Faculty, Istanbul, Turkey Osman Tiryakioglu, MD, tiryaki64@hotmail.com

Department of Cardiovascular Surgery, Dicle University Medical Faculty, Diyarbakir, Turkey Oguz Karahan, MD Sinan Demirtas, MD

Department of Cardiovascular Surgery, Şişli Etfal Education and Research Hospital, Istanbul, Turkey Fatih Gokalp, MD

Department of Cardiovascular Surgery, Medicalpark Bursa Hospital, Bursa, Turkey Kamuran Erkoc, MD

Department of Cardiology, Bahçeşehir University Medical Faculty, Istanbul, Turkey Hakan Özkan, MD

Department of Cardiovascular Surgery, Bursa Yüksek Ihtisas Education and Research Hospital, Bursa, Turkey Ahmet Ozyazicioglu, MD

Sixty-five symptomatic patients were included in this study during a mean 32-month period between January 2010 and December 2014. The mean age of patients was 64.11 years (20–82). Biodegradable peripheral stents (Remedy®, Kyoto Medical, Japan) were used. The stent-application procedure was performed in symptomatic patients who had ≥ 70% stenosis of the native artery diameter. An informed consent form was obtained from all patients prior to the procedure. Additionally, ethical approval was obtained from the local ethics committee of the University (2014-160). An exclusive stent-application team was used to place the peripheral stents into appropriate patients (according to diameter of vessel and length of lesion) in different cities. Only selected lesions were studied due to the limited diameter (5–8 mm) and length (36 mm) of the stents. Patients excluded from the study were those with long segments of stenosis, upper-extremity arterial interventions, large-diameter lesions, and if more than one stent was required. TASC type C and D patients who had had previous surgery and had appropriate vessel diameters and lesion lengths were included in the study. All patients were included in the study prospectively. In addition, some of the cases were re-evaluated retrospectively (especially patients with stent re-stenosis). The peripheral arterial lesions of patients were classified in accordance with the TASC classification scheme.9 All procedures

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were performed under spinal or local anaesthesia. The anaesthesia protocol was selected according to patient compliance. During the procedure, anticoagulation was provided by the administration of 100–300 U/kg of heparin. Following the procedure, 150 mg of clopidogrel (Plavix®, Sanofi, Turkey) was used as the anti-aggregant treatment protocol for patients who did not have any contraindications, and the treatment was subsequently continued with a dose of 75 mg/day. The patients were monitored post-operatively in the first week, and second and sixth months. After one year, there were no significant clinical symptoms in any of the 43 patients. The mean follow-up period was 32 months. Digital subtraction angiography (DSA) was used to examine nine randomly selected patients, whereas colour Doppler ultrasonography (USG) was used to examine 52 randomly selected patients. Four symptomatic patients were evaluated separately.

Statistical analysis For analysis of the demographic, pre-operative, operative and postoperative data, and comparison of the median ± t standard deviation results of the group, categorical data chi-squared tests were performed using the Statistical Package for the Social Sciences (SPSS Inc, Chicago, Illinois, USA) 17.0 statistics software; p < 0.05 was considered significant.

Results Twenty-seven patients had diabetes, 38 had hyperlipidaemia, and seven had chronic obstructive pulmonary disease. Fifteen patients underwent coronary bypass surgery or were diagnosed with coronary artery disease (Table 1). Sixty-five patients were examined with DSA or colour Doppler USG. DSA was performed in all symptomatic patients, and Doppler USG was performed in all patients. In-stent restenosis was defined as ≥ 50% stenosis in the stent area in asymptomatic patients. The procedure was repeated if the degree of stenosis was ≥ 70%. None of the nine asymptomatic patients who were examined with DSA had ≥ 50% lesions. Three patients who were examined with colour Doppler USG had 50–70% lesions (two patients had a superficial femoral artery lesion; one had a popliteal arterial stent). None of the patients had ≥ 70% lesions. In total, 46 patients had type A lesions, 12 had type B lesions, five had type C lesions, and two had type D lesions, according to the TASC classification scheme.9 Sixteen out of 65 patients (24.6%) were female and 49 (75.4%) were male (Table 1). The total number of stents used was 92, and the mean number of stents per patient was 1.42.1-3 The mean stent length was 53 mm per patient (36–108 mm). Twenty-two stents (24%) were implanted into the right superficial femoral artery, 31 (33.7%) into the left superficial femoral artery, 14 (15.2%) into the left iliac artery, 17 (18.4%) into the right iliac artery, five (7.8%) into the left popliteal artery, and three (3.2%) into the right popliteal artery. The size of the stents ranged between 5 and 8 mm; the most commonly used stent width was 6 mm. The procedure was performed under spinal anaesthesia in five patients (7.7%) and local anaesthesia in 60 patients (92.3%) (Table 2). There was 100% success rate of the procedure. In all patients, it was possible to palpate the pulse with a hand on the

Table 1. Laboratory data and vascular pathology of patients. Mean age ± SD = 64.11 ± 24.13 years n

49/16

75.4/24.6

TASC type A lesion

70.7

TASC type B lesion

20.0

TASC type C lesion

8.3

TASC type D lesion

3.3

Diabetes mellitus

41.5

Hyperlipidaemia (LDL > 130 mg/dl or 3.37 mmol/l)

58.4

Parameters Gender (M/F)

COPD Coronary artery disease (diagnosed)

10.7

TASC: TransAtlantic Inter-Society Consensus; LDL: low-density lipoprotein cholesterol; COPD: chronic obstructive pulmonary disease.

distal part of the stent zone after the procedure. There was no limb loss in any patient. The mean follow-up period was 32 months (18–48 months), and in-stent restenosis developed in three patients. These patients received a secondary balloon dilatation procedure. The superficial femoral artery stent was fully occluded in one patient, and the patient underwent femoropoliteal bypass. The remaining patients have had no complications from the stenting. During the follow-up period, DSA was performed in all patients who were admitted symptomatically (four patients). Total occlusion was observed in one patient and 70–90% stenosis was observed in three patients (all patients had a superficial femoral artery stent) (Table 3). All patients were alive, so the Kaplan–Meier life curve was not included in the text, and we used single-group analysis.

Discussion This was a unique study demonstrating the results of mid-term peripheral application of biodegradable stents. The findings of this study show that biodegradable stents had successful clinical outcomes during an average of 32 months’ follow-up period. It appears from this study that biodegradable stents have good mid-term patency rates in peripheral arterial occlusions. In patients with limb ischaemia, the first-line approach for revascularisation has shifted over the past decade from bypass surgery to endovascular intervention.10 Stenting for the treatment of lower-extremity arterial occlusive disease is an important tool and continues to evolve, with new stent designs and technologies being developed to provide superior patency rates and limb salvage.11,12 Table 2. Stent features and sites for stent therapy in patients Stents Total number of stents (n) Mean stent diameter (mean ± SD mm) Stent length per patient (mean ± SD mm) Restenosis (n/%)

92 6.9 ± 1.3 53.0 ± 14.6 7/7.6

Application area (n/%) Superficial femoral artery

53/57.6

Iliac artery

31/33.7

Popliteal artery

8/8.7

Anaesthesia (n/%) Local

60/92.3

Spinal

5/7.7

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Table 3. Clinical imaging and severity of stenosis according to symptoms < 50% stenosis

50–70% stenosis

≥ 70% stenosis 4*

Symptomatic patients: DSA (n = 4)

–

Asymptomatic patients: DSA (n = 9)

–

Asymptomatic patients: USG (n = 52)

3**

–

Total

*All were superficial femoral artery (SFA) lesions. **Two patients had SFA stents; one had a popliteal artery stent. USG: ultrasonography; DSA: digital subtraction angiography.

Short- and long-term outcomes of peripheral artery stents are available for metal, nitinol and e-PTFE-coated nitinol stents. Initially, efforts were directed at overcoming the negative outcomes of metal stents, especially in long lesions.13 Owing to the recent advances in stent technology, biodegradable stents have been produced to avoid the undesirable outcomes of classic stents, which lead to mechanical restenosis with stent fracture. This technology has been extensively evaluated in the coronary artery system, but not in peripheral arteries. The outcome of bio-absorbable stents remains unclear.14 We therefore investigated the patency rates of this technology in the peripheral arterial system. Outcomes of even classical stents have not been sufficiently documented. Short hospitalisation times, successful outcomes in stent patency, higher secondary patency rates, and comparable outcomes with grafts have increased the popularity of peripheral stent applications in TASC type A and B lesions. The primary and secondary patency rates of PTFE-coated stents in veins above the knee have equal efficacy to PTFE grafts, and are comparable with that of saphenous veins. Formation of the neo-intima layer in closed stents prevents the development of restenosis in the early period, and ensures that stents remain patent for a longer period.15,16 The use of stents for suitable lesions, especially in the case of iliac artery lesions, protects the patient from possible abdominal surgery and increases the incidence of long-term patency as the vein diameter is also suitable. In the present study, stenosis was not observed during the follow-up period after iliac artery stent implantation. The PLLA peripheral stent is the first fully bioresorbable stent to be implanted in humans, with complete degradation taking 12 to 24 months. This resulted in less vessel wall injury during implantation and therefore less initial thrombus formation and reduced intimal hyperplasia. The existing literature on the use of biodegradable peripheral stents on coronary arteries is limited, and their superiority over drug-releasing stents has not been proven to date. The existing data regarding the use of biodegradable peripheral stents in peripheral arteries is insufficient.17 The application of biodegradable peripheral stents has positive contributions to the collateral system in particular. The collateral system in the application area does not close, but rather increases in the long term, and supports development of the existing systems.18 Collateral closure was not observed during the procedure in our patient group, and we also observed that these veins stayed open in the patient group examined with DSA. The major advantages of using biodegradable stents include practical application, a suitable structure for secondary interventions, their absorbable and non-depositing nature,

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perfect tissue compatibility, and a lower rate of restenosis in experimental studies. A feature of the biodegradable stent is that it allows for re-intervention due to its structure (after implantation of the stent, integration occurs between the stent structure and vessel wall). Therefore re-intervention can easily be carried out (balloon and stent or surgery) in TASC type A lesions when they become TASC type C or D. Another advantage is that there is no possibility of stent fracture, which eliminates anti-aggregant use. They are also suitable for use in patients with a metal allergy. There are certain disadvantages, including insufficient number of clinical studies investigating long-term outcomes. Moreover, the length and diameter of the available stents are not suited for every clinical scenario. The absence of a control group was the main limitation of this study. The ethics committee approved the study protocol, which was created with the use of a single type of stent to avoid bias and to provide similar opportunities for all patients. This pilot study was therefore conducted without a control group.

Conclusion In this study, primary implantation of a Remedy® biodegradable stent for moderate-length lesions in lower-extremity arterial occlusive segments of patients with claudication was associated with better mid-term results. Our experience shows that the Remedy® peripheral stents were safe and effective in our cohort of patients, with acceptable patency rates.

References 1.

Schillinger M, Erich Minar E. Past, present and future of femoropopliteal stenting. J Endovasc Ther 2009; 16: I147–I152.

Ansel GM, Lumsden AB. Evolving modalities for femoropopliteal interventions. J Endovasc Ther 2009; 16: II82–II97.

Onuma Y, Serruys PW. Bioresorbable scaffold: the advent of a new era in percutaneous coronary and peripheral revascularization? Circulation 2011; 123: 779–797.

Waksman R, Pakala R. Biodegradable and bioabsorbable stents. Curr Pharm Des 2010; 16: 4041–4051.

Biamino G, Schmidt A, Scheinert D. Treatment of SFA lesions with PLLA biodegradable stents: results of the PERSEUS study. J Endovasc Ther 2005; 12: 5.

Garg S, Serruys P. Biodegradable stents and non-biodegradable stents. Minerva Cardioangiol 2009; 57: 537–565.

Macaya C, Moreno R. Bioabsorbable drug-eluting stents: the future of coronary angioplasty? Nat Clin Pract Cardiovasc Med 2008; 5: 598–599. doi: 10.1038/ncpcardio1306.

Wykrzykowska JJ, Onuma Y, Serruys PW. Advances in stent drug delivery: the future is in bioabsorbable stents. Expert Opin Drug Deliv 2009; 6: 113–126. doi: 10.1517/17425240802668495.

Norgren L, Hiatt WR, Dormandy JA, Nehler MR, Harris KA, Fowkes FG; TASC II Working Group. TASC II Working Group Inter-Society Consensus for the Management of Peripheral Arterial Disease (TASC II). J Vasc Surg 2007; 45: 5–67.

10. Biancari F, Kantonen I, Albäck A, Mätzke S, Luther M, Lepäntalo M. Limits of infrapopliteal bypass surgery for critical leg ischemia: when not to reconstruct. World J Surg 2000; 24: 727–733. 11. Scheinert D, Katsanos K, Zeller T, Koppensteiner R, Commeau P, Bosiers M, et al. ACHILLES Investigators. A prospective randomized

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multicenter comparison of balloon angioplasty and infrapopliteal stent-

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carrev.2015.08.005.

ing with the sirolimus-eluting stent in patients with ischemic peripheral

15. Schneider JR, Verta MJ, Alonzo MJ, Hahn D, Patel NH, Kim S. Results

arterial disease: 1-year results from the ACHILLES trial. J Am Coll

with Viabahn-assisted subintimal recanalization for TASC C and TASC

Cardiol 2012; 60: 2290–2295. doi: 10.1016/j.jacc.2012.08.989.

D superficial femoral artery occlusive disease. Vasc Endovascular Surg

12. El-Sayed HF. Endovascular techniques in limb salvage: stents. Methodist Debakey Cardiovasc J 2013; 9: 79–83. 13. Krankenberg H, Schlüter M, Steinkamp HJ, Bürgelin K, Scheinert

2011; 45: 391–397. 16. Saxon RR, Coffman JM, Gooding JM, Ponec DJ. Long-term patency and clinical outcome of the Viabahn stent-graft for femoropopliteal

D, Schulte KL, et al. Nitinol stent implantation versus percutaneous

artery obstructions. J Vasc Interv Radiol 2007; 18: 1341–1349; quiz 1350.

transluminal angioplasty in superficial femoral artery lesions up to 10

17. Sammel AM, Chen D, Jepson N. New generation coronary stent tech-

cm in length: the femoral artery stenting trial (FAST). Circulation 2007;

nology – is the future biodegradable? Heart Lung Circ 2013; 22: 495-506.

116: 285–292.

doi: 10.1016/j.hlc.2013.02.008.

14. Galyfos G, Geropapas G, Stefanidis I, Kerasidis S, Stamatatos I,

18. Vermassen F, Bouckenooghe I, Moreels N, Goverde P, Schroe H. Role

Kastrisios G, et al. Bioabsorbable stenting in peripheral artery disease.

of bioresorbable stents in the superficial femoral artery. J Cardiovasc

Cardiovasc Revasc Med 2015; pii: S1553-8389(15)00200-6. doi: 10.1016/j.

Surg (Torino) 2013; 54: 225–234.

Should the findings of the DAPT trial change my practice? One of AfricaPCR 2016’s final sessions discussed this question and a distinguished panel concluded, albeit with a few reservations and caveats, that the answer is ‘yes’. DAPT was the first large trial to evaluate the benefits of dual antiplatelet therapy prolonged beyond 12 months after percutaneous coronary intervention (PCI). DAPT set out to investigate, as an effectiveness endpoint, whether prolonged dual antiplatelet therapy was associated with a reduction in stent thrombosis and in major cardiovascular and cerebrovascular events. The safety endpoint was its effect on moderate or severe bleeding. It found that prolonging therapy up to 30 months was associated with a 71% relative risk reduction in stent thrombosis and a 29% relative risk reduction in major events. But there was a price to pay in that there was a 1.6–2.5% increase in rate of bleeding. Prior to DAPT, previous practice had been to give therapy for much shorter periods, usually one to six months. Reviewing what was known before the study, Dr Colin Schamroth, from Johannesburg, noted that most previous studies had suggested that there was no solid evidence to support treatment beyond 12 months. TRITON-TIMI had hinted that there might be a case for prolonging therapy, but had still concluded that most benefit was accrued over 12 months. The 2012 European guidelines currently recommend dual antiplatelet therapy for a minimum of one month in patients with bare-metal stents and six months in those with drugeluting stents, with three to six months for the former and six to 12 months for the latter being the preferred strategy. Drugeluting stents are associated with higher rates of thrombosis. Twelve months of therapy are recommended for unstented STEMI patients.

Drilling down into the details of DAPT, Dr Riaz Dawood, also from Johannesburg, noted that it was a well-designed trial with results that were valid and not underpowered. It included nearly 10 000 patients from 11 countries. However, none of these was in Africa, South America or Asia. He also underscored that it was a low-risk population, excluding patients with a high risk of bleeding or ischaemic events. The 29% relative risk reduction in major events was driven mainly by a decrease in myocardial infarction (MI), with no significant differences observed in respect of mortality and haemorrhagic stroke. The increase in bleeding risk was driven by moderate bleeding, with severe, life-threatening bleeding not significant. All-cause mortality, however, showed a 36% increase, but this was not driven by either cardiovascular death or by bleeding, but rather by cancer. Dr Dawood felt that this was most likely a ‘play of chance’, unrelated to dual antiplatelet therapy. In conclusion, DAPT found that prolonging therapy up to 30 months was associated with a significant reduction in rates of thrombosis and major events, notably MI, but at a risk of increased rates of bleeding. The MI benefit was not limited to stented sites. The findings were also consistent, regardless of which thienopyridine agents were used. So should these findings change clinical practice? Prof Pravin Manga, from Johannesburg, offered a cautious ‘yes’. ‘The evidence suggests that longer and maybe even indefinite therapy may be of benefit, but we need to be very careful when it comes to patient selection.’ Dr Dawood echoed this. ‘Bleeding worries us more than ischaemic events, and bleeding risk increases with age, so one needs to be especially cautious in elderly patients. But in patients without contra-indications who are at risk of thrombosis, but with a low bleeding risk, why not continue?’ Source: AfricaPCR 2016

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Humoral immune response and coated or uncoated oxygenators during cardiopulmonary bypass surgery Sedat Ozan Karakisi, Ayşe Gül Kunt, Şahin Bozok, İdil Çankaya, Mustafa Kocakulak, Uğur Muşabak, Mustafa Fevzi Sargon, Şaban Ergene, Gökhan İlhan, Hakan Karamustafa, NebiyeTufekci, Erol Şener

Abstract Aim: To investigate and compare uncoated and phosphorylcholine-coated oxygenators in terms of induction of humoral immune response during coronary artery bypass surgery. Methods: A total of 20 consecutive patients who underwent coronary artery bypass surgery were randomly distributed into two groups according to the type of oxygenator used during surgery. Group 1 consisted of 10 patients who were operated on using phosphorylcholine-coated oxygenators. Group 2 contained 10 patients who underwent surgery using uncoated oxygenators. Blood and oxygenator fibre samples were obtained and compared in terms of immunoglobulins (IgG, IgM), complements (C3c, C4), serum total protein and albumin levels using electron microscopy and flow cytometry. Results: In group 1, levels of IgM, IgG, total protein and serum albumin were significantly increased at the end of cardiopulmonary bypass (CPB) compared to those at the beginning of CPB. In group 2, C3c and C4 levels at the beginning of CPB were found to be significantly higher than at the end. Electron microscopic examination of oxygenator fibres demonstrated that phosphorylcholine-coated fibres were less likely to be adsorbed by serum proteins and complements than the uncoated fibres.

Department of Cardiovascular Surgery, Faculty of Medicine, Recep Tayyip Erdogan University, Training and Research Hospital, Rize, Turkey Sedat Ozan Karakisi, MD Şahin Bozok MD, sahinboz@yahoo.com Şaban Ergene, MD Gökhan İlhan, MD Hakan Karamustafa, MD NebiyeTufekci, MD

Department of Cardiovascular Surgery, Faculty of Medicine, Yıldırım Beyazıd University, Atatürk Training and Research Hospital, Ankara, Turkey Ayşe Gül Kunt, MD Erol Şener, MD

Department of Biomedical Engineering, Baskent University, Ankara, Turkey İdil Çankaya Mustafa Kocakulak

Department of Immunology, Gulhane Military Medical Academy and School of Medicine, Ankara, Turkey Uğur Muşabak, MD

Department of Anatomy, Faculty of Medicine, Hacettepe University, Ankara, Turkey Mustafa Fevzi Sargon, MD

Conclusion: Our results indicate that phosphorylcholine-coated oxygenators seemed to induce humoral immune response to a lesser extent than uncoated oxygenators during coronary artery bypass procedures. Keywords: cardiopulmonary bypass, oxygenator, phosphorylcholine, humoral inflammation Submitted 5/10/15, accepted 8/1/16 Cardiovasc J Afr 2016; 27: 242–245

www.cvja.co.za

DOI: 10.5830/CVJA-2016-003

Cardiopulmonary bypass (CPB) facilitates surgical procedures and provides adequate perfusion of other organs during cardiovascular surgery.1,2 Despite the advantages offered by CPB, a systemic inflammatory response may arise due to multiple components of the immune system, including cellular and humoral components. This inflammation may arise from contact of circulating blood cells with non-endothelial surfaces of extracorporeal circulation, as well as from ischaemia/reperfusion injury, hypothermia and other operative stresses.1,2 Cardiopulmonary and systemic hazards may occur owing to the outcomes of this inflammatory response, leading to morbidity and mortality.1,3 Modalities to manage this inflammatory response include medical agents such as steroids, complement inhibitors, monoclonal antibodies and protease inhibitors. In addition to these medications, it has been suggested that lining the inner surfaces of extracorporeal circulation systems with a relatively inert material may provide suppression of the immune response.4 The membranes of oxygenators are important in this aspect since they are directly in contact with the blood. Hence, coating these membranes is thought to aid in decreasing the inflammatory response.4,5 The objective of this study was to compare phosphorylcholinecoated and uncoated oxygenators in terms of the humoral immune response triggered during cardiopulmonary bypass surgery.

Methods This randomised, cross-sectional clinical study was performed in the cardiovascular surgery department of a tertiary care centre. Approval was obtained by the local institutional review board (2010/12) and all patients gave written informed consent. A total of 20 consecutive patients scheduled for CPB surgery were included. During CPB, a phosphorylcholine-coated oxygenator was used in 10 patients, constituting group 1, while the uncoated oxygenator was used for the remaining 10 cases, making up group 2. Participants were allocated to the two study groups according to a computerised block-randomisation process in order to keep the number of participants in the different groups equal.

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Serum study

243

Samples were gathered from the oxygenators with a sterile scalpel after opening the hard, protective cover surrounding the oxygenator with a Dremel cutting burr (Widget Supply Inc, Albany, Oregon, USA). The samples were obtained in two

different sizes, containing 300 fibres (6 cm) and 50 fibres (1 cm). Ultrasonic washing was performed on the 6-cm samples for mechanical cleaning. The fibres were maintained in 50-ml tubes containing 35 ml isotonic saline. Liquid nitrogen was added to the fibres prior to transection and electron microscopy. Electron microscopy was performed with the FEI Quanta 200 FEG scanning electron microscope (SEM) (FEI Europe, Nanoport, Eindhoven, The Netherlands) under an acceleration voltage of 22 kV.2 Fixation of the 1-cm fibres with 2.5% glutaraldehyde solution for 24 hours was followed by irrigation with Sorensen’s phosphate buffer (SPB). The next fixation was done with 1% osmium tetroxide, and the fibres were irrigated again with SPB solution. Increasing concentrations (25, 50, 75 and 100%) of acetone were used for dehydration. The samples were transferred to Petri dishes and dried for six hours. After drying, the material was adhered to metallic plates of the SEM and coated with a mixture gold and palladium of 100-Å thickness using a Bio-Rad sputter apparatus (Bio-Rad Laboratories headquarters, Hercules, CA, USA). After keeping the samples in a dry medium for 24 hours, electron microscopy was performed with a Jeol SEM ASID-10 device (Jeol Ltd, Tokyo, Japan) under 80-kV acceleration voltage. Electron microscopic views of the coated and uncoated oxygenator fibres are shown in Figs 1 and 2. Adsorption of proteins on the fibres of the coated and uncoated oxygenator fibres can be seen in Figs 3 and 4.

Fig. 1. E lectron microscopic view of a phosphorylcholinecoated oxygenator fibre.

Fig. 3. Protein adsorption on the surface of a phosphorylcholine-coated oxygenator fibre.

Fig. 2. E lectron microscopic view of an uncoated oxygenator fibre.

Fig. 4. Protein adsorption on the surface of an uncoated oxygenator fibre.

Complements (C3c, C4), immunoglobulins (IgG, IgM) and proteins were analysed from blood samples. A total of 5 ml of venous blood was drawn from each patient and these samples were rapidly transferred to acid–citrate–dextrose Adenin (ACD A) tubes (Becton Dickinson, Meylan, Cedex, France). Monoclonal antibodies (20 µl) of IgG1FITC/IgG1PE/ PerCP were added to each tube containing 1 × 106 cells. Erythrocytes were separated and removed with the addition of 2–3 ml of lysing solution (Becton Dickinson, San Jose, USA) after incubation in the dark at room temperature for 20 minutes. Subsequent to the lysing solution, the samples were irrigated with 2 ml of phosphate-buffered saline (PBS) and suspended in 500 µl PBS containing 1% paraformaldehyde. The samples were maintained at 2–8°C in the dark until analysis. Humoral analysis was done using the FACSCanto flow cytometry system and BD FACSDiva program (Becton Dickinson, Immunocytometry Systems, San Jose, CA 95131 USA).

Electron microscopy

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Statistical analysis The Statistical Package for Social Sciences program version 16.0 (SPSS Inc, Chicago, IL, USA) was used. Descriptive data were expressed as mean, standard deviation and range (minimum to maximum values). Numbers and percentiles were used for expression of categorical variables. Parametric tests were used for data with a normal distribution, and non-parametric tests were applied to data without a normal distribution. Distribution of normality was tested with the Kolmogorov– Smirnov test. The Mann–Whitney U- and Wilcoxon tests were used for comparing variables between groups. Chi-squared, Fisher’s and Mantel Haenszel tests were performed for comparison of categorical variables. Level of significance was set at p < 0.05.

Results The study group consisted of a total of 20 patients (3 females, 17 males) with average ages of 61.7 ± 13.2 years (range, 44–78) and 63.1 ± 9.6 years (range, 51–78), for groups 1 and 2, respectively. The descriptive data and peri-operative characteristics are shown in Table 1. The average values for serum albumin, total protein, C3c and C4 levels were higher in group 1 at the start of the pump (Table 2). No difference was observed between the groups in terms of these variables at the end of CPB (Table 2). In group 1, total protein levels were significantly higher at the start of the pump

Table 1. Descriptive and peri-operative parameters of the patients in groups 1 and 2 Group 1 (n = 10)

Group 2 (n = 10)

Age (years), median (range)

61.7 ± 13.25 (44–78)

63.1 ± 9.64 (51–78)

0.677

Gender (M/F), n (%)

8/2 (80/20)

9/1 (90/10)

0.531

2/8 (20/80)

3/7 (30/70)

1.000

Parameters

compared to at the end of CPB (p = 0.01). A significantly lower IgG level was noted at the end of CPB compared to at the start in group 1 (p = 0.012) (Table 2). IgM levels at the end of CPB were higher than IgM levels at the start in group 1 (p = 0.012). In group 1, mean levels of C3c were lower at the end of CPB compared to those at the start (p = 0.005) (Table 2).

Discussion In this study, we attempted to demonstrate whether there was a difference between the cellular immune responses of patients who underwent CPB using uncoated or phosphorylcholinecoated oxygenators. Our study indicated that a more prominent cellular immune response was observed in patients operated on using phosphorylcholine-coated oxygenators. In spite of the advantages it offers during cardiac surgery, CPB has the potential to cause a complex inflammatory response, initiated by the contact of heparinised blood with non-endothelial surfaces.6 Other factors, such as bleeding, ischaemia–reperfusion injury and rejection reactions further contribute to augmentation of the immune response via secretion of vasoactive and cytotoxic cytokines, resulting in alleviation of this inflammatory cascade.2-5 Recently, modification of the surfaces that come into contact with extracorporeal circulating blood has become popular and phosphorylcholine is one of the materials used for this purpose.7 In the literature, there are several reports on the impact of coated oxygenators on the immune response. It was suggested that leukocyte levels increased and platelet levels decreased in both phosphorylcholine-coated and uncoated oxygenators with regard to baseline values during induction of anaesthesia.8,9

p-value

Risk factors Diabetes mellitus, n (%)

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Hypertension, n (%)

6/4 (60/40)

1.000

COPD, n (%)

1/9 (10/90)

2/8 (20/80)

1.000

CRF, n (%)

1/9 (10/90)

2/8 (20/80)

1.000

Smoking habit, n (%)

5/5 (50/50)

7/3 (70/30)

0.361

Ejection fraction (%), median (range)

53.00 ± 3.58 (30–65)

47.08 ± 10.30 (30–65)

0.193

Duration of cross clamp (min), median (range)

49.60 ± 13.67 (30–77)

58.70 ± 23.55 (40–113)

0.405

Duration of CPB (min), median (range)

88.40 ± 26.28 (58–134)

102.1 ± 25.58 (75–146)

0.162

Heparin (units/ml), median (range)

4.20 ± 0.42 (4–5)

5.50 ± 0.84 (4–7)

0.001

Protamine (mg), median (range)

4.60 ± 0.51 (4–5)

5.70 ± 1.05 (4–8)

0.008

Duration of intubation (h), median (range)

8.00 ± 2.90 (4–12)

9.4 ± 3.74 (4–18)

0.488

ICU stay, (h), median (range)

34.00 ± 11.50 (20–48)

38.00 ± 11.19 (24–48)

0.384

Hospitalisation (days), median (range)

6.60 ± 0.96 (6–9)

8.00 ± 1.88 (6–12)

0.039

Drainage (ml), median (range)

645.00 ± 319.24 (200–1300)

530.00 ± 182.87 (250–900)

0.447

Transfusion (units), median (range)

920.00 ± 454.11 (600–1800)

900.00 ± 391.57 (600–1800)

0.934

Table 2. Levels of total protein, serum albumin, IgG, IgM, and complements C3c and C4. Variable Total protein (mg/dl)

Albumin (g/dl)

Postoperative features

M: male; F: female; CRF: chronic renal failure; COPD: chronic obstructive pulmonary disease; CPB: cardiopulmonary bypass; ICU: intensive care unit.

IgG (g/l)

IgM (mg/l)

C3c (mg/dl)

C4 (mg/dl)

Time of measurement

Group 1 median (range)

Group 2 median (range)

p-value

Start of pump

3.36 ± 0.31 (2.9–3.8)

4.08 ± 0.57 (3.1–4.7)

0.009§

End of pump

3.82 ± 0.61 (3.1–4.8)*

4.16 ± 0.37 (3.7–4.6)

0.223

Start of pump

1.96 ± 0.08 (1.9–2.1)

2.14 ± 0.31 (1.6–2.4)

0.030§

End of pump

2.24 ± 0.28 (1.9–2.7)*

2.24 ± 0.23 (1.9–2.5)

0.758

Start of pump

6.06 ± 1.71 (3.5–7.8)

7.94 ± 3.55 (3.6–12.4)

0.172

End of pump

7.48 ± 1.96 (4.9–10.3)*

6.72 ± 2.94 (3.2–9.9)

0.448

Start of pump

0.48 ± 0.13 (0.43–0.72)

0.64 ± 0.27 (0.21–0.98)

0.095

End of pump

0.54 ± 0.14 (0.39–0.73)*

0.55 ± 0.17 (0.23–0.68)

0.798

Start of pump

0.58 ± 0.11 (0.4–0.7)

0.81 ± 0.19 (0.59–1.12)

0.010§

End of pump

0.69 ± 0.10 (0.54–0.82)*

0.73 ± 0.07 (0.61–0.79)

0.601

Start of pump

0.13 ± 0.06 (0.08–0.23)

0.16 ± 0.02 (0.13–0.19)

0.022§

End of pump

0.29 ± 0.31 (0.09–0.87)

0.18 ± 0.04 (0.12–0.24)

0.315

IgG: immunoglobulin G; IgM: immunoglobulin M; C3c: complement factor C3c; C4: complement c4. *Statistically significant difference within the same group; §statistically significant difference between the two groups.

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Similarly, Sohn et al. stated that there was a decrease in the postoperative platelet counts of patients who were operated on using a phosphorylcholine-coated oxygenator.10 Our results have shown that IgG and IgM levels were increased in group 1 at the end of CPB. This finding suggests that phosphorylcholine-coated oxygenators may induce humoral immunity. Lante et al. however found that both IgG and IgM concentrations were decreased after cardiac surgery.11 The complement system may be activated due to factors such as ischaemia, hypoxia, haemodilution or contact with foreign bodies.12 In our series, C3c levels were found to be significantly increased in group 1 at the end of CPB. Adsorption of complements to the uncoated surfaces of fibres may be an explanation for this difference. It must be remembered that even the simple circulation of blood in extracorporeal systems may lead to activation of complements.6 Similar to our results, de Somer et al. found no difference between uncoated and phosphorylcholine-coated systems with regard to C3 and C4 levels. The increase in C3 levels up to the first postoperative day in the phosphorylcholine-coated group was assumed to be associated with prevention of protein adsorption by the coating.9 Baksaas et al. found no difference with regard to levels of C3 and C4 between patients operated on using uncoated and bio-passive coated surfaces.13 Watanabe et al. reported that there was an increase in C3 levels in both groups in the postoperative period.14 In another study, comparison of phosphorylcholine- and heparin-coated oxygenators demonstrated a rise in C3 levels in both groups.15 Suhara et al. found thromboses on the surface of uncoated oxygenator fibres.16 Niimi et al. reported decreased adherence of platelets to the fibres in heparin-coated systems.17 However, no difference was detected with regard to protein adsorption in the same study.17 Gunaydin reported that less protein adsorption was observed on phosphorylcholine-coated oxygenators.18 The results of our study have shown that levels of albumin and total protein appeared significantly higher at the end of the operation. This increase was more in the uncoated group in our series, which is to be expected since proteins and albumin have a greater tendency to adhere to uncoated surfaces. Electron microscopy also exhibited a thicker protein layer on the surface of uncoated oxygenator fibres and this finding is in conjunction with the increased likelihood of adherence of immune system elements to uncoated surfaces. Some limitations of this study must be noted. First, our sample size was small and strict criteria for inclusion of patients were not adhered to. Moreover, the impact of metabolic, environmental, genetic, racial and geographic factors, which could have influenced the results, could not be completely controlled. Therefore, interpretations and extrapolations must be made with caution. However, we hope that the results of this study will pioneer further trials on this topic.

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Paparella D, Yau TM, Young E. Cardiopulmonary bypass induced inflammation: pathophysiology and treatment. An update. Eur J Cardiothorac Surg 2002; 21: 232–244.

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Karakisi S, Kunt A, Çankaya I, Kocakulak M, Bozok Ş, Muşabak U, et al. Do phosphorylcholine-coated and uncoated oxygenators differ in terms of elicitation of cellular immune response during cardiopulmonary bypass surgery? Perfusion 2015, Jan 6. (e-pub).

Chenoweth DE, Cooper SW, Hugli TE, et al. Complement activation during cardiopulmonary bypass: evidence for generation of C3a and C5a anaphylactoxins. N Engl J Med 1981; 304: 497–503.

Pappalardo F, Della Valle P, Crescenzi G, Corno C, Franco A, Torracca L, et al. Phosphorylcholine coating may limit thrombin formation during high-risk cardiac surgery: a randomized controlled trial. Ann Thorac Surg 2006; 81: 886–891.

Gunaydin S. Clinical significance of coated extracorporeal circuits: a review of novel technologies. Perfusion 2004; 19: S33–S41.

De Somer F, François K, van Oeveren W, Poelaert J, de Wolf D, Ebels T. Phosphorylcholine coating of extracorporeal circuits provides natural protection against blood activation by the material surface. Eur J Cardiothorac Surg 2000; 18: 602–606.

10. Sohn N, Marcoux J, Mycyk T, Krahn J, Meng QH. The impact of different biocompatible coated cardiopulmonary bypass circuits on inflammatory response and oxidative stress. Perfusion 2009; 24: 231–237. 11. Lante W, Franke A, Weinhold C, Markewitz A. Immunoglobulin levels and lymphocyte subsets following cardiac operations: further evidence for a T-helper cell shifting. Thorac Cardiovasc Surg 2005; 53: 16–22. 12. Moore FD, Warner KG, Assousa S, Valeri CR, Khuri SF. The effects of complement activation during cardiopulmonary bypass – the attenuation by hypothermia, heparin, and hemodilution. Ann Surg 1988; 208: 95–103. 13. Baksaas ST, Videm V, Fosse E, Karlsen H, Pedersen T, Mollnes TE, et al. In vitro evaluation of surface coatings for extracorporeal circulation. Perfusion 1999; 14: 11–19. 14. Watanabe H, Hayashi J, Ohzeki H, Moro H, Sugawra M, Eguchi S. Biocompatibility of a silicone-coated polypropylene hollow fiber oxygenator in an in vitro model. Ann Thorac Surg 1999; 67: 1315–1319. 15. Thiara AS, Andersen VY, Videm V, Mollnes TE, Svennevig K, Hoel TN, et al. Comparable biocompatibility of phisio- and bioline-coated cardiopulmonary bypass circuits indicated by the inflammtory response. Perfusion 2010; 25: 9–16. 16. Suhara H, Sawa Y, Nishimura M, Oshiyama H, Eng B, Yokoyama K, et al. Efficacy of a new coating material, PMEA, for cardiopulmonary bypass circuits in a porcine model. Ann Thorac Surg 2001; 71:

Conclusion Despite the fact that phosphorylcholine-coated oxygenators were developed to decrease the immune response during coronary artery bypass surgery, our results have shown that a notable humoral immune response still exists with the use of these materials.

1603–1608. 17. Niimi Y, Ichinose F, Ishiguro Y, Terui K, Uezono S, Morita S, at al. The effects of heparin coating of oxygenator fibres on platelet adhesion and protein adsorption. Anesth Analg 1999; 89: 573–579. 18. Gunaydın S. Emerging technologies in biocompatible surface modifying activities: quest for physiologic cardiopulmonary bypass. Curr Med Chem Cardiovasc Hematol Agents 2004; 2: 187–196.

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High aortic pulse-wave velocity may be responsible for elevated red blood cell distribution width in overweight and obese people: a community-based, cross-sectional study Ibrahim Halil Altiparmak, Muslihittin Emre Erkus, Aydemir Kocarslan, Hatice Sezen, Ozgur Gunebakmaz, Yusuf Sezen, Zekeriya Kaya, Ali Yildiz, Recep Demirbag

Abstract Background: Obesity and overweight are risk factors for atherosclerosis. Red blood cell distribution width (RDW) is associated with subclinical cardiac diseases. The aim of this study was to investigate the association between RDW and aortic stiffness in overweight or obese subjects. Methods: A total of 101 overweight or obese subjects without overt cardiovascular disorders, and 48 healthy controls were enrolled. RDW, aortic pulse-wave velocity (PWV) and augmentation index 75 (Aix75) were evaluated. The case subjects were divided into two sub-groups according to PWV values; ≥ 10 m/s in group I, and < 10 m/s in group II. Bivariate correlation and multiple regression analyses (stepwise) were performed. Results: RDW and PWV were considerably increased in the case groups compared with the controls. RDW was significantly increased in group I compared with group II and the controls [median 12.0 m/s, interquartile range (IQR): 10.5–17.5; median 11.7 m/s, IQR: 10.2–14.2, and median 11.4 m/s, IQR: 9.6–15.5, p < 0.05, respectively]. Resting heart rate and age were higher in group I than group II (81 ± 11 vs 74 ± 12 beats/min and 41 ± 120 vs 36 ± 9 years, respectively, p < 0.05). Regression analyses revealed that while log-RDW, age and resting heart rate were independent predictors for aortic PWV, log-RDW was the most important predictor in the final model.

Department of Cardiology, Faculty of Medicine, Harran University, Sanliurfa, Turkey Ibrahim Halil Altiparmak, MD, ihaltiparmak@gmail.com Muslihittin Emre Erkus, MD Ozgur Gunebakmaz, MD Yusuf Sezen, MD Zekeriya Kaya, MD Recep Demirbag, MD

Department of Cardiovascular Surgery, Faculty of Medicine, Harran University, Sanliurfa, Turkey Aydemir Kocarslan, MD

Department of Clinical Biochemistry, Faculty of Medicine, Harran University, Sanliurfa, Turkey Hatice Sezen, MD

Department of Cardiology, Faculty of Medicine, Hacettepe University, Ankara, Turkey Ali Yildiz, MD

Conclusions: RDW, resting heart rate and age independently predicted arterial stiffness, and RDW may be useful to provide an early recognition of subclinical atherosclerosis in overweight and obese individuals. Keywords: erythrocyte indices, vascular stiffness, overweight, obese Submitted 18/9/15, accepted 19/1/16 Published online 19/2/16 Cardiovasc J Afr 2016; 27: 246–251

www.cvja.co.za

DOI: 10.5830/CVJA-2016-005

In addition to traditional risk factors, overweight and obesity are important risk factors for the development of atherosclerosis and cardiovascular events.1 Previous studies demonstrated that arterial stiffness (AS) was impaired in these populations.2 AS, which is a result of functional and structural disorders of the arterial wall, signifies end-organ damage and increased risk of cardiovascular events.3 Several indicators provide valuable information about AS. Among these, aortic pulse-wave velocity (PWV) and augmentation index occupy an important place. Many studies have demonstrated that aortic PWV is associated with subclinical coronary atherosclerosis, significant coronary artery disease, hypertension and kidney disease, and it has a predictive value for cardiovascular events.3-6 Red cell distribution width (RDW), part of a routine complete blood count, is a laboratory evaluation of the variability in the volume and size of circulating erythrocytes. It is usually used for a differential diagnosis of anaemia.7,8 Many recent studies have revealed relationships between high RDW levels and adverse cardiovascular conditions, such as heart failure,9 severity and complexity of coronary artery disease,10 coronary slow flow,11 isolated coronary artery ectasia,8 acute myocardial infarction,12 and lack of coronary collateral vessels in acute coronary syndromes.7 The exact mechanisms causing elevated RDW are uncertain in these clinical events. However, it has been asserted that inflammation and oxidative stress may be possible pathophysiological mechanisms underlying increased RDW levels in cardiovascular diseases.11 To our knowledge, the association of RDW with the markers of AS in overweight and obese individuals is unknown. The aim of the study was to determine whether there was any relationship between RDW and AS in this population.

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Methods A total of 159 consecutive overweight or obese volunteers without overt cardiovascular disorders was enrolled as the case group and 48 healthy volunteers formed the control group in this community-based, cross sectional study (age between 18 and 75 years, mean 38 ± 11 and 37 ± 7 years, respectively). Fiftyeight of the case group who had systemic diseases, were on any medication, and consumed alcohol or smoked were excluded from the study. The aortic PWV and aortic normalised augmentation index to 75 beats/min heart rate (Aix75) were measured in the remaining participants. The case subjects were divided into two groups based on aortic PWV values; those with aortic PWV ≥ 10 m/s were included in group I, and those with aortic PWV < 10 m/s were included in group II. The study design is shown in Fig. 1. The study conformed to the recommendations of the Declaration of Helsinki on biomedical research involving human subjects. The study protocol was approved by the ethics committee and each participant provided written, informed consent.

Measurement of RDW and AS Samples of peripheral venous blood were drawn from the antecubital vein on admission after local antisepsis. Complete blood counts were measured using an autoanalyser (Sysmex K-1000, Block Scientific, USA) within five minutes of sampling. RDW (%) is one of the parameters automatically calculated during a full blood count. It is an index of the size distribution of red blood cells.

159 subjects with ≥ 25 BMI without overt cardiovascular disease and 48 healthy volunteers (BMI < 25) 58 subjects with ≥ 25 BMI were excluded due to: • Diabetes mellitus • Hypertension • Rheumatological disorders • Chronic renal insufficiency • Smoking • Alcohol consumption • Any medication 149 subjects enrolled in the study

Case group n = 101

Group I (PWV ≥ 10 m/s) n = 47

Control group n = 48

Group II (PWV < 10 m/s) n = 54

Fig. 1. D iagram of the study design. BMI: body mass index; PWV: pulse-wave velocity.

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The following measurements were assessed on a TensioMedTM arteriograph (TensioMed Ltd, Hungary): aortic PWV (m/s), aortic Aix75 (%), resting heart rate (beats/min), and systolic and diastolic blood pressures (mmHg). These parameters were measured according to the recommendations of the European Society of Hypertension for blood pressure and resting heart rate measurements.13 Subjects rested in the sitting position for at least five minutes and measurements were taken non-invasively using an appropriate cuff on the TensioMedTM arteriograph. The choice of cuff size (small, medium and large) was automatically determined by the arteriography according to the arm circumference, and the jugulum symphysis size reflects the interval from the aortic root to the bifurcation. Arteriography perceives brachial artery pulsations based on the oscillometric principle, and assesses parameters of AS by analysing pulse-wave forms. Aortic PWV is determined by calculating the travelling time of the pulse wave between two reference points.14 As there is a linear relationship between heart rate and Aix, the Aix is standardised to a heart rate of 75 beats/ min (Aix75). Aix (%) represents the ratio of reflected wave to primary wave, which moves from heart to tissues. It is inversely associated with arterial or aortic compliance.15 All parameters were measured by the investigators in the morning between 8:00 and 10:00, after 12 hours of fasting, and the average of at least three measurements was used.

Statistical analysis All analyses were done using SPSS version 20.0 (IBM Corporation, USA). The parameters were expressed as numbers (percentage) for categorical data, mean ± SD for parametric data, and median with interquartile range (IQR) for non-parametric data. We used the one-sample Kolmogorow–Smirnow test to assess normality of the data. The Student’s t-test (for parametric data) and the Mann– Whitney U-test (for non-parametric data) were used to compare variables between the two groups. Also, ANOVA and the Kruskal– Wallis tests were used for comparisons between the three groups (for parametric and non-parametric variables, respectively). To test gender differences between the groups, the chi-squared test or Fisher’s exact test, where appropriate, were used. Statistical significance between the variables was set at p < 0.05. We used bivariate correlation analysis to determine the correlation between statistically significant variables. Pearson’s correlation was used for data with normal distributions, and Spearman’s correlation was used for data with a skewed distribution. After definition of the factors that were associated with aortic PWV in bivariate analysis, independent predictors for estimation of aortic PWV were determined using multiple linear regression analyses with stepwise exclusion of these factors, using a criterion of p < 0.05 for retention of factors in the model. When evaluating RDW with skewed distribution in these analyses (correlation and regression), the variable was log-transformed (ln) and this logarithmic value was entered into the correlation and regression models.

Results The clinical and laboratory parameters of the two groups are presented in Table 1. Aortic PWV, RDW, body mass index (BMI)

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Table 1. Comparison of case group with control group

Table 2. Comparison of variables between the three groups

Case group (n = 101)

Control group (n = 48)

p-value

Male, n (%)

72 (71)

32 (67)

0.566

Age, years

38 ± 11

37 ± 7

0.564

Body mass index, kg/m2

28.8 ± 3.3

22.4 ± 1.6

< 0.001

Waist–hip ratio

0.91 ± 0.10

0.82 ± 0.07

< 0.001

Systolic blood pressure, mmHg

121 ± 9

120 ± 8

0.655

Diastolic blood pressure, mmHg

78 ± 5

77 ± 6

0.110

Resting heart rate, beats/min

77 ± 11

76 ± 12

0.719

Aortic pulse wave velocity, m/s

9.9 ± 2.0

9.1 ± 2.3

0.027

Aortic augmentation index 75, %

19 ± 11

17 ± 12

0.275

90 ± 5 (5 ± 0.28)

88 ± 6 (4.88 ± 0.33)

0.056

Fasting glucose, mg/dl (mmol/l) Urea, mg/dl Creatinine, mg/dl (mmol/l) Alanine aminotransferase, U/l Triglycerides, mg/dl (mmol/l) Total cholesterol, median (IQR), mg/dl (mmol/l) LDL cholesterol, mg/dl (mmol/l) HDL cholesterol, median (IQR), mg/dl

28 ± 5

26 ± 5

0.058

0.80 ± 0.08

0.78 ± 0.08

0.175

(70.72 ± 7.07)

(68.95 ± 7.07)

26 ± 5

24 ± 4

0.059 0.919

150 ± 19

149 ± 21

(1.70 ± 0.21)

(1.68 ± 0.24)

177 (128–244)

176 (116–227)

0.201*

109 ± 19

102 ± 17 (2.64 ± 0.44)

42 (33–52)

42 (38–53)

0.154 0.184*

Control (n = 48)

Males, n (%)

34 (72)

34 (63)

32 (67)

Age, years

41 ± 12a

36 ± 9

37 ± 7

Body mass index, kg/m2

29.1 ± 3.4b

28.3 ± 2.9c

22.4 ± 1.6

Waist–hip ratio

0.94 ± 0.10b

0.90 ± 0.09c

0.82 ± 0.07

Systolic blood pressure, mmHg

122 ± 8

120 ± 10

120 ± 8

Diastolic blood pressure, mmHg

80 ± 5

78 ± 6

77 ± 6

Resting heart rate, beats/min

81 ± 11 11.6 ± 1.4

Aortic pulse wave velocity, m/s

a,b

23 ± 13

Aortic augmentation index 75, % Fasting glucose, mg/dl (mmol/l) Urea, mg/dl Creatinine, mg/dl (mmol/l) Alanine aminotransferase, U/l Triglycerides, mg/dl Total cholesterol, median (IQR), mg/dl (mmol/l)

(mmol/l)

[1.09 (0.85–1.35)] [1.09 (0.98–1.37)] 11.9 (10.2–17.5)

11.4 (9.6–15.5)

0.005*

White blood cell, 103/µl

8.1 ± 1.6

7.6 ± 1.6

0.086

44 ± 5

43 ± 5

0.219

IQR: interquartile range; HDL: high-density lipoprotein; LDL: low-density lipoprotein; RDW: red cell distribution width. *p-value of Mann–Whitney U-test.

and waist–hip ratio were considerably higher in the case groups than the controls (p < 0.05) (Table 1). In subgroup analyses, RDW was significantly increased in group I compared with group II and the controls (p = 0.003 and p < 0.001, respectively), although there was no statistically significant change between group II and the controls (p > 0.05) (Table 2, Fig. 2). Aortic PWV was significantly higher in group I than in group II and the controls (p < 0.001 for both); however, it was similar in group II and the controls (p > 0.05). Although age was similar in group II and the controls (p > 0.05), it was considerably higher in group I than in group II (p = 0.046). While BMI and waist–hip ratio were significantly higher in groups I and II than in the controls, as expected (p < 0.001), there was no statistically significant difference between groups I and II (p > 0.05) (Table 2). Bivariate correlation analyses showed positive correlations of log-RDW, resting heart rate, age and BMI with aortic PWV (p < 0.001, p < 0.001, p = 0.022, and p = 0.007, respectively) (Table 3, Fig. 3A–C). Also, there was a positive correlation between log-RDW and BMI (Fig. 3D). However, multiple regression analyses (stepwise) using variables with significant correlation revealed that log-RDW, resting heart rate and age independently predicted aortic PWV. Of these, log-RDW was a stronger predictor than age and resting heart rate in the final model (Table 3).

HDL cholesterol, median (IQR), mg/dl

The main findings of this community-based, prospective study were: (1) the subjects with high aortic PWV had advanced

76 ± 12

8.4 ± 0.9

9.0 ± 2.3

17 ± 9

17 ± 12

90 ± 5

89 ± 6

88 ± 6

(5 ± 0.28)

(4.94 ± 0.33)

(4.88 ± 0.33)

28 ± 5

28 ± 4

26 ± 5

0.81 ± 0.06

0.80 ± 0.09

0.78 ± 0.08

(71.60 ± 5.30)

(70.72 ± 7.96)

(68.95 ±7.07)

25 ± 5

27 ± 4

24 ± 4

150 ± 19

148 ± 17

149 ± 21

(1.70 ± 0.21)

(1.67 ± 0.19)

(1.68 ± 0.24)

177 (128–244)

178 (136–223)

176 (116–227)

106 ± 18

108 ± 18

102 ± 17

(2.75 ± 0.47)

(2.80 ± 0.47)

(2.64 ± 0.44)

41 (34–52)

42 (33–48)

42 (38–53)

(mmol/l)

[1.06 (0.88–1.35)] [1.09 (0.85–1.24)] [1.09 (0.98–1.37)]

RDW, median (IQR), %

12.0 (10.5–17.5)a,b 11.7 (10.2–14.2)

White blood cell, 103/µl Haematocrit, %

11.4 (9.6–15.5)

8.3 ± 1.7

7.9 ± 1.6

7.6 ± 1.6

44 ± 7

45 ± 4

43 ± 5

Group I: pulse-wave velocity ≥ 10 m/s; group II: pulse-wave velocity < 10 m/s; IQR: interquartile range; HDL: high-density lipoprotein; LDL: low-density lipoprotein; RDW: red cell distribution width. a p < 0.05 between group I and II; bp < 0.05 between group I and control; cp < 0.05 between group II and control.

age, increased RDW and increased resting heart rate; (2) these parameters were positively correlated with aortic PWV; and (3) log-RDW, resting heart rate and age were independent predictors for increased aortic stiffness (aortic PWV ≥ 10 m/s), 13.50 13.00 12.50

p > 0.05

12.00 11.50 p = 0.003 p < 0.001

11.00 Group I

Discussion

74 ± 12a

[4.58 (3.32–6.32)] [4.61 (3.52–5.78)] [4.56 (3.0–5.88)]

LDL cholesterol, mg/dl (mmol/l)

RDW, median (IQR), % Haematocrit, %

Group II (n = 54)

(mmol/l)

[4.58 (3.32–6.32)] [4.56 (3.0–5.88)] (2.82 ± 0.49)

Group I (n = 47)

Parameters

95% CI, red cell distribution width, %

Parameters

Group II Groups

Control

Fig. 2. The error bar graph showing RDW difference between the groups. RDW: red cell distribution width.

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14 12 10 8

n = 149 β = 0.469 p < 0.001

6 0.95

1.00

1.05

1.10 1.15 Log-RDW

1.20

16 14 12 10 8

1.15

Log-RDW

1.20

10 8

n = 149 β = 0.188 p < 0.022

6 20

40 60 Age, years

80 100 Heart rate, beats per minute

120

1.25

n = 149 β = 0.417 p < 0.001

1.25

18 Pulse wave velocity, m/s

Pulse wave velocity, m/s

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1.10 1.05 1.00

n = 149 β = 0.239 p < 0.004

0.95 15

20 25 30 35 Body mass index, kg/m2

Fig. 3. S catter plot showing correlations of related parameters with each other. Aortic PWV with log-RDW (A), heart rate (B), age (C), and log-RDW with BMI (D). BMI: body mass index; PWV: pulse-wave velocity; RDW: red cell distribution wave.

and in the final model the strongest predictor among them was log-RDW. We know that increased body fat mass can lead to atherosclerotic vascular diseases.2 If there is no evidence of cardiovascular disease or disorders such as hypertension, hyperlipidaemia and diabetes in overweight or obese individuals, they are not treated with medication, but lifestyle changes are Table 3. Bivariate correlations and regression analyses (stepwise) of aortic PWV with related parameters Stepwise regression analyses

Parameters

Bivariate correlation

Model I (R2=0.304)

Model II (R2=0.456)

Model III (R2=0.492)

Aortic PWV Log-RDW 0.469 < 0.001 0.551 < 0.001 0.506 < 0.001 0.457 < 0.001 Resting heart rate

0.417 < 0.001

Age

0.188

0.022

BMI

0.223

0.007

WHR

0.128

0.174

0.393 < 0.001 0.421 < 0.001 0.196

0.003

For regression analyses of aortic PWV; predictors in model I: log-RDW; predictors in model II: log-RDW and resting heart rate; predictors in model III: logRDW, resting heart rate and age. Aix75: augmentation index 75; BMI: body mass index; PWV: pulse-wave velocity; RDW: red cell distribution width; WHR: waist–hip ratio.

suggested.16 Despite this, cardiovascular events appear more frequently in the obese population than in individuals with a lower BMI.1 In this study, we speculated that RDW may be used to determine persons at high risk for atherosclerosis among obese and overweight individuals without overt cardiovascular disorders. Thus individuals who are prone to atherosclerosis could more easily be identified using RDW and more closely followed up. RDW, which is usually overlooked and almost only used in the differential diagnosis of anaemia in daily practice, is typically raised in some cardiac and non-cardiac conditions, such as coronary artery disease, peripheral artery disease, acute coronary syndromes, heart failure, malnutrition, neoplastic metastases to bone marrow and inflammatory bowel disease.7,17-19 RDW generally reflects ineffective red cell production or their increased rate of destruction. However, it has been demonstrated that oxidative stress and chronic inflammation also cause elevated RDW. By destruction of the erythrocytes, haemolytic conditions lead to elevation of RDW, and impaired erythropoiesis, haemoglobinopathies, folic acid or cyanocobalamin deficiency, and iron deficiency also cause raised RDW.19,20 On the other hand, inflammatory cytokines have an effect on iron metabolism and the production or activity of erythropoietin.

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They also impair the erythrocyte membrane and maturation of red blood cells. As a result, they lead to the release of juvenile erythrocytes from bone marrow into the circulation.21 Additionally oxidative stress can affect the production of red blood cells. These cells have tremendous antioxidant capacity and are a primary oxidative sink, nevertheless, they are susceptible to oxidative damage. The oxidative stress associated with many clinical conditions brings about raised RDW.8 Activation of neurohumoral mediators (such as angiotensin II and the sympathetic system hormones) as well as inflammation and oxidative stress stimulate abnormal erythropoiesis. Therefore the survival of red blood cells is diminished on account of ineffective erythropoiesis.7,21 All of these conditions can be observed in atherosclerotic cardiovascular disease, leading to elevation of RDW. In the present study, we aimed to investigate whether RDW had an association with AS, which is an indicator of subclinical atherosclerosis. We hoped to recognise atherosclerosis at an early stage using RDW level, which is a routine parameter of a full blood analysis in most patients. Because RDW could be affected by many clinical conditions, we included healthy volunteers with a high BMI in the study. Those with low haematocrit levels or who had undergone blood transfusions in the previous six month were excluded. We did not investigate whether there were iron, folic acid or cyanocobalamine deficiencies in the subjects, since the mean haematocrit level of the study population was normal. The results of this study support our hypothesis, and indicate the strong positive correlation between log-RDW and aortic PWV. Also, age, resting heart rate and BMI were positively correlated with aortic PWV. However, multiple regression analysis suggested that log-RDW, resting heart rate and age were independent predictors for aortic PWV. It is known that elasticity of the arteries is impaired with advanced age.22,23 Similarly, it has been reported that increased resting heart rate is associated with aortic stiffness and atherosclerosis.24,25 Our study supports these results. However, an interesting result of our study was that RDW was the most important independent predictor for aortic PWV in the stepwise regression analyses. In clinical practice, we believe that RDW and resting heart rate, especially with advanced age, can indicate AS or subclinical atherosclerosis in obese and overweight individuals. Therefore, these patients may be identified at an early stage of atherosclerosis and medically treated earlier and more aggressively, before the development of any symptoms of atherosclerotic disease. Aortic PWV, which is the travelling time of the pulse wave from one point to another, indicates arterial stiffness, and high values reflect atherosclerotic cardiovascular disease with or without symptoms.4,6,26 Atherosclerotic disease may be determined by various symptoms or acute events; however, subclinical atherosclerosis is not be easily recognised. Therefore, some determinants of AS, such as aortic PWV and Aix75, have been used to detect subclinical atherosclerosis. Aortic PWV is the gold standard to evaluate AS and cardiovascular risk, as has been shown in many studies.13,16,25,26 A number of recently published studies have demonstrated elevated aortic PWV in several cardiovascular disorders. Kullo et al. showed that aortic PWV was related to subclinical coronary atherosclerosis, detected by the presence of coronary artery calcium using computed tomography.4 Another study by

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Catalano et al. pointed out that augmentation index in patients with peripheral artery disease, which is another parameter of AS, was higher than in the normal population.27 Our study showed no changes in aortic Aix75 between the case subjects and controls, unlike aortic PWV. Obesity is an important risk factor for various health problems, including atherosclerosis. It has recently been demonstrated that people using high-fat dairy products had a higher aortic PWV and carotid media thickness compared with those using low-fat dairy products.28 VayĂĄ et al. and Fujita et al. reported an elevated RDW in obese subjects, and our results are compatible with these.29,30 However, previous studies asserted that different causes might be responsible for this elevation. Among them, inflammation and hyposideraemia were blamed as responsible mechanisms.29,30 There is no consensus on the pathogenesis of the rise in RDW in obese and overweight populations. We also demonstrated a raised RDW and PWV in obese and overweight persons compared with the healthy controls. However, when the case group was divided into two subgroups according to aortic PWV, the RDW of the overweight and obese subjects with low aortic PWV was similar to that of the controls. Conversely, those with high aortic PWV had a considerably elevated RDW compared with the controls and the other group (obese and overweight with low PWV). Also, although there was a linear correlation between BMI and RDW as well as aortic PWV, stepwise regression analyses showed that log-RDW was an independent predictor in the final models for the estimation of aortic PWV. These results indicated that impaired aortic elastic properties with high aortic PWV may contribute to increased RDW in overweight and obese patients, apart from the previously stated reasons. There are some limitations of our study. The main limitation is the relatively small sample size. Another is that iron, folic acid and cyanocobalamin levels were not determined in the participants. However, volunteers with a haematocrit lower than 35% were not included in the study. Lastly, we did not evaluate adipocytokines, such as leptin, adiponectin, resistin, chemerin and visfatin, knowing their relationship with atherosclerosis and angiogenesis.

Conclusion This study indicated that raised aortic stiffness may be responsible for elevated RDW values in obese and overweight persons. Therefore, RDW may help to provide an early recognition of atherosclerosis using a simple test instead of more sophisticated devices, particularly if evaluated together with resting heart rate in the older population.

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without obstructive coronary disease: a preliminary study. Eur Rev Med

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Chinese population. Intern Med 2011; 50(24): 2941–2945. 19. Tseliou E, Terrovitis JV, Kaldara EE, Ntalianis AS, Repasos E, Katsaros

Independent prognostic value of the ambulatory arterial stiffness index

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Li XL, Hong LF, Jia YJ, Nie SP, Guo YL, Xu RX, et al. Significance

22. Strazhesko I, Tkacheva O, Boytsov S, Akasheva D, Dudinskaya E,

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telomere length in adults free of cardiovascular diseases. PLoS One

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Felker GM, Allen LA, Pocock SJ, Felker GM, Allen LA, Pocock SJ, et

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al.; CHARM investigators. Red cell distribution width as a novel prog-

23. Hodson B, Norton GR, Booysen HL, Sibiya MJ, Raymond A, Maseko

nostic marker in heart failure: data from the CHARM program and the

MJ, et al. Brachial pressure control fails to account for most distend-

Duke databank. J Am Coll Cardiol 2007; 50(1): 40–47.

ing pressure-independent, age-related aortic hemodynamic changes in

10. Isik T, Uyarel H, Tanboga IH, Kurt M, Ekinci M, Kaya A, et al.

adults. Am J Hypertens 2015 Aug 20. pii: hpv140. [Epub ahead of print].

Relation of red cell distribution width with the presence, severity, and

24. Whelton SP, Blankstein R, Al-Mallah MH, Lima JA, Bluemke DA,

complexity of coronary artery disease. Coron Artery Dis 2012; 23(1):

Hundley WG, et al. Association of resting heart rate with carotid and

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aortic arterial stiffness: multi-ethnic study of atherosclerosis. Hypertension

11. Luo SH, Jia YJ, Nie SP, Qing P, Guo YL, Liu J, et al. Increased red cell

2013; 62(3): 477–484. doi: 10.1161/HYPERTENSIONAHA.113.01605.

distribution width in patients with slow coronary flow syndrome. Clinics

25. Reference values for arterial stiffness collaboration. Determinants of

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Red cell distribution width is associated with acute myocardial infarction in young patients. Cardiol J 2012; 19(6): 597–602. doi: 10.5603/ CJ.2012.0111. 13. Mancia G, Fagard R, Narkiewicz K, Redon J, Zanchetti A, Böhm M, et al. 2013 ESH/ESC guidelines for the management of arterial hyper-

J 2010; 31(19): 2338–2350. doi: 10.1093/eurheartj/ehq165. 26. Mitchell GF, Hwang SJ, Vasan RS, Larson MG, Pencina MJ, Hamburg NM, et al. Arterial stiffness and cardiovascular events: the Framingham Heart Study. Circulation 2010; 121(4): 505–511. doi: 10.1161/ CIRCULATIONAHA.109.886655.

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15. Hlimonenko I, Meigas K, Viigimaa M, Kozlíková K, Janiga I. Assessment of pulse wave velocity and augmentation index in different arteries in patients with severe coronary heart disease. Conf Proc IEEE Eng Med Biol Soc 2007; 2007: 1703–1706. 16. Perk J, De Backer G, Gohlke H, Graham I, Reiner Z, Verschuren M, et al. European Association for Cardiovascular Prevention &

velocity and carotid intima–media thickness in adults. Nutr J 2014; 13: 37. doi: 10.1186/1475-2891-13-37. 29. Vayá A, Alis R, Hernandez-Mijares A, Solá E, Cámara R, Rivera L, et al. Red blood cell distribution width is not related with inflammatory parameters in morbidly obese patients. Clin Biochem 2014; 47(6): 464–466. doi:10.1016/j.clinbiochem.2014.01.020.

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Electrocardiographic findings in a cross-sectional study of human immunodeficiency virus (HIV) patients in Enugu, south-east Nigeria PO Njoku, EC Ejim, BC Anisiuba, SO Ike, BJC Onwubere

Abstract Background: Electrocardiographic (ECG) abnormalities are prevalent in subjects with human immunodeficiency virus (HIV) infection. In this study, three groups of subjects were investigated and the prevalence of ECG abnormalities was analysed. Methods: A cross-sectional study was carried out on adults between November 2010 and November 2011 at the University of Nigeria Teaching Hospital, Enugu, Nigeria. One hundred HIV-infected patients on highly active anti-retroviral therapy (HAART), 100 HIV-infected HAART-naïve patients and 100 HIV-negative controls were recruited. Twelve-lead electrocardiograms were done on all subjects. Data were analysed using the chi-squared, Student’s t-, one-way ANOVA and Duncan post hoc tests. Results: Left-axis deviation was seen in 15 (16%) of the HIV-positive subjects on HAART, 10 (13.7%) of the HAART-naïve subjects and eight (21%) of the controls (p = 0.265). Eight (11%) subjects with left ventricular hypertrophy (p < 0.001) and two (2.7%) with ST-segment elevation were found among the HIV-positive HAART-naïve subjects (p = 0.134). Prolonged QTc interval was seen in 17 (18.2%) of the HIV-positive patients on HAART, 12 (16.4%) of the HIV-positive HAART-naïve patients and four (10.5%) of the controls (p = 0.012). Conclusion: The prevalence of ECG abnormalities was higher in the HIV-positive patients on HAART (93%) and the HIV-positive HAART-naïve patients (73%) compared to the controls. Keywords: abnormalities, electrocardiogram, highly active antiretroviral therapy, human immunodeficiency virus Submitted 19/11/15, accepted 26/1/16 Cardiovasc J Afr 2016; 27: 252–257

www.cvja.co.za

DOI: 10.5830/CVJA-2016-007

Department of Medicine, University of Nigeria Teaching Hospital, Ituku-Ozalla, Enugu, Nigeria PO Njoku, MB BS, MSc, FMCP, passyokuchi@yahoo.com EC Ejim, MB BS, FMCP BC Anisiuba, MB BS, FMCP SO Ike, MB BS, FMCP BJC Onwubere, MB BS, FMCP

Department of Medicine, Federal Medical Centre, Keffi, Nasarawa State, Nigeria PO Njoku, MB BS, MSc, FMCP, passyokuchi@yahoo.com

The global prevalence of HIV/AIDS in people aged 15–49 years was 0.8% in 2011,1 affecting approximately 34 million people. An estimated 23.5 million (22.1–24.8 million) of these people, representing 69% of the global HIV burden, reside in sub-Saharan Africa.2 Worldwide, Nigeria has the second highest number of new infections reported each year, and an estimated 3.7% of the population or 3.4 million people are living with HIV infection.3,4 With increased access to anti-retroviral therapy (ART) in resource-poor countries, longevity has increased among people living with HIV/AIDS.5 Indeed, dramatic reductions in morbidity and mortality rates have been noted in these patients since the introduction of ART in 1996. In Nigeria, this has prolonged and improved the quality of life of HIV/AIDS patients, with survival data of 68.3% of adults and children who were on ART in 2009, alive and healthy after 12 months.6 Although the use of highly active anti-retroviral therapy (HAART) is associated with virological suppression and immunological recovery in people living with HIV infection, HIV and HAART, especially protease inhibitors, induce disorders of lipid metabolism,7,8 such as diabetes and dyslipidaemia, which are implicated in the increased incidence of cardiovascular disease in this patient population.9,10 The electrocardiogram (ECG) identifies abnormalities in HIV/AIDS patients whether or not they were suspected of having cardiac disease.11 Barbaro and colleagues found ECG abnormalities, including supraventricular and ventricular ectopic beats, as well as non-specific ST–T-wave abnormalities in about 57% of asymptomatic HIV-infected patients.12 Sani and colleagues13 documented abnormal ECG findings in 81% of 100 AIDS patients, 65% of 78 HIV-positive asymptomatic subjects and 37.5% of 80 HIV-negative subjects. The ECG abnormalities described included different types of arrhythmias, low-voltage QRS complexes, non-specific ST-segment and T-wave changes, poor R-wave progression, right bundle branch block, axis deviations, enlargement of various heart chambers and QTc prolongation. Asymptomatic ECG findings specific for myocardial ischaemia (Q waves and ST-segment depression, and T-wave inversion in men) signify an increased risk of myocardial infarction or death in HIV-uninfected adults,14 and may have worse consequences in HIV-infected individuals. The ECG, which is fairly widely available in resource-poor countries, is helpful in evaluating HIV subjects for cardiovascular disease. Considering the huge impact of the HIV/AIDS disease burden in sub-Saharan Africa, increasing access to HAART as well as the improved longevity of people living with HIV/ AIDS, there is a need for proper cardiovascular evaluation of this patient population. Early identification and management of cardiovascular diseases in this population would optimise patient

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care and provide the much-needed information on this subject in sub-Saharan Africa. This study analysed the ECG findings in HIV-infected individuals, including those on HAART, and HIV-negative subjects residing in Enugu, Nigeria.

Methods This cross sectional study was carried out from November 2010 to November 2011 at the University of Nigeria Teaching Hospital (UNTH), Enugu, south-east Nigeria. The procedures followed were in accordance with the ethical standards of the Helsinki Declaration (1964, amended most recently in 2008) of the World Medical Association. This study was approved by the ethics committee of the UNTH, Enugu, and written consent was obtained from the subjects. Information obtained was anonymised as far as possible. Inclusion criteria were adult Nigerians aged 18 years and older with confirmed HIV-positive serology. HIV screening was by enzyme-linked immunoassay (ELISA) and confirmed by Western-blot electrophoresis, while CD4 T-lymphocytes (CD4 cells) were quantified by flow cytometry. Sample size was calculated using the Fisher’s formula:15

Table 1. Comparison of mean ± standard deviations of parameters across the three groups using one-way ANOVA HIV-positive on HAART

Parameters

HIV-positive HAART-naïve

Age (years)

35.85 ± 8.94

Weight (kg)

65.77 ± 13.92* 62.40 ± 12.45 1.66 ± 0.07*

Height (m)

Heart rate (bpm)

F-value p-value 0.716

0.490

68.69 ± 8.67*

7.007

0.001

1.71 ± 0.79

17.886

< 0.001

6.301

0.002

1.81 ± 0.15

4.420

0.013

82.92 ± 14.08* 84.28 ± 16.79* 68.77 ± 8.02

40.232

< 0.001

1.77 ± 0.17*

BSA (m2)

1.66 ± 0.09*

Control 35.76 ± 9.74

24.18 ± 3.32*

24.14 ± 4.55*

BMI (kg/m2)

34.43 ± 9.49

22.47 ± 3.65 1.75 ± 0.18*

*Duncan post hoc multiple comparison test indicating means for groups in homogenous subsets (means not significantly different). BMI = body mass index, BSA = body surface area.

Echocardiography was also carried out on each of the subjects using the SonoScape SS1-5000 machine and transducer of frequency 3.5 MHz. M-mode, two-dimensional, pulsed-wave, continuous-wave, tissue Doppler imaging and colour Doppler assessments were done with the subject in the left lateral decubitus position. Measurements were taken (in cm) using the American Society of Echocardiography guidelines (leading-edge methodology).18

z2pq

n = ____   d      2

where n = minimum sample size; z = 95% confidence level i.e. 1.96; d = level of precision (0.075)13; p = maximum prevalence reported in a study of a similar population16 (13.6%); and q = 1–p. A minimum sample size of 80 was calculated. For the purpose of the study, 100 HIV-positive patients who had not taken HAART were recruited into the HIV-positive, HAART-naïve group. One hundred patients who had received HAART for at least three months were enrolled in the group of HIV-positive patients on HAART. Another 100 controls with already known HIV-negative serology were recruited from those being screened for blood donation, marriage and insurance purposes. Patients in end-stage AIDS disease, classified as category C by the Centre for Disease Control, 1993, were excluded.17 Other subjects excluded were those under 18 years of age, individuals with arterial hypertension, coronary artery disease or active symptoms suggestive of ischaemic heart disease or congestive cardiac failure, cardiomyopathy, peripheral or cerebrovascular disease or diabetes mellitus. Further exclusion criteria were patients who were pregnant or in pueperium, as well as those with a significant history of tobacco and or alcohol use, or those who used drugs known to affect the cardiovascular system. All subjects were evaluated clinically and anthropometric parameters such as height (m), weight (kg), body mass index (kg/ m2) and body surface area (m2) were assessed. Qualifying subjects had a resting 12-lead surface ECG recording in the supine position at a speed of 25 mm/s using a two-channel automated Techmel ECG machine (USA), ECG-1101 model. ECG tracings from each participant were analysed in the standard fashion with the long lead II tracing serving as the rhythm strip. Parameters analysed were heart rate, rhythm, P wave (duration, shape), height (paroxysmal atrial complexes), PR interval, QRS wave (duration, shape, height, axis), paroxysmal ventricular complexes, QT interval, QTc, Q wave, T wave (shape), ST-segment (shape), and R and S waves for ventricular hypertrophy.

Statistical analysis Statistical analysis of data was done using EPI INFO version 6 software. The chi-squared test was used to test the association between categorical variables. Continuous variables were analysed using the Student’s t-test. Comparison of mean ± standard deviations of parameters across the three groups was done using one-way ANOVA, and the Duncan post hoc multiple comparison test was done to indicate means for groups in homogenous subsets (means not significantly different) (Table 1). A p-value < 0.05 was taken as statistically significant.

Results Three hundred adults were recruited for the study, comprising the group of 100 HIV-positive patients on HAART, made up of 49 males and 51 females, 100 HIV-positive HAART-naïve patient group, made up of 48 males and 52 females, and 100 apparently healthy adults (control group), made up of 52 males and 48 females. There was no significant difference in the gender distribution of these three groups (χ2 = 0.347, p = 0.841) (Table 2). The mean age of the HIV-positive patients on HAART was 35.85 ± 8.94 years, that of the HIV-positive HAART-naïve patients was 34.43 ± 9.49 years, while that of the control group was 35.76 ± 9.74 years. There was no significant difference in the mean age of the three groups (F = 0.72, p = 0.49). There was no significant difference in the age groups of the patients and controls (χ2 = 4.74, P = 0.19) (Table 3).

Table 2. Gender distribution of the study groups Male, n (%)

Female, n (%)

HAART

51 (51.0)

49 (49.0)

100 (100)

HAART-naive

48 (48.0)

52 (52.0)

100 (100)

52 (52.0)

48 (48.0)

100 (100)

151 (50.3)

149 (49.7)

300 (100)

Groups

Control Total

Total, n (%)

χ2 = 0.347, p = 0.841. HAART = highly active antiretroviral therapy.

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Table 3. Age distribution in the study groups HIV-positive on HAART, n (%)

HIV-positive HAART-naïve, n (%)

HIV-negative control, n (%)

< 26

11 (23.9)

19 (41.3)

16 (34.8)

46 (100)

26–30

22 (37.9)

19 (32.8)

17 (29.3)

58 (100)

31–35

21 (30.0)

24 (34.3)

25 (35.7)

70 (100)

36–40

17 (35.4)

16 (33.3)

15 (31.3)

48 (100)

41–45

13 (46.4)

7 (25.0)

8 (28.6)

28 (100)

46–50

9 (31.0)

11 (37.9)

29 (100)

> 50

7 (33.3)

6 (28.6)

8 (38.1)

21 (100)

100 (33.3)

300 (100)

Age (years)

Total

Total, n (%)

χ2 = 4.739, p = 0.192. HIV = human immunodeficiency virus; HAART = highly active antiretroviral therapy.

The mean duration of HAART medication for the HIV-positive patients on HAART was 4.0 ± 2.4 years, with minimum and maximum durations of one and 10 years, respectively. Of these patients, 7% took the HAART regimen containing protease inhibitors (PIs), while 93% took HAART that did not contain PIs. However, those on PIs received it for less than six months. Table 4 shows ECG abnormalities in the study groups and controls. T-wave inversion (< 3 mm) in leads V1–V3 (anterior leads) was the commonest abnormality in all study groups. It was seen in 44 (65.7%) of the HIV-positive subjects on HAART, 22 (45.8%) of the HIV-positive HAART-naïve subjects and 14 (29.2%) of the controls. T-wave inversion (< 3 mm) in leads II, III and aVF (inferior leads) was also seen in two (2.2%) HIV-positive subjects on HAART, one (1.4%) HIV-positive HAART-naïve subject, and six (15.8%) control subjects. Left-axis deviation (LAD), that is, QRS axis of < 0° to 90°, was seen in 15 (16%) of the HIV subjects on HAART, 10 Table 4. ECG abnormalities in the study population HIVpositive on HAART, n (%)

HIVpositive HAART naïve, n (%)

Control, n (%)

LAD

15 (16)

10 (13.7)

8 (21)

2.656

0.265

T wave inversion in leads V1 –V3

44 (47)

22 (30.4)

14 (36.8)

24.682

< 0.001

Low QRS voltage complex

1 (1.1)

0 (0)

2.007

0.367

1st-degree heart block

3 (3.2)

1 (1.4)

2 (5.3)

1.020

0.600

T-wave inversion in leads II, III, aVF (inferior leads)

2 (2.2)

1 (1.4)

6 (15.8)

4.811

0.090

VEB

1 (1.1)

1 (1.4)

0 (0)

2.007

0.367

0 (0)

2 (2.7)

2 (5.3)

2.027

0.363

LBBB

1 (1.1)

0 (0)

2.007

0.367

RBBB

1 (1.1)

0 (0)

2 (5.3)

2.020

0.364

LVH

0 (0)

8 (11)

0 (0)

16.438

< 0.001

Sinus tachycardia

8(8.6)

14 (19.2)

0 (0)

2.020

0.364

ST-segment elevation

0 (0)

2 (2.7)

0 (0)

4.027

0.134

Sinus bradycardia

0 (0)

2 (5.3)

25.000

< 0.001

Mean QTc ± SD

0.42 ± 0.04

0.41 ± 0.04

0.39 ± 0.03

*15.779

< 0.001

Prolonged QTc

17 (18.2)

12 (16.4)

4 (10.5)

8.784

0.012

Total

93 (100)

73 (100)

38 (100)

ECG abnormalities

T-wave inversion in leads I, aVL,V5–V6 (lateral leads)

χ2

p-value

*F-value. For QTc, F = 15.779; p < 0.001. Duncan post hoc multiple comparison test showed all significantly different. LAD = left axis deviation, VEB = ventricular ectopic beat, LBBB = left bundle branch block, RBBB = right bundle branch block, LVH = left ventricular hypertrophy.

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(13.7%) of the HAART-naïve subjects and eight (21%) of the controls. First-degree heart block (PR interval > 0.2 s) was seen in three (3.2%) HIV-positive subjects on HAART, one (2.1%) HIV-positive HAART-naïve subject, and two (5.3%) control subjects. One (1.1%) subject with low QRS-wave voltage complex (amplitude of QRS complex < 5 mm in limb leads or > 10 mm in precordial leads), left bundle branch block (LBBB), and right bundle branch block (RBBB), respectively, was found in the group of HIV-positive subjects on HAART, while eight (11%) subjects with left ventricular hypertrophy (LVH) and two (2.7%) with ST-segment elevation were found among the HIV-positive HAART-naïve subjects. Sinus tachycardia (heart rate > 100 beats per minute) was seen in eight (8.6%) HIV-positive subjects on HAART, and 14 (19.2%) control subjects, while sinus bradycardia (heart rate < 60 beats per minute) was seen in only two (5.3%) of the control subjects. Prolonged QTc (QTc > 0.44 in males or > 0.46 in females) was seen in 17 (18.2%) of the HIV-positive patients on HAART, 12 (16.4%) of the HIV-positive HAART-naïve patients and four (10.5%) of the control subjects.

Discussion The prevalence of ECG abnormalities was higher among HIV-positive patients on HAART (93%) and HIV-positive HAART-naïve patients (73%), compared to the controls (38%) (Table 4). Okoye19 found a similar prevalence of ECG abnormalities in 80% of AIDS patients with CD4 cell counts < 200 cells/mm3, 60% of HIV-positive subjects with CD4 cell counts > 200 cells/mm3 and 35% of HIV-negative healthy controls. The prevalence of ECG abnormalities found in both HIV-positive patients on HAART and HIV-positive HAARTnaïve patients in this study also compared favourably with the rate of 86% reported by Mouanodji et al.20 Conversely, the prevalence of ECG abnormalities in this study was significantly higher than the 53% found by Levy et al.21and the 55% documented by Herst et al.22 The difference may have been due to the small sample sizes of 32 and 21 patients in the studies done by Levy et al. and Herst et al., respectively. Also, not matching the numbers in the study groups, as well as inclusion of patients at advanced stages of AIDS by Levy et al. could have been reasons for the discrepancy. LAD, T-wave inversion in leads V1–V3 and prolonged QTc were the three most common ECG abnormalities found in HIV-positive patients on HAART and HIV-positive HAARTnaïve patients, respectively, in this study (Table 4). LAD and T-wave inversion in leads II, III, avF and V1–V3 were the three most common ECG abnormalities in the controls. LAD was found more often in both HIV-positive patients on HAART and HIV-positive HAART-naïve patients than in the control subjects. T-wave inversion in leads V1–V3 was the commonest ECG abnormality in all the groups, occurring in 47% of the HIV-positive patients on HAART, 30.4% of the HIV-positive HAART-naïve patients and 36.8% of the controls. It also occurred in smaller proportions in leads II, III, avF, I and avL. T-wave inversion may signify ischaemia but is often a non-specific finding, especially in women.23 No relationship between ECG abnormalities and gender was found in the study. ECG evidence of asymptomatic ischaemic heart disease (IHD) (Q wave or ST-segment depression) was not found in

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any of the study groups, although ST-segment elevation was seen in leads V2–V3 in two of the HIV-positive HAARTnaïve patients. Early repolarisation abnormality, commonly seen in blacks,24,25 may be an explanation for this. In a similar study involving 4 831 HIV-positive adults, including those with hypertension and diabetes mellitus, T-wave inversion was observed in 11.1% of participants and it was substantially more common in women than men. Shikuma et al. found ECG evidence of asymptomatic IHD in 10.9% of participants with no known IHD.23 Although the exact reason is not known, a higher occurrence of myocarditis, coronary artery vasculitis and ischaemia,26 which cause cardiac abnormalities in HIV-positive patients, may explain this higher prevalence. Sinus tachycardia was seen more often in the HIV-positive groups (8.6% in HIV-positive patients on HAART, 19.2% in HIV-positive HAART-naïve patients) compared to 0% in the controls. This may have been due to inter-current febrile illness, anaemia, myocarditis and increased metabolic demand in HIV patients.27 These conditions induce autonomic dysfunction and increased sympathetic activity. The higher prevalence of sinus tachycardia in HIV-positive HAART-naïve patients may have been due to their higher immunosuppression and the prevalence of these identified factors. Sinus bradycardia, noted in 2% of the controls and 0% of the HIV-positive patients, may have been due to increased vagal tone, commonly seen in healthy young people.28 First-degree heart block was found in both patients and controls, although slightly more often in patients than in the control group. Although currently out of phase, febrile illness such as leptospirosis, which occurs commonly in HIV-positive patients, has been reported to cause complete heart block.29 The use of HAART, especially those containing PIs, in HIV infection is known to cause prolonged QRS duration, firstdegree atrioventricular block and complete bundle branch block. The conduction abnormalities seen in this study were not unexpected since most of the patients had recurrent fever, and were often not evaluated for leptospirosis. Moreover, some of the HIV-positive patients on HAART in this study were on PIs. Charbit et al.30 reported a higher number of patients showing prolonged QRS duration, first-degree atrioventricular block and complete bundle branch block in those taking PIs. Left ventricular hypertrophy (LVH) assessed by Sokolow and Lyon criteria, as well as Araoye’s voltage criteria,31 was found in eight (11%) of the HIV-positive HAART-naïve patients but in none of the HIV-positive patients on HAART or the controls. Four HIV-positive patients on HAART, four HIV-positive HAART-naïve patients and one control subject had LVH, derived with Devereux’s formula. We found left ventricular mass was increased in HIV infection (Table 5). In similar studies to ours, Barbaro et al.32 and Lipshultz et al.33 reported increased LV mass in HIV-positive patients. On the other hand, Martinez-Garcia et al.34 found decreased LV mass in asymptomatic HIV-infected patients, and Samaan et al.35 found decreased LV mass among patients with AIDS wasting syndrome. The mechanisms by which these adverse effects on LV mass occur in HIV-positive patients are not fully understood, but are thought to be related to mitochondrial toxicity.36 Many studies have shown that HIV virions directly affect myocardial cells and are associated with local release of cytokines and other factors leading to inflammation, myocarditis and dilated

cardiomyopathy.37 Also, increase or decrease in LV mass has been suggested to be associated with opportunistic infections and malnutrition,35 therefore a lower nadir CD4 cell count has been associated with higher LV mass index. Meng et al.38 reported greater interventricular septal and posterior wall thicknesses among patients exposed to PIs compared to those who were not exposed. The finding of higher numbers of HIV-positive patients with LVH in our study was not surprising, given the pathogenesis and sequela of HIV infection as well as the use of HAART. Pewsner et al. and Devereux found LVH, assessed by ECG, only in HIV-positive HAART-naïve patients, probably because of the high specificity and low sensitivity of the ECG.39,40 On the other hand, Michael found four patients with LVH, using echocardiography, in both HIV-positive patients on HAART and the HIV-positive HAART-naïve group, and one patient in the control group. This may have been because echocardiography is much more sensitive than ECG.41 QTc interval, corrected for heart rate using Bazett’s formula, was more prolonged in our HIV-positive patients on HAART and HIV-positive HAART-naïve patients than in the controls (Table 4). The prevalence of prolonged QTc was 34.6% in HIV-positive patients and 10.5% in the controls. A breakdown of this showed a higher prevalence of 18.2% in HIV-positive patients on HAART, compared to 16.4% in the HIV-positive HAART-naïve group. This is similar to the 45% reported by Okoye22 and 34.7% reported by Ogunmodede.42 Villa et al.43 reported a high prevalence of prolonged QTc interval of 65% in a highly selected cohort of HIV-positive patients who had already developed autonomic dysfunction. On the other hand, Kocheril et al.44 reported a prevalence of 29% in 42 AIDS patients in the absence of any known cause. QTc prolongation in HIV-positive patients has been attributed to electrolyte imbalance from poor nutrient intake and recurrent diarrhoea, drugs including zidovudine,45 protease inhibitors,46 pentamidine,46 halofantrin,47 trimethoprim-sulfamethoxazole,48 and autonomic dysfunction due to HIV-associated neuropathy.43 Okoye14 documented hypocalcaemia as the cause of QTc prolongation in 35% of AIDS patients. In our study, the higher QTc prolongation in the patients may have been due to electrolyte imbalance from vomiting and diarrhoea, PIs, Table 5. Comparison of echocardiographic parameters measured across the groups using one-way ANOVA Parameters

HIV-positive on HAART

HIV-positive HAART-naïve

AO (cm)

2.71 ± 0.40*

2.41 ± 0.37

2.74 ± 0.42*

21.363

< 0.001

LA (cm)

3.27 ± 0.62

2.68 ± 0.51

3.11 ± 0.47

31.385

< 0.001

EDD (cm)

4.73 ± 0.70*

4.41 ± 0.55

4.75 ± 0.42*

11.240

< 0.001

ESD (cm)

3.01 ± 0.51

2.84 ± 0.57

2.92 ± 0.43

2.616

0.075

IVS (cm)

0.77 ± 0.17*

0.85 ± 0.17

0.78 ± 0.15*

6.098

0.003

PW (cm) EF (%)

Control

F-value p-value

0.82 ± 0.16

0.87 ± 0.17

0.82 ± 0.13

2.878

0.058

68.95 ± 12.43*

72.81 ± 11.70

67.36 ± 9.04*

6.223

0.002

36.51 ± 8.64

37.77 ± 6.53

FS (%)

36.77 ± 9.81

0.623

0.537

LVM (g)

141.94 ± 49.75 138.61 ± 48.53 131.26 ± 31.55

1.540

0.216

LVMI (g/m2)

79.95 ± 26.25

2.760

0.065

77.55 ± 25.91

72.37 ± 16.52

*Duncan post hoc multiple comparison test indicating means for groups in homogenous subsets (means not significantly different). AO = aorta; LA = left atrium; EDD = end-diastolic diameter; ESD = end-systolic diameter; IVS = interventricular septum; PW = posterior wall of left ventricle; EF = ejection fraction; FS = fractional shortening; LVM = left ventricular mass; LVMI = left ventricular mass index.

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zidovudine, trimethoprim-sulfamethoxazole and antimalarial medications. The higher prevalence seen in HIV-positive patients on HAART, compared to the HIV-positive HAART-naïve group may have been from the use of lopinavir and zidovudine medications. There were some limitations of this study. First, we were unable to carry out biochemical tests such as serum electrolytes and lipograms in any of the subjects. Second, we were unable to determine the actual time of HIV infection, and by extension, the duration of HIV infection. Third, since this was an observational, cross-sectional study, we were unable to infer causality.

15. Araoye MO. Reseach Methodology with Statistics for Health and Social

Conclusion

19. Okoye IC. Electrocadiographic abnormalities of newly diagnosed

Dciences. Ilorin: Nathadex, 2003: 117–121. 16. Danbauchi SS, Sani BG, Alhassan AM, et al. Echocardiographic features of HIV/AIDS subjects on 1–2 years of ARV drugs in Nigeria. Available at http://www2.umdnj.edu/shindler/hivecho.html. 17. Centre for Disease Control. 1993 revised classification system for HIV infection and expanded surveillance case definition for AIDS among adolescents and adults. HIV/AIDS surveillance report, Atlanta 1993; 1–23. https://www.cdc.gov/mmwr/preview/mmwrhtml/00018871.htm. 18. Henry WL, De Maria A, Gramiak R, et al. Report of the American Society of Echocardiography committee on nomenclature and standards in 2-D echocardiography. Circulation 1980; 62: 212–217.

The prevalence of ECG abnormalities was higher in the HIV-positive patients on HAART (93%) and HIV-positive HAART-naïve patients (73%) compared to the apparently healthy controls. The use of ECG is helpful in cardiovascular evaluation of this patient population, especially in resource-poor countries.

patients with HIV/AIDS who are not on HAARTS at UNTH, Enugu. Part II dissertation. West Africa College of Physicians April, 2010. 20. Mouanodji M, Mbaingnro D, Brendon P. Clinical outline of AIDS patients with cardiac manifestations in Africa. Int conf AIDS 1994; 10: 266. 21. Levy WS, Simon GL, Rios JC, Ross AM. Prevalence of cardiac abnormality in HIV infection. Am J Cardiol 1989; 63: 86–89.

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United Nation’s General Assembly Special Session (UNGASS) Country Progress Report for Nigeria (Jan 2008 to Dec 2009) in March 2010, indicator 24: 35. http://data.unaids.org/pub/Report/2010/nigeria_2010_ country_progress_report_en.pdf.

14: 21–27. 23. Shikuma CM, Zackin R, Sattler F, et al. Changes in weight and lean body mass during highly active antiretroviral therapy. Clin Infect Dis 2004; 39: 1223–1230. 24. Araoye MA. Basic Electrocardiography. 2nd edn. Ibadan: Spectrum Books, 2012: 43. 25. Di Paolo FM, Schmied C, Zerguini YA. The athlete’s heart in adolescent Africans: An electrocardiographic and echocardiographic study. J Am Coll Cardiol 2012; 59: 1029–1036. 26. Paton P, Tabib A, Loire R, Tete R. Coronary artery lesions and human immunodeficiency virus infection. Res Virol 1993; 144: 225–231. 27. Okeahialam BN, Anjorin FL. Echocardiographic study of the heart in AIDS. The Jos experience. Trop Card 2000; 26: 3–6. 28. Aaron LB, Malissa JW. Athlete’s heart and cardiovascular care of the athlete, scientific and clinical update. Circulation 2011; 123: 2723–2735.

factors in HIV patients –Association with antiretroviral therapy. Results

29. Trivedi SV, Bhattacharya A, Amichandwala K. Evaluation of cardiovascular status in severe leptospirosis. J Assoc Physic Ind 2003; 51: 951–953.

Frerichs FC, Dingemans KP, Brinkman K. Cardiomyopathy with

30. Charbit B, Gayat E, Voiriot P, et al. Effects of HIV protease inhibitors

mitochondrial damage associated with nucleoside reverse-transcriptase

on cardiac conduction velocity in unselected HIV-infected patients. Clin

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tion in patients with Kaposi sarcoma and AIDS. Clin Inv Med 1991;

Friis-Moller N, Weber R, Reiss P, et al. Cardiovascular disease risk from the DAD study. AIDS 2003; 17: 1179–1193.

22. Herst JA, Shepherd FA, Liu P. Prospective assessment of cardiac func-

Powderly WG. Long-term exposure to lifelong therapies. J Acquir Immune Defic Syndr 2002; 29(Suppl 1): S28–40.

Pharmacol Therapeut 2011; 90: 442–448. 31. Araoye MA. Basic Electrocardiography. 2nd edn. Ibadan: Spectrum Books, 2012; 102–103.

10. Bozzette SA, Ake CF, Tam HK, et al. Cardiovascular and cerebro-

32. Barbaro G, Barbarini G, Di Lorenzo G. Early impairment of systolic

vascular events in patients treated for human immunodeficiency virus

and diastolic function in asymptomatic HIV-positive patients: a multi-

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center echocardiographic and echo-Doppler study. AIDS Res Hum

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AIDS. Internet J Cardiol 2008. Available at http://www.ispub.com/ jour-

33. Lipshultz SE, Easley KA, Orav EJ, et al. Left ventricular structure

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34. Martinez-Garcia T, Sobrino JM, Pujol E, et al. Ventricular mass and

per lo Studio Cardiologico dei pazienti affetti da AIDS Investigators.

diastolic function in patients infected by the human immunodeficiency

Cardiac involvement in the acquired immunodeficiency syndrome: a multicenter clinical-pathological study. AIDS Res Hum Retroviruses 1998; 14: 1071–1077. 13. Sani MU, Okeahialam BN, Ukoli CO. Electrocardiographic abnormalities in Nigerian AIDS patients. Cardiologie Tropicale 2004; 30: 3–6. 14. Andrew C, Birgit G, Jacqueline N, et al. Strategies for Management of Antiretroviral Therapy (SMART) study. AIDS 2008; 22: 257–267.

virus. Heart 2000; 84: 620–624. 35. Samaan SA, Foster A, Raizada V, et al. Myocardial atrophy in acquired immunodeficiency syndrome-associated wasting. Am Heart J 1995; 130: 823–827. 36. Lund KC, Wallace KB. Direct effects of nucleoside reverse transcriptase inhibitors on rat cardiac mitochondrial bioenergetics. Mitochondrion 2004; 4: 193–202.

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37. Barbaro G, Di Lorenzo G, Grisorio B, Barbarini G. Incidence of

43. Villa A, Foresti V, Confalonieri F. Autonomic neuropathy and prolon-

dilated cardiomyopathy and detection of HIV in myocardial cells of

gation of QT interval in human immunodeficiency virus infection. Clin Auton Res 1995; 5: 48–52.

HIV-positive patients. N Engl J Med 1998; 339: 1093–1099. 38. Meng Q, Lima JA, Lai H, et al. Use of HIV protease inhibitors is asso-

44. Kocheril AG, Bokhari SAJ, Bastsford WP, Sinusas AJ. Role of antiret-

ciated with left ventricular morphologic changes and diastolic dysfunc-

roviral treatment in prolongating QTc interval in HIV positive patients. J Infect 2007; 54(6): 597–602.

tion. J AIDS 2002; 30; 306–310. 39. Pewsner D, Jüni P, Egger M, et al. Accuracy of electrocardiography

45. Alessandra F, Nicola P, Angelo DS, et al. QTc interval prolongation

in diagnosis of left ventricular hypertrophy in arterial hypertension:

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systematic review. Br Med J 2007; 335: 711. 40. Devereux RB. Is the electrocardiogram still useful for detection of left

46. Anson BD, Weaver JG, Ackerman MJ, et al. Blockade of HERG channels by HIV protease inhibitors. Lancet 2005; 365: 682–686.

ventricular hypertrophy? Circulation 1990; 81: 1144–1146. 41. Michael AB. Left ventricular hypertrophy: An overlooked cardiovascu-

47. Eisenhauer MD, Eliasson AH, Taylor AJ, et al. Incidence of cardiac

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arrhythmias during intravenous pentamidine therapy in HIV-infected patients. Chest 1994; 105: 389–395.

42. Ogunmodede A. Electrocardiographic and echocardiographic abnormalities in HIV patients seen at University of Ilorin Teaching Hospital,

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Gender differences in the control of multiple cardiovascular disease risk factors in type 2 diabetes patients Adjusted mean diastolic blood pressure levels were found to be significantly higher in women compared to men, but other risk factors were almost the same between genders, according to recent research. Williams and colleagues conducted a cross-sectional study in which patients were randomly recruited from three primary care clinics in the south-eastern USA and asked to complete a self-report survey yielding data relevant to gender differences in cardiovascular disease (CVD) risk-factor control. The primary outcomes were individual diabetes-related risks, which were defined as not having an HbA1c level < 7%, blood pressure of < 130/80 mmHg, and low-density lipoprotein (LDL) cholesterol level < 100 mg/dl (2.59 mmol/l), and composite control defined as having all three outcomes under control simultaneously. Of the patients enrolled, 56% were men, 67% were non-Hispanic black, and 78% made less than $35 000 per year. Unadjusted mean systolic blood pressure (134 vs 13 mmHg, p = 0.005) and LDL cholesterol levels [99.7 vs 87.6 mg/dl (2.58 vs 2.27 mmol/l), p < 0.001] were much higher in women than in men; however, after adjusting for relevant confounders, differences in systolic blood pressure and LDL cholesterol levels were not significant. Adjusted mean diastolic blood pressure levels were found to be significantly higher in women compared to men (β = 3.09, 95% CI = 0.56–5.63). Regarding the gender differences in composite control, the

results showed that women had poorer control of multiple CVD risk outcomes than men (β = 2.90, 95% CI = 1.37–6.13). Other primary outcomes were not statistically significantly different, including glycaemic control in both genders. Limitations of this study included the fact that the crosssectional study design did not prove causal associations. Also, confounders not controlled for included diabetes knowledge, self-management practices, medication adherence, co-morbidity burden, social support, duration of diabetes, medications used to treat diabetes, and hypertension. In addition, high triglyceride level was an independent risk factor for coronary heart disease, particularly for women. In conclusion, further study is needed. In the meantime, both genders, but especially women, need to be encouraged to adopt healthy lifestyle habits with a view to modifying their risk factors and achieving better outcomes.

References 1.

Williams J, Lunch C, Winchester R, et al. Gender differences in composite control of cardiovascular risk factors among patients with type 2 diabetes. Diabetes Technol Therapeut 2014; 16(7): 421–427.

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Comparison of primary repair and patch plasty procedure on the P wave in adult atrial septal defect closure Alper Ucak, Veysel Temizkan, Murat Ugur, Ahmet Erturk Yedekci, Omer Uz, Arif Selcuk, Ahmet Turan Yilmaz

Abstract Introduction: In this study we compared the effects of two different surgical procedures for closure of adult atrial septal defect (ASD) on postoperative P-wave changes. Methods: Patients who underwent cardiac surgery for secundum type ASD closure were evaluated retrospectively. Seventy-two patients with primary repair of ASD and 29 patients with pericardial patch plasty repair were compared according to Pmax, Pmin and P-wave dispersions (Pd). Results: In each group, the increases in postoperative maximum P-wave duration (Pmax) and minimum P-wave duration (Pmin) were statistically significant. There was no statistically significant difference between post- and pre-operative Pd values. In the comparison between group 1 and group 2 in terms of postoperative P-wave changes (Pmax, Pmin, Pd) there was no statistically significant difference. Conclusion: Comparing patch plasty and primary repair for the surgical closure of ASD in the early to mid-postoperative period, no difference was found and both surgical procedures can be performed in adult ASDs. Keywords: atrial septal defect, P-wave analysis, arrhythmia Submitted 17/12/15, accepted 17/2/16 Published online 4/5/16 Cardiovasc J Afr 2016; 27: 258–261

www.cvja.co.za

DOI: 10.5830/CVJA-2016-013

Atrial septal defect (ASD) is one of the most common congenital heart defects in adulthood. It can be repaired percutaneously or surgically, depending on the defect size. Primary surgical repair or patch plasty (closure with patch) are the two surgical ASD closure techniques used depending on diameter of the defect. Department of Cardiovascular Surgery, GATA Haydarpasa Training Hospital, Istanbul, Turkey Alper Ucak, MD, dralperucak@gmail.com Veysel Temizkan, MD Murat Ugur, MD Arif Selcuk, ND Ahmet Turan Yilmaz, MD

Department of Anesthesiology, Kyrenia Military Hospital, Cyprus Ahmet Erturk Yedekci, MD

Department of Cardiology, GATA Haydarpasa Training Hospital, Istanbul, Turkey Omer Uz, MD

Atrial arrhythmias may develop in repaired and unrepaired ASD patients. The inter-atrial conduction pathway may influence this, and postoperative arrhythmias may develop due to increased size of the right atrium in unrepaired patients, or tension in the suture line in the postoperative period.1 Prolonged maximum P-wave duration (Pmax) and increased P dispersion (Pd) may be pioneer indicators of disturbance of the inter-atrial conduction pathway and atrial fibrillation in ASD patients.2 Many different studies have been carried out for P-wave changes following ASD closure surgeries. These studies evaluated the effects of percutaneous or surgical closure techniques on the P wave after defect repair.3-6 In the literature, there is no study investigating the effects of surgical ASD closure techniques on the P wave in adults. In our study, we retrospectively investigated the electrocardiograms of adult patients who underwent surgical closure of ASDs and evaluated the effects of both primary closure and pericardial patch plasty techniques on the P wave.

Methods The study design was approved by the institutional review board. Patients who underwent surgical ostium secundum type ASD closure between the years 2004 and 2014 in the cardiovascular surgery clinic of the GATA Haydarpasa Training Hospital were included in the study. Patients with primum ASD and patients with cardiac pathologies requiring additional surgical treatment were excluded from the study. Twelve-lead surface electrocardiograms were collected from patients’ records who underwent surgical ostium secundum type ASD closure. The 101 patients were divided into two groups (Table 1) according to their closure procedure; primary surgical repair (group 1) and pericardial patch plasty (group 2). Seventy-two patients without increased atrial stretch and with a small- to medium-sized defect diameter were evaluated as suitable for primary surgical repair (group 1) and they underwent primary surgical repair for ASD closure. Twenty-nine patients with a larger defect diameter and/or patients with accompanying sinus venosus type ASD (group 2) underwent the patch plasty technique for ASD closure. The demographic data of the patients in each group and their pre- and postoperative five- to seven-day and three-month 12-lead surface electrocardiogram P waves were compared. All surgeries were carried out under general anaesthesia. A median sternotomy, mini-thoracotomy and mini-sternotomy were performed in 69, 14 and 18 patients, respectively (Table 2). Cardiopulmonary bypass was achieved by cannulation of the aorta and double venous cannulation in the right atrium following median sternotomy, where femoral arterial cannulation and femoral vein to selected superior vena cava cannulation were achieved following mini-thoracotomy/mini-sternotomy.

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Table 1. Comparison of pre-operative patients’ characteristics Demographics

Table 3. P-wave changes in primary repair procedure for ASD closure

Group 1 (n = 72) Group 2 (n = 29)

Age (years)

21.8 ± 2.4

22.3 ± 3.7

BMI (kg/m2)

25.2 ± 1.2

24.7 ± 1.3

Diameter of defect (mm)

20.6 ± 8.5

23.9 ± 10.3

Qp/Qs

2.0 ± 0.7

2.3 ± 0.9

EF (%)

60.9 ± 6.2

64.3 ± 5.2

PASB (mmHg)

30 ± 10.7

32.3 ± 11.8

≤ Mild tricuspid regurgitation (n)

≤ Mild pulmonary regurgitation (n)

Anomalous pulmonary venous return (n)

≤ Mild pulmonary stenosis (n)

≤ Mild aortic regurgitation (n)

BMI: body mass index, EF: ejection fraction, PASB: pulmonary artery systolic pressure.

Cardiopulmonary bypass was established and following cardioplegic arrrest, 32°C blood–body temperature was provided. Following right atriotomy, the defect was evaluated in the presence of pre-operative echocardiographic findings. In group 1, the defect was closed with a primary continuous suture technique with 4/0 prolene. In group 2, the defect was closed by means of a fresh autologous pericardial patch using a continuous suture technique with 4/0 prolene. Pre-operative, fifth- and seventh-day postoperative and threemonth 12-lead surface electrocardiograms were provided from patients’ records, which were obtained at a paper speed of 50 mm/s with 1-mV/cm standardisation. Electrocardiograms were scanned for evaluation of the P waves. Electrocardiograms exhibiting P waves at least nine in 12 derivations were analysed for the existing P waves.4 Measuring the length of the P waves using Photoshop® (Adobe), the longest P wave was denoted as Pmax whereas the shortest P wave was Pmin. The difference between Pmax and Pmin was the P-wave dispersion (Pd) (Pd = Pmax – Pmin).

Statistical analysis The 5.0 version of the GraphPad Prism program was used for statistical analysis. Data are shown as mean ± standard deviation. Postoperative P-wave changes of the patients in group 1 and 2 were compared with pre-operative values and the differences were evaluated. Continuous variables were compared using Mann–Whitney U- and Student’s t-tests. A p-value ≤ 0.05 was considered statistically significant.

Results Pre-operative demographic characteristics of the two groups were similar. Demographic characteristics and transthoracic Table 2. Comparison of surgical data Surgery

Mini-sternotomy (n)

CPB time (min)

38.5 ± 12.2

42.4 ± 16.6

Cross-clamp time (min)

20.3 ± 7.6

23.3 ± 10.6

Revision (n) Hospital stay (days) CPB: cardiopulmonary bypass.

Postoperative 5th day

Postoperative 3rd month

Pmax

205.9 ± 29.4

220.6 ± 31.5

231.1 ± 39.4

0.0033

0.0001

Pmin

108.1 ± 29.4

121.2 ± 32.7

129.5 ± 36.9

0.0162

0.0003

97.2 ± 33.1

98.8 ± 35.9

101.7 ± 42.2

0.7011

0.4432

p1: comparison of the pre-operative period and the fifth day postoperatively; p2: comparison of the pre-operative period and the third month postoperatively.

echocardiographic data of our patients are shown in Table 1. All of the patients were in sinus rhythm in the pre-operative period. Arrhythmia was not observed in the postoperative follow up and there was no need for pace implementation. The patients were discharged 6.2 ± 1.4 days postoperatively. In group 1, compared to the pre-operative period, Pmax was significantly increased in the five to seven days postoperatively, and Pmax was still significantly longer three months after the procedure (Table 3). Pmax was also increased in the postoperative period in group 2 but this change gained statistical significance at three months following the procedure (Table 4). In the evaluation of Pmin, compared to the pre-operative period, Pmin was significantly increased in the five to seven days postoperatively and three months after the procedure in both groups (Tables 3, 4). Evaluation of the P-wave dispersion revealed that in group 1, compared to the pre-operative period, no significant difference was found in the five to seven days postoperatively and three months after the procedure. Similarly, in group 2, compared to the pre-operative period, no significant difference was found in any postoperative follow-up periods. No statistically significant difference was found in a comparison of group 1 and 2, both pre-operatively and at the postoperative follow up in terms of P-wave analysis (Pmax, Pmin and Pd) (Table 5).

Discussion The presence of ASD causes volume overload and increased stretch induces right heart dilatation and dysfunction. In these patients, Pmax and Pd extension develops as a result of prolongation of the atrial depolarisation time.2,4,7 The prolongation of Pd and Pmax reflect non-homogeneous and discontinuous sinus stimulation and may be predictors of atrial fibrillation.2 In the literature, P-wave changes have been compared in repaired and unrepaired ASDs, or in repaired ASDs by surgical or percutaneous means. There is no study comparing the effects of two different surgical techniques on the P wave. In our study, two different surgical ASD closure techniques were compared in terms of P-wave analyses using primary repair and pericardial patch plasty techniques. The incidence of dysrhythmia increases with increasing age in patients with unrepaired ASD.8 Additionally, the risk

Group 1 (n = 72) Group 2 (n = 29)

Median sternotomy (n) Mini-thoracotomy (n)

Pre-operative

1 6.2 ± 1.4

0 6.4 ± 1.4

Table 4. P-wave changes in pericardial patch plasty procedure for ASD closure Postoperative Postoperative P2 Pre-operative 5th day 3rd month P1 0.1092 0.0089 Pmax 219.4 ± 37.7 236.1 ± 39.4 254.3 ± 51.1 0.06 0.043 Pmin 107.1 ± 28.7 120.3 ± 27.7 132.8 ± 35.6 0.5659 0.2796 Pd 110.6 ± 43.6 115.9 ± 39.9 121.7 ± 47.9 p1: comparison of the pre-operative period and the fifth day postoperatively; p2: comparison of the pre-operative period and the third month postoperatively.

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Table 5. Comparison of primary repair procedure and pericardial patch plasty procedure for ASD closure Pre-operative Pmax Pmin Pd Postoperative 5th day Pmax Pmin Pd Postoperative 3rd month Pmax Pmin Pd

Primary repair

Patch plasty

p-value

205.9 ± 29.4 108.1 ± 29.4 97.2 ± 33.1

219.4 ± 37.7 108.8 ± 28.7 110.6 ± 43.6

0.1112 0.9436 0.1742

220.6 ± 31.5 121.2 ± 32.7 98.8 ± 35.9

236.1 ± 39.4 120.3 ± 27.7 115.9 ± 39.9

0.5805 0.983 0.0929

231.1 ± 39.4 129.5 ± 36.9

254.3 ± 51.1 132.7 ± 35.6

0.0639 0.6696

101.7 ± 42.2

121.7 ± 47.9

0.0711

of arrhythmia increases due to right atrial volume overload and atrial remodelling related to atrial hypertrophy.3 One of the indications of ASD closure is to avoid arrhythmia, which increases the risk of morbidity and mortality.2,7 The risk of atrial arrhythmia decreases with recovery of right atrial dilatation and electrophysiological changes after surgical closure of the ASD.9 In our study, we compared the effects of two surgical techniques on atrial conduction pathways and remodelling caused by inflammatory fibrosis, by analysis of the P wave. In both primary repair and patch plasty groups, Pmax and Pmin values increased but Pd values did not change. Increase in Pmax and Pmin values in the third month may have been related to early remodelling, however because of the unchanged Pd, we believe that the risk of arrhythmia will not increase in the long term. In the postoperative period, the Pd value decreased in patients who had regressed right atrial dilatation. However, Pmax and Pd were longer in patients with persistent atrial dilatation.10 From the results of Fang et al.,10 longer Pd values in patients with permanent atrial dilatation support the notion that Pd may be used as a non-invasive parameter for prediction of atrial arrhythmia. Our findings showed that the risk of atrial arrhythmia was similar in both primary closure and patch plasty techniques, since there was no difference between Pd values. By contrast, Thilen et al.11 declared that increased P-wave duration did not decrease with surgical repair in older patients and it was not related to atrial dilatation. In haemodynamically significant ASD, the increase in P-wave duration may depend on regional damage of the atrial conduction pathways rather than atrial enlargement.12,13 In our study, we were able to obtain three-month follow-up records of the patients. During the follow up, although Pmax values had increased, Pd values did not change and there was no arrhythmia. From our results, we believe that in repair of ASD in adults, the Pd value could be a more meaningful predictor for the risk of arrhythmia. The age of the patient at ASD closure may be a risk factor for the development of arrhythmia in the follow-up period.11,14 ASD repair in childhood prevents permanent changes in the atrial myocardium and regresses Pmax and Pd, thus decreasing the risk for atrial fibrillation in older patients.2,15 In adult patients, the chance of returning to normal atrial size is lower with ASD repair. Repair of ASD after 25 years of age is a risk factor for the development of atrial fibrillation in the long term.14,16

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In our study we investigated 101 patients who had had a diagnosis of ASD. Ninety of them were younger than 25 years and their mean age was 21.9 ± 2.8 years. None of the patients had arrhythmia before the surgery and none had atrial arrhythmia during the three-month follow-up period. In our patients, the Pd value did not change after surgical repair and the rate of returning normal atrial size may have been higher since most of our patients were young adults. The ASD closure technique did not affect the P wave in a comparison of different percutaneous techniques, or surgical repair and percutaneous techniques in previous studies.3,11 Javadzadegan et al.3 suggested that in both percutaneous ASD closure and surgical treatment, the P-wave duration was decreased at six months’ follow up and this decrease was not related to the defect size. In comparison, surgical and transcatheter ASD closure, Baspınar et al.7 declared that in the surgical group, decrease in the Pd was more meaningful in the early period. We compared primary closure and patch plasty techniques as two different surgical techniques, to analyse the effects on the P wave, and there were no differences between the two surgical techniques. When considering the anatomy of the right atrial electrophysiological conduction pathway, there are no significant conduction pathways into the atrial septum. We believe that different surgical closure techniques do not affect the Pd value or cause changes. In long-term follow up after ASD closure, increase in Pd values could be a sign of atrial arrhythmia.2 In ASD repair of young adults, there was no increase in Pd values following ASD closure. Neither primary repair nor patch plasty techniques had any effect on the P-wave length or dispersion. Both surgical techniques can therefore be performed, depending on the defect anatomy and size. There are limitations to this study. The majority of patients were young and they were in the second and third decades of their lives. After three months’ postoperative period, we had limited access to long-term follow-up records, because they were carried out in their homelands. We need long-term results to assess whether the increased Pmax and Pd values in the first three months continue in the long term and whether these changes led to arrhythmias in the follow-up period.

Conclusion In this study we compared patch plasty and primary repair for the surgical closure of ASD in the early to mid-postoperative period. No differences were found between the methods in terms of postoperative P-wave changes, and we concluded that both surgical procedures can be performed in adult ASDs.

References 1.

Berger F, Vogel M, Kretschmar O, Dave H, Prêtre R, Dodge-Khatami A. Arrhythmias in patients with surgically treated atrial septal defects. Swiss Med Wkly 2005; 135: 175–178.

Guray U, Guray Y, Mecit B, Yilmaz MB, Sasmaz H, Korkmaz S. Maximum p wave duration and p wave dispersion in adult patients with secundum atrial septal defect: the impact of surgical repair. Ann Noninvasive Electrocardiol 2004; 9: 136–141.

Javadzadegan H, Toufan M, Sadighi AR, Chang JM, Nader ND. Comparative effects of surgical and percutaneous repair on P-wave and

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atrioventricular conduction in patients with atrial septal defect – ostium 4.

10. Fang F, Luo XX, Lin QS, Kwong JS, Zhang YC, Jiang X, et al.

secundum type. Cardiol Young 2013; 23: 132–137.

Characterization of mid-term atrial geometrical and electrical remod-

Pac FA, Balli S, Topaloglu S, Ece I, Oflaz MB. Analysis of maximum

eling following device closure of atrial septal defects in adults. Int J Cardiol 2013; 168: 467–471.

P-wave duration and dispersion after percutaneous closure of atrial septal defects: comparison of two septal occluders. Anadolu Kardiyol 5.

11. Thilen U, Carlson J, Platonov PG, Olsson SB. Atrial myocardial

Derg 2012; 12: 249–254.

pathoelectrophysiology in adults with a secundum atrial septal defect is

Ozyilmaz I, Ozyilmaz S, Tola HT, Saygi M, Kiplapinar N, Tanıdır C,

unaffected by closure of the defect. A study using high resolution signal-

et al. Holter electrocardiography findings and P-wave dispersion in

averaged orthogonal P-wave technique. Int J Cardiol 2009; 132: 364–368.

pediatric patients with transcatheter closure of atrial septal defects. Ann

12. Morton JB, Sanders P, Vohra JK, Sparks PB, Morgan JG, Spence SJ, et

Noninvasive Electrocardiol 2014; 19: 174–181. 6.

al. Effect of chronic right atrial stretch on atrial electrical remodeling in patients with an atrial septal defect. Circulation 2003; 107: 1775–1782.

Johnson JN, Marquardt ML, Ackerman MJ, Asirvatham SJ, Reeder GS, Cabalka AK, et al. Electrocardiographic changes and arrhythmias

13. Thilen U, Carlson J, Platonov PG, Havmöller R, Olsson SB. Prolonged

following percutaneous atrial septal defect and patent foramen ovale

P wave duration in adults with secundum atrial septal defect: a marker

device closure. Catheter Cardiovasc Interv 2011; 78: 254–261.

of delayed conduction rather than increased atrial size? Europace 2007; 9(Suppl 6): 105–108.

Baspinar O, Sucu M, Koruk S, Kervancioglu M, Ustunsoy H, Deniz H, et al. P-wave dispersion between transcatheter and surgical closure of

14. Murphy JG, Gersh BJ, McGoon MD, Mair DD, Porter CJ, Ilstrup DM,

secundum-type atrial septal defect in childhood. Cardiol Young 2011;

et al. Long-term outcome after surgical repair of isolated atrial septal defect. Follow-up at 27 to 32 years. N Engl J Med 1990; 323: 1645–1650.

21: 15–18. 8.

Berger F, Vogel M, Kramer A, Alexi-Meskishvili V, Weng Y, Lange PE,

15. Yavuz T, Nisli K, Oner N, Dindar A, Aydogan U, Omeroglu RE, et

et al. Incidence of atrial flutter/fibrillation in adults with atrial septal

al. The effects of surgical repair on P-wave dispersion in children with secundum atrial septal defect. Adv Ther 2008; 25: 795–800.

defect before and after surgery. Ann Thorac Surg 1999; 68: 75–78. 9.

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Gatzoulis MA, Freeman MA, Siu SC, Webb GD, Harris L. Atrial

16. Oliver JM, Gallego P, González A, Benito F, Mesa JM, Sobrino JA.

arrhythmia after surgical closure of atrial septal defects in adults. N Engl

Predisposing conditions for atrial fibrillation in atrial septal defect with

J Med 1999; 340: 839–846.

and without operative closure. Am J Cardiol 2002; 89: 39–43.

Diabetes is often overlooked after a myocardial infarction At least 10% of people who have a myocardial infarction (MI) may also have undiagnosed diabetes. Yet many doctors fail to look for diabetes in these patients, a recent study has found. Dr Suzanne Arnold, assistant professor at Saint Luke’s Mid-America Heart Institute and the University of Missouri in Kansas City, and her team studied data from 2 854 patients who experienced an MI and had never been diagnosed with type 2 diabetes. The study tested the patients’ HbA1c levels. It revealed that doctors often failed to recognise and begin treating diabetes in patients who have experienced MIs with no prior history of diabetes, even when the patient tested positive for diabetes. The researchers found that 287 or 10.1% of the patients who experienced MIs tested positive for diabetes. Out of the 287 patients who tested positive for diabetes, less than one-third received education or medication when discharged from hospital. According to the results, doctors failed to recognise diabetes in 198 or 69% of the previously undiagnosed patients. The researchers noted that when a patient’s HbA1c test results were checked while they were being treated for their MI, there was a 17-fold greater chance that the diabetes would be diagnosed. In a press release, Dr Arnold stated, ‘Diagnosing diabetes

in patients who have had a heart attack is important because of the role diabetes plays in heart disease. By recognising and treating diabetes early, we may be able to prevent additional cardiovascular complications through diet, weight loss and lifestyle changes, in addition to taking medications. Another important reason to diagnose diabetes at the time of heart attack is that it can guide the treatments for the patient’s coronary artery disease.’ According to Dr Arnold and her team, two in three patients with diabetes die from heart-related conditions. Patients with diabetes experience a significantly higher risk for MI. The authors concluded that people who have an MI should ask for a diabetes test if they present with other risk factors such as being overweight, having high blood pressure or a family history of diabetes. This study was presented on 3 June at the American Heart Association’s Quality of Care and Outcomes Research Scientific Sessions 2014.

Reference 1.

http://www.diabetesincontrol.com/articles/diabetes-news/16453diabetes-often-times-overlooked-after-heart-attack.

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Carotid characteristics of black South Africans with five-year sustained hypertension Melissa Maritz, Carla MT Fourie, Johannes M van Rooyen, Hugo W Huisman, Aletta E Schutte

Abstract Introduction: An important feature of hypertension is a reduction in large artery distensibility, which may be due to structural and functional adaptations. Black populations are particularly prone to the development of hypertension. We therefore compared the carotid characteristics between fiveyear sustained hypertensive and normotensive black South Africans, and investigated how carotid characteristics relate to cardiometabolic risk factors, inflammation, endothelial activation and health behaviours. Methods: We included HIV-free black South Africans who were either consistently hypertensive (n = 351) or normotensive (n = 241) from 2005 to 2010. We assessed carotid characteristics, including intima–media thickness (IMT), distensibility and lumen diameter with B-mode ultrasound, and calculated Young’s elastic modulus, cross-sectional wall area and beta-stiffness index. We measured the carotid dorsalis pedis pulse-wave velocity, brachial and central systolic blood pressure (cSBP) and determined metabolic, inflammatory and endothelial activation markers from blood samples. Health behaviours were reported in questionnaires. Results: The hypertensive group presented with higher brachial and central blood pressure, thicker IMT and stiffer carotid arteries (all p < 0.001). However, after adjustment for cSBP but not mean arterial pressure (MAP), all significant differences in carotid characteristics were lost. The carotid thickness measurements did not differ after adjustment for MAP. After adjustment, metabolic, inflammatory and endothelial activation markers did not differ between the two groups. Conclusion: Our results suggest that besides structural changes, functional adaptations are also involved in deterioration of the carotid wall characteristics of hypertensive black South Africans. These results highlight the importance of proper hypertension control in Africa. Keywords: ethnicity, large artery, stiffness, distensibility, hypertension, central pressure Submitted 30/10/15, accepted 5/5/16 Cardiovasc J Afr 2016; 27: 262–269

www.cvja.co.za

DOI: 10.5830/CVJA-2016-059

Hypertension in Africa Research Team (HART), North-West University, Potchefstroom, South Africa Melissa Maritz, MSc Carla MT Fourie, PhD, carla.fourie@nwu.ac.za Johannes M van Rooyen, DSc Hugo W Huisman, PhD Aletta E Schutte, PhD

MRC Research Unit for Hypertension and Cardiovascular Disease, North-West University, Potchefstroom, South Africa Aletta E Schutte, PhD

Worldwide, cardiovascular disease is the leading cause of death.1 Arterial stiffness is implicated in the development of cardiovascular disease, which results in stroke, coronary heart disease and heart failure.2,3 Arterial stiffness refers to the reduced ability of an artery to expand and recoil with pressure changes, and the arterial distensibility is a measure of vessel stiffness.4 Decreased distensibility may raise central systolic blood pressure (cSBP) and consequently decrease the amplification of pulse pressure.5,6 This results in inadequate coronary perfusion, increased afterload on the heart, as well as an increased pulsatile load on the microcirculation.3 The artery is able to resist strain during blood pressure increases via recruitment of collagen fibres in the arterial wall,7 but sustained high pressure predisposes the vessel to progressive changes in the wall shape and composition, ultimately leading to several clinical complications such as arterial fibrosis and stiffening.8,9 Recently, van Sloten et al. reported that in a European population, carotid artery stiffness independently predicts incident cardiovascular (CV) events and all-cause mortality.3 The higher incidence of stroke, heart failure and renal failure in black populations are a consequence of the higher prevalence of hypertension,10,11 and arterial stiffness12,13 in black compared to white populations. Despite the high prevalence of hypertension and stroke among black South Africans,14 there is limited knowledge on carotid wall properties in this population. We therefore compared the characteristics of the carotid arteries between normotensive and five-year sustained hypertensive black individuals, along with brachial and central blood pressure and conventional cardiometabolic risk factors, markers of inflammation, endothelial activation and health behaviours.

Methods This sub-study forms part of the South African leg of the multi-national Prospective Urban and Rural Epidemiology (PURE) study.15 The participants of the PURE-SA study were from urban and rural localities in the North West Province, and baseline data collection took place in 2005 (n = 2 010 participants), while follow-up data was collected five years later, in 2010 (n = 1 288 participants). For this sub-study we included only the HIV-free black participants with two consecutive (2005, 2010) blood pressure measurements in either the hypertensive or normotensive range (n = 592 participants), consisting of a group of five-year sustained normotensive (n = 241participants) and hypertensive (n = 351 participants) black South Africans (Fig. 1). The Health Research Ethics Committee of the North-West University approved the protocol of the PURE-SA study, as well as this sub-study (ethics number: NWU-00016-10-A1), and it complies with the Declaration of Helsinki. Participants completed structured demographic, lifestyle and physical

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activity questionnaires with the assistance of trained African field workers from the communities. Before measurements commenced, all procedures were explained to the participants in their home language. The participants then gave written informed consent. Height (Invicta stadiometer IP 1465, Leicester, UK), weight (Precision health scale, A & D Company, Tokyo, Japan) and waist circumference (WC) (Holtain unstretchable metal tape, Apex Tool Group, Apex, USA) were measured using standardised methods and calibrated instruments.16 Body mass index (BMI) was calculated with the formula: weight (kg)/height (m2). The validated OMRON HEM-757 (Omron Healthcare, Kyoto, Japan) device was used to measure blood pressure. Each participant was fitted with the correct cuff size. During the measurements, the participant was seated in a relaxed upright position with legs uncrossed. After a resting period of 10 minutes, the brachial systolic (bSBP) and diastolic blood pressure (bDBP) were measured on the right upper arm, followed by a five-minute resting period and a second blood pressure measurement. The value of the second measurement was used for analysis. Participants were classified as hypertensive or normotensive according to standard guidelines.17 The cSBP was measured with the OMRON 9000AI device (Omron Healthcare, Kyoto, Japan), which uses the second systolic peak (reflected wave) as basis for the calculation of cSBP. Diastolic blood pressure is assumed to be consistent

throughout the body, therefore the central pulse pressure (cPP) was calculated by subtracting the bDBP from the cSBP.18,19 Pulse-wave velocity (PWV) was measured non-invasively on the left side of each participant while in a supine position. The Complior SP device (Artech-Medical, Pantin, France) uses superficial pulses over the carotid dorsalis pedis (cdPWV) section of the arterial tree to estimate PWV. The carotid characteristics were measured non-invasively using B-mode ultrasonography with the SonoSite Micromaxx system (SonoSite, Inc., Bothel, WA, USA), using a six- to 13-MHz linear array transducer. A minimum of two optimal angles were used from images of the right and left common carotid arteries. These images were then measured by a single reader according to protocols, and digitised and imported into the automated software of the Artery Measurements System (Gothenburg, Sweden). The carotid intima–media thickness (IMT) was analysed by a single reader on a good-quality image of a maximum 10-mm segment. The borders of the inner diameter of the blood vessel and the near and far wall of the intima–media were identified by an automated function of the program, but user intervention was possible. Approximately 100 discrete measurement points within a 10-mm segment of the carotid artery were used to obtain the mean IMT and the carotid diameter for each participant. The carotid cross-sectional wall area (CSWA) was calculated according to the following formula: LD

___ ___ ___ CSWA = [3.14 × ( ___ 2    + IMTf) × (  2    + IMTf)] – [3.14 × (  2   )  × (  2   ) ]

Normotensive participants 2005 (n = 520)

Remained normotensive 2005–2010 (n = 241)

Excluded hypertensive participants 2010 (n = 191) Excluded missing BP data 2010 (n = 18) Excluded medication use (statins, anti-inflammatory, anti-hypertensive) 2010 (n = 70) Hypertensive participants 2005 (n = 541)

Sustained hypertensive 2005–2010 (n = 351)

Excluded normotensive participants 2010 (n = 170) Excluded missing BP data 2010 (n = 20)

Fig. 1. S tudy population; all participants were HIV-free. BP, blood pressure.

where LD = lumen diameter and IMTf = carotid intima–media thickness of the far wall. A single reader analysed video-clips of the carotid artery in order to determine the maximum and minimum lumen diameters (LD). The carotid distensibility (CD) coefficient was calculated according to the following formula: [(2 × delta LD × min LD) + (delta LD)2)

_______________________________     CD =       (cPP × min LD2]

where LD = lumen diameter, delta LD = maximum LD – minimum LD, min LD = minimum LD, cPP = central pulse pressure.20 Young’s elastic modulus was calculated according to the following formula: min LD

_________________     Young’s elastic modulus =     (IMT × distensibility)

where IMT = carotid intima–media thickness, min LD = minimum lumen diameter.21 The beta-stiffness index was calculated according to the following formula: ln (cSBP/dDBP)

______________     Beta-stiffness index =     delta LD/min LD

where delta LD = maximum LD – minimum LD, min LD = minimum LD.22 Participants were required to fast for at least eight hours and blood samples were taken from the ante-brachial vein with a sterile winged infusion set and syringes. The preparation of the serum and plasma was done according to standardised methods, snap frozen on dry ice and stored in the laboratory at –80°C. In the case of blood collection in a rural area, serum and plasma were snap frozen and stored at –20°C for not more than five days. The serum was then transported to the laboratory and stored at –80°C for further analysis.

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The Cobas Integra 400 (Roche® Clinical System, Roche Diagnostics, Indianapolis, IN) was used to assess the quantitative aspect of total cholesterol (TC), high-density lipoprotein cholesterol (HDL-C), triglycerides (TG), gamma-glutamyltransferase (GGT), creatinine and high-sensitivity C-reactive protein (hsCRP) levels in the serum samples, and glucose levels in fluoride plasma samples. Glycosylated haemoglobin (HbA1c) levels were determined from EDTA whole blood with ion-exchange high-performance liquid chromatography (D-10 haemoglobin testing system, Bio-Rad #220-0101). The Friedewald formula was used to calculate the quantitative aspect of low-density lipoprotein cholesterol (LDL-C).23 The estimated creatinine clearance (CrCl) rate was calculated with the Cockcroft–Gault formula.24 We determined serum interleukin-6 (IL-6) levels with an electro-chemiluminescence immunoassay (Cobas e411 analyzer, Roche, Basel, Switzerland). Serum intercellular adhesion molecule 1 (sICAM-1) and vascular cell adhesion molecule 1 (sVCAM1) concentrations were assessed by sandwich ELISAs (human sICAM-1 and human sVCAM-1 assay, IBL, Hamburg, Germany).

Statistical analysis Statistical analyses were performed using Statistica® 12 (StatSoft, Inc, Tulsa, OK, USA). Descriptive statistics, including the mean and standard deviation, were performed on data with a normal distribution. If not normally distributed, the data were logarithmically transformed and presented as the geometric mean and the fifth and 95th percentiles. We used independent t-tests to determine differences between normotensives and hypertensives or chi-squared tests for categorical variables. We compared the groups using ANCOVA while adjusting for age, gender, WC, GGT, tobacco and antihypertensive medication use. We plotted quartiles of cSBP against carotid features, and compared carotid features using ANOVA and ANCOVA while adjusting for cSBP. We used single and linear regression analyses to determine associations between carotid measures and cardiometabolic risk factors, health behaviours, inflammation and endothelial activation. Multiple regression analyses were done to determine independent associations between the carotid characteristics and cardiometabolic risk factors, with the dependent variables including CD and IMT, and CSWA and max LD. The following co-variates were included in the regression model: locality, gender, age, WC, cSBP, heart rate, LDL-C, HbA1c, CrCl, hsCRP, ICAM-1, GGT, tobacco use and additionally for the hypertensive group, anti-hypertensive medication use.

Results The characteristics of normotensive and hypertensive black Africans are shown in Table 1. The hypertensives were older, had a higher BMI and WC, and a larger percentage were from an urban area than the normotensives. All the cardiovascular measurements, as well as carotid characteristics, glycaemic and inflammatory markers, ICAM-1, TG, GGT and self-reported alcohol use were significantly higher in the hypertensive group. After adjustments for age, gender, WC, GGT, tobacco and anti-hypertensive medication use (Table 2), all carotid characteristics except IMT remained significantly different.

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Table 1. Characteristics of normotensive and hypertensive black Africans Normotensive (n = 241)

Hypertensive (n = 351)

Men, n (%)

90 (37.3)

129 (36.8)

0.88

Urban, n (%)

78 (32.4)

178 (50.7)

< 0.001

52.8 ± 8.93

59.0 ± 10.0

< 0.001

Age, years

p-value

Anthropometry Waist circumference, cm

78.7 ± 11.9

84.7 ± 13.5

< 0.001

Body mass index, kg/m2

24.7 ± 6.87

26.6 ± 7.85

0.003

Cardiovascular measures Brachial SBP, mm Hg

119 ± 11.9

157 ± 22.4

< 0.001

Brachial DBP, mm Hg

78.9 ± 7.23

100 ± 12.5

< 0.001

Heart rate, bpm

61.8 ± 14.7

67.8 ± 17.9

< 0.001

Central SBP, mm Hg

116 ± 12.8

150 ± 22.2

< 0.001

Central PP, mm Hg

37.4 ± 11.8

50.9 ± 20.4

< 0.001

Carotid dorsalis pedis PWV, m/s

8.25 ± 1.35

10.0 ± 1.89

< 0.001

Carotid characteristics Distensibility × 10-3, 1/kPa

4.72 ± 2.00

3.02 ± 1.83

< 0.001

Young’s elastic modulus × 103, kPa

2.17 ± 1.07

3.72 ± 2.20

< 0.001

Beta-stiffness index

7.10 ± 2.62

9.37 ± 4.51

< 0.001

Intima–media thickness, mm

0.68 ± 0.13

0.77 ± 0.17

< 0.001

Cross-sectional wall area, mm2

14.2 ± 4.41

17.4 ± 5.30

< 0.001

Lumen diameter maximum, mm

6.18 ± 0.77

6.60 ± 0.85

< 0.001

Lumen diameter minimum, mm

5.74 ± 0.72

6.22 ± 0.83

< 0.001

HDL-C, mmol/l

1.45 ± 0.65

1.51 ± 0.60

0.30

LDL-C, mmol/l

2.86 ± 1.17

2.96 ± 1.14

0.29

1.09 (0.57–2.11)

1.20 (0.59–2.89)

0.039

Lipids

Triglycerides, mmol/l Glycaemia Glucose, mmol/l

4.86 (3.91–6.06)

5.27 (3.96–7.90) < 0.001

HbA1c (%)

5.94 (5.30–6.80)

6.08 (5.20–7.80)

Interleukin-6, pg/ml

3.39 (0.75–23.5)

4.43 (0.75–20.0) < 0.001

C-reactive protein, mg/l

2.96 (0.21–32.6)

3.84 (0.38–28.4)

286 ± 92.7

321 ± 111

< 0.001

732 (442–1388)

762 (443–1735)

0.23

91.4 (53.7–163)

91.6 (47.7–164)

0.94

0.038

Inflammatory markers 0.022

Adhesion molecules Intercellular adhesion molecule-1, pg/ml Vascular adhesion molecule-1, pg/ml Renal function Creatinine clearance, ml/min Health behaviours γ-glutamyl transferase, U/l

33.7 (11.9–167)

52.7 (13.6–347)

< 0.001

Self-reported alcohol intake, n, total (%)

60/220 (27.3)

159/336 (47.3)

< 0.001

Self-reported tobacco use, n, total (%)

117/229 (51.1)

159/341 (46.6)

0.56

Anti-hypertension medication, n, total (%)

–

124/351 (35.3)

Lipid-lowering medication, n, total (%)

–

5/351 (1.42)

Anti-inflammatory medication, n, total (%)

–

21/351 (5.98)

Data are arithmetic means ± SD or geometric mean (fifth and 95th percentile intervals) for logarithmically transformed. n, number of participants; SBP, systolic blood pressure; DBP, diastolic blood pressure; PP, pulse pressure; PWV, pulse wave velocity; HDL-C, high-density lipoprotein; LDL-C, low-density lipoprotein; HbA1c, glycated haemoglobin.

The inflammatory and glycaemic markers, lipids and adhesion molecules did not differ however after the above adjustments. The carotid characteristics plotted against quartiles of cSBP are shown in Fig. 2. All the carotid characteristics changed significantly with increasing cSBP before and after the adjustments.

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Table 2. Adjusted characteristics of normotensive and hypertensive black Africans Normotensive (n = 241)

Hypertensive (n = 351)

p-value

Brachial SBP, mm Hg

120 ± 20.0

155 ± 20.7

< 0.001

Brachial DBP, mm Hg

79.2 ± 11.1

99.8 ± 11.6

< 0.001

Heart rate, bpm

62.7 ± 17.5

66.8 ± 18.0

0.017

Central SBP, mm Hg

117 ± 20.2

148 ± 20.9

< 0.001

Central PP, mm Hg

38.4 ± 17.9

49.4 ± 18.5

< 0.001

Carotid dorsalis pedis PWV, m/s

8.87 ± 2.06

9.37 ± 2.13

< 0.001

Cardiovascular measures

Distensibility (1/kPa × 10–3)

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p-trend <0.001

6.0 4.0 2.0 0.0

<119

Unadjusted

Carotid characteristics Distensibility × 10-3, 1/kPa

4.58 ± 1.98

3.15 ± 2.07

< 0.001

Young’s elastic modulus × 103, kPa

2.26 ± 1.98

3.59 ± 2.07

< 0.001

Beta-stiffness index

7.48 ± 3.98

8.97 ± 4.16

0.002

0.9 0.8

0.72 ± 0.14

0.73 ± 0.18

0.42

14.8 ± 4.71

16.7 ± 4.86

< 0.001

Lumen diameter maximum, mm

6.24 ± 0.85

6.51 ± 0.92

0.005

Lumen diameter minimum, mm

5.80 ± 0.85

6.12 ± 0.76

< 0.001

HDL-C, mmol/l

1.46 ± 0.60

1.50 ± 0.73

0.47

LDL-C, mmol/l

2.93 ± 1.19

2.90 ± 1.27

0.81

1.16 (1.08–1.24)

1.11 (1.05–1.19)

0.50

Lipids

Triglycerides, mmol/l

5.01 (4.86–5.16)

5.10 (4.97–5.23)

0.43

HbA1c, %

6.05 (5.93–6.15)

5.99 (5.89–6.08)

0.44

Interleukin-6, pg/ml

3.62 (3.19–4.11)

4.05 (3.64–4.51)

0.22

C-reactive protein, mg/l

3.41 (2.84–4.08)

3.18 (2.72–3.72)

0.62

293 ± 101

308 ± 105

0.13

748 (706–792)

747 (711–785)

0.98

90.8 (87.8–94.0)

92.3 (89.6–95.0)

0.54

0.5 <119

Vascular adhesion molecule-1, pg/ml Creatinine clearance, ml/min

Data are arithmetic means ± SD or geometric mean (fifth and 95th percentile intervals) for logarithmically transformed. Data are adjusted for age, gender, waist circumference, γ-glutamyl transferase, tobacco use and anti-hypertensive medication use. Pulse-wave velocity and carotid intima–media thickness additionally adjusted for mean arterial pressure. n, number of participants; SBP, systolic blood pressure; DBP, diastolic blood pressure; PP, pulse pressure; HDL-C, high-density lipoprotein cholesterol; LDL-C, low-density lipoprotein cholesterol; HbA1c, glycated haemoglobin.

We finally adjusted for cSBP when comparing the normotensive and hypertensive groups (Table 3), resulting in no significant differences in the carotid characteristics between the groups. When cSBP was substituted with either brachial SBP (p = 0.029) or mean arterial pressure (MAP) (p = 0.0012), the difference in distensibility remained, but the more physical measures such as IMT, CSWA and LD did not differ. In sensitivity analysis, we further compared hypertensives who were not using anti-hypertensive therapy (n = 227) with treated hypertensives (n = 124) and normotensives, applying similar adjustments, including cSBP. We found results similar to those in Table 3 (Table 4). Furthermore, excluding participants on antihypertensive medication and comparing the normotensive group to the treated hypertensive group did not change the results. The types of medication used by the hypertensive participants are shown in Table 5. Table 6 reports the forward stepwise multiple regression analyses performed in the normotensive and hypertensive groups with either CD or IMT as dependent variables. As expected, CD associated with cSBP in both groups (p < 0.001), however, IMT associated independently with cSBP in the hypertensive group only (p = 0.016).

119–133

133–151

>151

Central systolic blood pressure (mmHg) Unadjusted

Adjusted

p-trend <0.001

20 CSWA (mm2)

18 16 14 12 10

<119

119–133

133–151

>151

Central systolic blood pressure (mmHg) Unadjusted

Adjusted

p-trend <0.001

7.0 Max LD (mm)

Intracellular adhesion molecule-1, pg/ml

Adjusted

0.7

0.4

Inflammatory markers

Adhesion molecules

>151

0.6

Glycaemia Glucose, mmol/l

133–151

p-trend <0.001

cIMT (mm)

Intima–media thickness, mm Cross-sectional wall area, mm2

119–133

Central systolic blood pressure (mmHg)

6.0 5.0 4.0 3.0

<119

119–133

133–151

>151

Central systolic blood pressure (mmHg) Unadjusted

Adjusted

Fig. 2. Quartiles of central blood pressure plotted against measures of the carotid artery in the total group (n = 592), unadjusted and adjusted for age, gender, waist circumference, GGT, tobacco and anti-hypertensive medication use. cIMT, carotid intima–media thickness; CSWA, cross-sectional wall area; Max LD, maximum lumen diameter.

Table 7 reports the forward stepwise regression analyses performed in the normotensive and hypertensive groups with either CSWA or maximum LD as the dependent variables. CSWA associated with cSBP (p < 0.001) in the hypertensive group only, whereas maximum LD associated with cSBP in both the normotensive and hypertensive groups.

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Table 3. Carotid characteristics of normotensive and hypertensive black Africans, adjusted for potential confounders Normotensive (n = 241)

Hypertensive (n = 351)

p-value

Carotid characteristics after adjustment for central SBP Distensibility × 10-3, 1/kPa

3.77 ± 1.98

3.95 ± 2.22

0.50

Young’s elastic modulus × 103, kPa

3.04 ± 1.84

2.80 ± 2.05

0.34

Beta-stiffness index

8.43 ± 4.21

8.02 ± 4.61

0.47

Intima–media thickness, mm

0.71 ± 0.14

0.73 ± 0.17

0.19

Cross-sectional wall area, mm2

15.2 ± 4.96

16.2 ± 5.25

0.09

Lumen diameter maximum, mm

6.30 ± 0.82

6.41 ± 1.04

0.34

Lumen diameter minimum, mm

5.85 ± 0.94

6.02 ± 0.89

0.12

Carotid characteristics after adjustment for mean arterial pressure Distensibility × 10-3, 1/kPa

4.38 ± 2.45

3.35 ± 2.66

Young’s elastic modulus × 103, kPa

2.46 ± 2.30

3.39 ± 2.64

Beta-stiffness index

7.13 ± 4.68

9.32 ± 5.20

0.0034 0.0082 < 0.001

Intima–media thickness, mm

0.71 ± 0.14

0.73 ± 0.18

0.42

Cross-sectional wall area, mm2

15.6 ± 5.57

16.0 ± 6.07

0.35

Lumen diameter maximum, mm

6.30 ± 0.97

6.44 ± 1.07

0.32

Lumen diameter minimum, mm

5.86 ± 0.97

6.06 ± 0.92

0.17

Data are arithmetic means ± SD. Data adjusted for age, gender, waist circumference, γ-glutamyl transferase, tobacco and anti-hypertensive medication use. Table 4. Carotid characteristics of normotensive, untreated and treated hypertensive black Africans, adjusted for potential confounders including central systolic blood pressure Untreated Treated hyperhypertensives tensives (n = 227) (n = 124) p-value

Carotid characteristics

Normotensive (n = 241)

Distensibility × 10-3, 1/kPa

3.53 ± 2.10 3.70 ± 1.77 3.47 ± 1.70

0.59

Young’s elastic modulus × 103, kPa 3.28 ± 2.07 3.04 ± 1.74 3.29 ± 1.70

0.44

8.83 ± 4.56 8.42 ± 3.80 8.62 ± 3.67

0.75

Beta-stiffness index Intima–media thickness, mm

0.73 ± 0.14 0.75 ± 0.14 0.73 ± 0.10

0.15

Cross-sectional wall area, mm2

15.9 ± 5.38 16.8 ± 4.35 15.9 ± 4.34

0.09

Lumen diameter maximum, mm

6.34 ± 0.94 6.46 ± 0.83 6.50 ± 0.80

0.48

Lumen diameter minimum, mm

5.89 ± 0.94 6.06 ± 0.83 6.12 ± 0.80

0.17

Data are arithmetic means ± SD. Data adjusted for age, gender, waist circumference, γ-glutamyl transferase, tobacco use and central systolic blood pressure.

Discussion As expected, we found that hypertensive black Africans presented with reduced carotid distensibility when compared to normotensives. In fact, we found significant differences for all carotid wall thickness and distensibility measurements between the hypertensives and normotensives prior to adjustments. However, upon adjustment for cSBP, all differences disappeared. The direct physical measures, such as IMT, CSWA and LD, were similar between the hypertensive and normotensive groups after adjustments for both cSBP and MAP. This similarity suggests that the decreased carotid distensibility and increased carotid crossTable 5. Anti-hypertensive medication use in the hypertensive group

Type of anti-hypertensive medication

Hypertensive participants using medication (n = 124)

Unspecified, n, total, (%)

61/124 (49.2)

Beta-blockers, n, total, (%)

12/124 (9.68)

Anti-adrenergics, n, total, (%)

2/124 (1.61)

Calcium channel blockers, n, total, (%)

30/124 (24.2)

Class 2 ACE inhibitors, n, total, (%)

49/124 (39.5)

Diuretics, n, total, (%)

54/124 (43.5)

n, number of participants. Unspecified: only indicated as high blood pressure pills or hypertension medication.

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Table 6. Forward stepwise multiple regression analyses with carotid distensibility and carotid intima–media thickness as dependent variables Normotensive (n = 241) β (95% CI)

p-value

Hypertensive (n = 351) β (95% CI)

p-value

Distensibility (1/kPa) Adjusted R2

0.27

0.37

Locality (urban)

–0.16 (–0.31– –0.01)

0.031

Age, years

–0.14 (–0.29–0.003)

0.058 –0.20 (–0.35– –0.05)

Waist circumference, cm

0.22 (0.06–0.39)

0.009 0.007

Central SBP, mm Hg –0.44 (–0.59– –0.29) < 0.001 –0.56 (–0.68– –0.45) < 0.001 Heart rate, bpm

–0.10 (–0.25–0.04)

0.16

LDL-C, mmol/l HbA1c (%) CrCl, ml/min ICAM-1, pg/ml

0.11 (–0.04–0.25)

Tobacco use (no/yes) –0.14 (–0.28–0.003)

–0.12 (–0.23– –0.01)

0.030

0.08 (–0.03–0.19)

0.18

–0.14 (–0.26– –0.02)

0.018

–0.10 (–0.28–0.07)

0.24

0.16 0.058

IMT (mm) Adjusted R2

0.25

0.35

Locality (urban)

–0.10 (–0.20– –0.01)

Gender (male)

0.26 (0.13–0.39)

< 0.001

Age, years

0.33 (0.21–0.46)

Waist circumference, cm

0.14 (0.01–0.28)

< 0.001

0.44 (0.34–0.54)

< 0.001

0.033

0.08 (–0.02–0.19)

0.13

0.12 (0.02–0.22)

0.016

Central SBP, mm Hg Heart rate, bpm

–0.08 (–0.20–0.05)

0.21

LDL-C, mmol/l

0.07 (-0.05–0.19)

0.29

HbA1c (%) C-reactive protein, pg/ml

0.16 (0.03–0.30)

0.017

ICAM-1, pg/ml

0.12 (–0.01–0.24)

0.079

γ-glutamyl transferase, U/l

0.030

0.25 (0.15–0.36)

0.15 (0.05–0.25)

0.005

0.07 (–0.03–0.18)

0.17

0.06 (–0.04–0.16)

0.27

–0.08 (–0.18–0.02)

0.12

Tobacco use (no/yes)

–0.08 (–0.18–0.02)

0.11

Anti-hypertension medications (no/yes)

–0.10 (–0.19– –0.003)

0.044

Data expressed as beta-values and 95% confidence intervals, p-values obtained with forward stepwise multiple regression analyses. Included in each model: locality, age, gender, waist circumference, heart rate, cSBP, LDL-C, HbA1c, C-reactive protein, ICAM-1, creatinine clearance, γ-glutamyl transferase, tobacco and anti-hypertensive medication use. IMT, carotid–intima media thickness; CSWA, cross-sectional wall area; Max LD, maximum lumen diameter; SBP, systolic blood pressure; LDL-C, low-density lipoprotein cholesterol; HbA1c, glycated haemoglobin; ICAM-1, intercellular adhesion molecule-1.

sectional wall area of five-year sustained hypertensive Africans are, besides structural changes due to arterial degeneration, also dependent on the distending pressure, and that functional changes in the carotid artery may be more prominent than structural changes in this population. Our findings are consistent with evidence in white populations that show increased stiffness to be due to the increased distending pressure that accompanies hypertension, suggesting a functional adaptation, and not only structural alterations of the arterial wall.25,26 In contrast to these and our findings, one study found that the acute reduction in blood pressure by nitroglycerin does not normalise large artery stiffness in essential hypertensives.27 It is expected that sustained high blood pressure, as seen in hypertension, would cause vascular damage by, for instance, altering the collagen–elastin ratio of the arterial wall in favour of collagen.28,29 Indeed, after adjustment for mean arterial pressure, the difference in carotid distensibility between the hypertensives and normotensives remained, therefore suggesting the presence of structural alterations. Nevertheless, in light of the significant

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Table 7. Forward stepwise multiple regression analyses with CSWA and max LD as dependent variables Normotensives (n = 241) β (95% CI)

p-value

Hypertensives (n = 351) β (95% CI)

p-value

CSWA (mm2) Adjusted R2

0.23

0.32

Locality (urban)

–0.11 (–0.21– –0.01)

0.022

Gender (male)

0.29 (0.15–0.42)

< 0.001

0.26 (0.15–0.37)

< 0.001

Age, years

0.29 (0.17–0.42)

< 0.001

0.43 (0.30–0.57)

< 0.001

Waist circumference, cm

0.18 (0.05–0.31)

0.008

0.02 (–0.12–0.17)

0.77

Central SBP, mm Hg

0.18 (0.08–0.28)

< 0.001

LDL-C, mmol/l

0.12 (0.01–0.23)

0.026

HbA1c (%)

0.08 (–0.02–0.19)

0.13

Creatinine clearance, ml/min

0.11 (–0.05–0.26)

0.18

C-reactive protein, pg/ml

0.16 (0.02–0.29)

0.025

0.09 (–0.01–0.20)

0.090

γ-glutamyl transferase, U/l

0.11 (–0.02–0.24)

0.10

–0.08 (–0.18–0.02)

0.11

–0.12 (–0.22– –0.02)

0.013

Anti-hypertension medication (yes) Max LD (mm) Adjusted R2

0.27

0.10

Locality (urban) Gender (male)

0.26 (0.10–0.42)

0.001

Age, years

0.13 (–0.03–0.29)

0.12

Waist circumference, cm Central SBP, mmHg

0.20 (0.05–0.35)

0.010

Heart rate, bpm

–0.09 (–0.24–0.06)

LDL-C, mmol/l

–0.16 (–0.30– –0.02)

Creatinine clearance, ml/min

0.21 (0.05–0.37)

0.011

ICAM-1, pg/ml

0.16 (0.01–0.30)

0.037

γ-glutamyl transferase, U/l

0.17 (0.03–0.32)

0.023

–0.08 (–0.21–0.05)

0.21

0.22 (0.08–0.36)

0.002

0.14 (-0.005–0.28)

0.061

0.14 (0.007–0.27)

0.039

0.22 0.027 –0.14 (–0.2– –0.003)

C-reactive protein, pg/ml

0.045

0.09 (–0.05–0.23)

0.23

0.08 (–0.05–0.21)

0.25

Data expressed as beta-values and 95% confidence intervals, p-values obtained with forward stepwise multiple regression analyses. Included in each model: locality, age, gender, WC, HR, cSBP, LDL-C, HbA1c, C-reactive protein, ICAM-1, creatinine clearance, γ-glutamyl transferase, tobacco and anti-hypertensive medication use. CSWA, cross-sectional wall area; Max LD, maximum lumen diameter; SBP, systolic blood pressure; LDL-C, low-density lipoprotein cholesterol; HbA1c, glycated haemoglobin; ICAM-1,intracellular adhesion molecule-1.

cardiovascular burden that black populations carry,10,11 our results suggest that treatment that effectively lowers central pressure may also significantly lower the risk for stroke and other cardiovascular events. Our results were similar for treated and untreated hypertensives; therefore treatment seems to be largely ineffective in this population. Indeed, the treatment and control of hypertension in low-income countries are largely inadequate despite half of those sampled being aware of their condition.14,30 South Africa has one of the highest hypertension rates (78%) for people over 50 years of age, but only 38% are aware of their hypertensive status and only 7.8% of those treated for hypertension have controlled hypertension.14 It therefore remains to be seen whether effective anti-hypertensive treatment in black Africans will result in improved carotid distensibility. Surprisingly, the IMT was similar between the two groups after adjustments, suggesting a lack of visible structural changes

267

in the hypertensive blacks. IMT is an important marker of the atherosclerotic burden of the carotid artery,31 but it may also indicate non-atherosclerotic compensatory remodelling of the arterial wall in response to hypertension.32 However, neither of these possibilities seems to be the case in this black population. On the other hand, IMT was independently associated with cSBP in the hypertensive group only, therefore suggesting that the continued high pulsatile load of uncontrolled hypertension may eventually mediate structural changes in the carotid artery. This result shows a similar trend to the findings of Wang et al.,33 confirming the relevance of central blood pressure to IMT. We observed no differences in the inflammatory and endothelial activation markers, lipid levels and glycaemic status between the normotensives and hypertensives. Africans are generally not prone to atherosclerosis and coronary heart disease,34 and exhibit a favourable lipid profile,35,36 possibly explaining the similar lipid levels between the two groups. However, our results confirm the commonly found association between IMT and LDL-C level,37,38 and an association between CD and HbA1c level in the hypertensive group only. Although we did not observe structural differences after adjustment for cSBP, these results suggest glucose metabolism and lipid abnormalities may play a role in the arterial changes, although these are not yet detectable with ultrasound. Inflammation and endothelial activation (as indicated by the adhesion molecules) may not play a major role in the mediation of central arterial stiffness at this stage of disease progression. These results are unexpected in the light of previous findings, which indicate that acute and chronic inflammation are associated with stiffness of the large arteries,39 and that endothelial activation may be an important mediator of hypertensive vascular injury.40 The findings of this study should be interpreted in the context of its limitations and strengths. Our study population consisted of individuals from specific urban and rural areas in the North West Province of South Africa, and may not be representative of the whole population. We were not able to use echo-tracking techniques to determine local arterial stiffness in our field study; however, the procedures of ultrasound assessment are standardised41 and were performed by a single reader in a large study population. Carotid distensibility was calculated with a formula that includes cSBP, and we adjusted for cSBP. However, neither direct measurements such as IMT nor indirect variables such as carotid distensibility differed after adjustments for cSBP. Due to the cross-sectional study design, causality cannot be inferred. Although the results were consistent after several adjustments, we cannot exclude residual confounding.

Conclusion Although differences existed in terms of carotid structure and function between the normotensive and hypertensive Africans, it seemed to be partially accounted for by the increased distending pressure of the hypertensive group. Despite their hypertensive status, structural adaptations, such as IMT thickening, were not detectable in this African population after adjustment for potential confounders, and even before cSBP or MAP were taken into account. The classic cardiometabolic risk factors, markers of inflammation, endothelial activation and health behaviour seemed to play only a minor role in the mediation of carotid distensibility in this population at this stage of disease

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development. These results suggest that interventional strategies and the use of medication targeted at effectively lowering blood pressure may also lower the risk for adverse cardiovascular events in black South Africans.

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to hypertensive target organ damage. J Hypertens 2004; 22: 1731–1737. 13. Morris AA, Patel RS, Binongo JNG, Poole J, Mheid I, Ahmed Y, et al. Racial differences in arterial stiffness and microcirculatory function between black and white Americans. J Am Heart Assoc 2013; 2: e002154. 14. Lloyd-Sherlock P, Minicuci N, Beard J, Ebrahim S, Chatterji S.

This work was financially supported by the SANPAD (South Africa– Netherlands Research Programme on Alternatives in Development), PHRI

Hypertension among older adults in low and middle income countries: prevalence, awareness and control. Int J Epidemiol 2014; 43: 116–128.

(Population Health Research Institute), Medical Research Council of

15. Teo K, Chow C, Vaz M, Rangarajan S, Yusuf S. The Prospective Urban

South Africa, South African National Research Foundation (NRF) (GUN

Rural Epidemiology (PURE) study: examining the impact of societal

numbers 2069139 and FA2006040700010), North-West University and Roche

influences on chronic non-communicable diseases in low-, middle- and

Diagnostics. Any opinion, findings and conclusions or recommendations expressed in this material are those of the authors, and therefore the NRF do not accept any liability in regard thereto. The authors thank all supporting staff and the participants of the PURE study and in particular:

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and office staff in the Africa Unit for Transdisciplinary Health Research

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(AUTHeR) and the Hypertension in Africa Research Team (HART), Faculty

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PURE International: Dr S Yusuf and the PURE project office staff at the Population Health Research Institute, Hamilton Health Sciences and McMaster University, ON, Canada.

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Atherosclerosis 2014; 235: 576–584.

Conﬁdence Through Clinical and Real World Experience1-3 #1 NOAC prescribed by Cardiologists* Millions of Patients Treated Across Multiple Indications4 References: 1. Patel M.R., Mahaffey K.W., Garg J. et al. Rivaroxaban versus warfarin in non-valvular atrial fibrillation. N Engl J Med. 2011;365(10):883–91. 2. Tamayo S., Peacock W.F., Patel M.R., et al. Characterizing major bleeding in patients with nonvalvular atrial fibrillation: A pharmacovigilance study of 27 467 patients taking rivaroxaban. Clin Cardiol. 2015;38(2):63–8. 3. Camm A.J., Amarenco P., Haas S. et al. XANTUS: A Real-World, Prospective, Observational Study. 4. Calculation based on IMS Health MIDAS, Database: Monthly Sales December 2015. For full prescribing information, refer to the package insert approved by the Medicines Regulatory Authority (MCC). S4 XARELTO ® 10 (Film-coated tablets). Reg. No.: 42/8.2/1046. Each film-coated tablet contains rivaroxaban 10 mg. PHARMACOLOGICAL CLASSIFICATION: A.8.2 Anticoagulants. INDICATION: Prevention of venous thromboembolism (VTE) in patients undergoing major orthopaedic surgery of the lower limbs. S4 XARELTO ® 15 and XARELTO ® 20 (Film-coated tablets). Reg. No.: XARELTO ® 15: 46/8.2/0111; XARELTO ® 20: 46/8.2/0112. Each film coated tablet contains rivaroxaban 15 mg (XARELTO ® 15) or 20 mg (XARELTO ® 20). PHARMACOLOGICAL CLASSIFICATION: A.8.2 Anticoagulants. INDICATIONS: (1) Prevention of stroke and systemic embolism in patients with non-valvular atrial fibrillation (SPAF); (2) Treatment of deep vein thrombosis (DVT) and for the prevention of recurrent deep vein thrombosis (DVT) and pulmonary embolism (PE); (3) Treatment of pulmonary embolism (PE) and for the prevention of recurrent pulmonary embolism (PE) and deep vein thrombosis (DVT). HCR: Bayer (Pty) Ltd, Reg. No.: 1968/011192/07, 27 Wrench Road, Isando, 1609. Tel: 011 921 5044 Fax: 011 921 5041. L.ZA.MKT.GM.01.2016.1265 *Impact RX Data Oct - Dec 2015 NOAC: Non Vitamin K Oral Anticoagulant

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Cardio News Frontline initiatives in early myocardial reperfusion with ST-elevation myocardial infarction Concern has been expressed by leading cardiologists in Africa about the lack of preparedness of healthcare services on this continent in relation to the management of non-communicable diseases and, specifically, cardiovascular disease.1 This may be attributable to a paucity of surveillance data and registries, a shortage of physicians and cardiologists, interventional measures not being in place, inadequate diagnostic capabilities, and misguided opinions, as reported.2 From the South African 2011 census,3 we know that low household income compounds the problem of inadequate healthcare provision, and also lack of transport to facilities where optimal care can be provided timeously. Public sector clinic services are utilised by 61.2% of households, public hospitals by 9.5%, and private hospitals, private clinics and other services by only about 5% of households. A disparity is evident between the health facility used and the population group, in that 17% of black South Africans versus 88% of white and 64% of Indian households visit private health facilities. The Government report explains the preference for private health institutions

by long waiting times and unavailability of drugs in the public healthcare system. However, of the total population, 41% would be able to reach the health facility normally used within 30 minutes, and an additional 17% within 90 minutes. Disparity is also observed among population groups concerning coverage by medical aid or medical benefit schemes and other private health insurance. The most recent report from a study performed at a public academic hospital in Pretoria in 2015 states that ‘Only 37% of patients received fibrinolytic therapy and only 3% received the medication within one hour’.4 Similarly, 44.7% of ST-elevation myocardial infarction (STEMI) cases reportedly received fibinolytic therapy at the Groote Schuur Hospital in Cape Town (2012),5 and 36% of South African STEMI cases captured in the ACCESS registry (2007–2008) received fibrinolytic therapy.6 Baseline data for the STEMI Early Reperfusion Project, undertaken in private hospitals in the Tshwane metropolis (May – October 2012) to establish time intervals along the referral pathways from onset of symptoms to percutaneous coronary

Declaration of intent to fulfil the Stent-for Life mission in South Africa. From left to right: Dr William Wijns (SFL initiative founder and past president of EAPCI), Prof Petr Kala (SFL chairman), Prof Rhena Delport (SFL South Africa) and Prof Stephan Windecker (president of EAPCI).

intervention (PCI), showed that system delays were evident with inter-facility transport (IFT) compared with direct access (DA) to a PCI facility (median 3.7 vs 30.4 hours; p < 0.001). Doorto-balloon times of ≤ 90 minutes were achieved in a mere 22% of DA and 33% of IFT patients, and fibrinolysis within ≤ 30 minutes was only achieved in 50% of DA and 20% of IFT patients.7 The South African Heart Association Early Reperfusion Project for ST-elevation myocardial infarction commenced in 2012 as an initiative of the South African Heart Association (SAHA), with Dr Adriaan Snyders as president of the association. The pilot study in the Tshwane metropolis private sector7 informed an observational multi-centre study in South African hospitals, launched in the last quarter of 2015, to identify factors that contribute to delays in early reperfusion for STEMI. A sub-study will be launched in 2017 to investigate whether implementation of a hub-and-spokes model (hub hospitals: PCI-capable hospitals, and spokes: referral hospitals), with the application of an ICT (an ECG-capturing, patient-monitoring, communication and data-capturing

Signing the intent. From left to right: Prof Stephan Windecker (president of EAPCI), Prof Rhena Delport (SFL South Africa) and Prof Petr Kala (SFL chairman).

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device) could contribute towards more effective management of STEMI in South Africa. In addition, the societal cost of an undertreated STEMI population will be estimated to determine the potential financial impact of the intervention as well the cost benefits of the treatment modalities (PCI and fibrinolysis). International collaboration has been established between the South African Society of Cardiovascular Intervention (SASCI), STEMI India (who developed the software and the model) and Stentfor-Life (SFL) Europe to pursue these objectives. SAHA applied for membership as affiliated country of SFL Europe, an initiative of the European Association of Percutaneous Cardiovascular Interventions (EAPCI) and Prof Rhena Delport (regional editor for South Africa of the Cardiovascular Journal of Africa and project manager for SFL South Africa) signed the declaration of intent, on behalf of SASCI, on 27 February 2016,

to fulfil the SFL mission in South Africa. SAHA has thus positioned itself in the frontline of STEMI care and hopefully, with concerted action among all role players in STEMI management in South Africa, STEMI outcomes will improve and the cardiovascular disease-related burden of disease will be managed appropriately.

Report no. 03-00-05 (2011).http://www. statssa.gov.za/publications/Report-03-00-05/ Report-03-00-052011.pdf. 4.

Meel R, Gonçalves R. Time to fibrinolytics for acute myocardial infarction: Reasons for delays at Steve Biko Academic Hospital, Pretoria, South Africa. S Afr Med J 2015; 106(1): 92–96.

Rhena Delport

Maharaj RC, Geduld H, Wallis LA. Doorto-needle time for administration of fibrinolytics in acute myocardial infarction in Cape

primary care system for non-communicable

Town. S Afr Med J 2012; 102(4): 241–244.

Kengne AP, Mayosi BM. Readiness of the 6.

tigators. Management of acute coronary

Health 2014; 2(5): e247–248.

syndrome in South Africa: insights from

Hertz JT, Reardon JM, Rodrigues CG, de

the ACCESS (Acute Coronary Events – a

Andrade L, Limkakeng AT, Bloomfield GS,

multinational survey of current management

Lynch CA. Acute myocardial infarction in

strategies) registry. Cardiovasc J Afr 2012; 7: 365–370.

sub-Saharan Africa: the need for data. PLoS One 2014; 9(5): e96688. 3.

Schamroth C; ACCESS South Africa inves-

diseases in sub-Saharan Africa. Lancet Glob

Snyders A, Delport R. The SA Heart

Use of health facilities and levels of

STEMI Early Intervention Project. Referral

selected health conditions in South Africa:

pathways for reperfusion of STEMI – devel-

Findings from the General Household

oping strategies for appropriate intervention.

Survey, 2011. Statistics South Africa.

SA Heart J 2015; 12: 72–80.

SA Heart Association Early Reperfusion Project

TIME TO REPERFUSION IS CRITICAL Primary-PCI capable centre

Patient has chest pain Preferably < 10 min

Monitor every 10 min if chest pain persists

Diagnosis with 12 lead ECG

EMS or non-primary-PCI capable centre

STEMI diagnosis*

Preferably < 60 min

PCI possible in < 120 min?

MEDICATION

• •

to administer before primary PCI Loading dose Aspirin 150 - 300 mg orally or 80 - 150 mg IV Loading dose Clopidogrel 600 mg orally if aged ≤ 75 years OR Loading dose Prasugrel 600 mg orally

Primary-PCI

Coronary Angiography

Preferably 3 - 24 hours

CONTACT INFORMATION:

YES

Successful Fibrinolysis?

Immediate transfer to PCI center

MEDICATION

to administer with fibrinolytic therapy • Loading dose Aspirin 150-500 mg orally or 250 mg IV • Loading dose Clopidogrel 300 mg orally if aged ≤ 75 years • Tenecteplase Single i.v. bolus: (TNK–tPA) 30 mg if < 60 kg 35 mg if 60 to < 70 kg 40 mg if 70 to < 80 kg 45 mg if 80 to < 90 kg 50 mg if ≥ 90 kg • Streptokinase 1.5 million units over (SK) 30 – 60 min i.v.

Preferably < 90 min ≤ 60 min for early presenters

Immediately

NO Preferably < 30 min

Immediate transfer to PCI center

Rescue PCI ECG = electrocardiogram FMC = first medical contact STEMI = ST-segment elevation myocardial infarction PCI = Percutaneous coronary intervention

YES

Thrombolysis ABSOLUTE Contra-indications • Previous intracranial haemorrhage or stroke of unknown origin at any time • Ischaemic stroke in the preceding 6 months • Central nervous system damage or neoplasms or atrioventricular malformation • Recent major trauma/surgery/head injury (within the preceding 3 weeks) • Gastrointestinal bleeding within the past month • Known bleeding disorder (excluding menses) • Aortic dissection • Non-compressible punctures in the past 24 h (e.g. liver biopsy, lumbar puncture)

Immediate Fibrinolysis

Dr Adriaan Snyders - National Champion for project 082 446 1558 Prof Rhena Delport - Project Manager 082 445 4500

Reference: 1. ESC Guidelines for the management of acute myocardial infarction in patients presenting with ST-segment elevation. Eur Heart J. 2012 Oct;33(20):2569-619. doi: 10.1093/eurheartj/ehs215. 2. http://www.heartfoundation.co.za/how-your-heart-works/symptoms-heart-attack (accessed on 7 Feb 2013)

Thrombolysis RELATIVE Contra-indications • Transient ischaemic attack in the preceding 6 months • Oral anticoagulant therapy • Pregnancy or within 1 week postpartum • Refractory hypertension (systolic blood pressure > 180 mmHg and/or diastolic blood pressure > 110 mmHg) • Advanced liver disease • Infective endocarditis • Active peptic ulcer • Prolonged or traumatic resuscitation

Novo Nordisk South Africa

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Through the proper use of long-acting (basal) insulins, fast-acting insulins and combinations of these (including pre-mix insulins), it became possible to more closely replicate the natural production of insulin in the body, allowing people with diabetes a greater degree of treatment flexibility.5 p219a, p220b However, this often means a person who is on insulin treatment needs to take several injections throughout the day to address the body’s varying insulin needs.7 p15c Most recently, technology has moved a step closer to the concept of an ideal insulin regimen in that novel insulins mimic the profile of natural insulin production more closely than conventional insulins.7 p15c Because they are modified to have an ultra-long duration of action, timing of the dose is flexible.7 p19a Furthermore, because of their novel chemical properties, for the first time it is possible to combine two different insulin analogues in a single pen device.7 p16b Through this innovation, an ultra-long-acting insulin and a rapidacting insulin can be combined and administered with one injection device, potentially reducing the number of injections that a person would need to control their blood glucose throughout the day.7 p16b,p18b

Device updates In tandem with these breakthroughs in the development of insulin, the devices used to administer the insulin have also progressed. The advent of the insulin pen has made these once-feared injections as easy as the click of a button. First launched in 1985 by Novo Nordisk8 p505a, the world’s largest diabetes company, insulin pens today are either durable devices (that can be reused) with replaceable cartridges, or prefilled disposable pens. These pens use shorter, finer and sharper needles than those of traditional syringes, making the process of injecting oneself easy, smooth and virtually painless.8,9 8. p508 9. p12a

Insulin pens have allowed injection of insulin to become convenient and discreet, while improving lifestyle flexibility and making it easier for people with diabetes to adhere to their therapy. They have also removed the potential stigma of drawing up insulin in a syringe and injecting it in a public place.9 p12b,p15a Over the years, advances in technology have included the development of pens that deliver smaller more accurate doses specifically designed to suit the needs of children and adolescents. Colourful, creative, interchangeable ’skins’ allow the devices to be personalised. In 2010, there was a newly developed pen with a memory function that records the dose and the time delivered, providing assurance for parents and school staff that the correct dose was given at the right time.9 p11a Novo Nordisk’s continued focus on innovative solutions to adequately meet the needs of the world’s growing population of people with diabetes has now resulted in another world-first in injector devices; aimed at improving the

convenience and safety of insulin injections. Equipped with a series of new easy-to-use, patient-friendly safety features to ensure that the correct dose is administered, the latest pens are disposable devices that are prefilled with different types of insulin according to the requirements of individual users. In addition, many of the latest features have been designed specifically to accommodate people with visual impairment or reduced dexterity, making it much easier for them to use an insulin pen rather than a vial and syringe.9,10 10. p523a; 9.p14b

Like any other innovation, progress in insulin development not only solves some of the problems posed by older treatments and administration devices, but also highlights new ones. These ’treatment gaps’ are a constant focus to the scientists involved in making the lives of people with diabetes safer, easier and happier. For the first time, being able to combine a true long-acting insulin analogue with a short-acting one in the same novel administration pen is a further encouraging step in this journey. Both insulin and device will soon be available in South Africa. This information is supported by Novo Nordisk – the world’s largest diabetes company. Novo Nordisk is committed to innovation, and continues to evolve insulin and diabetes care to improve the lives of the millions of people living with diabetes around the world.

References 1. 2.

Aras M. Ninety years of insulin – Canada celebrates. Diabetes Voice. 2012; 57(1): 41-42. Rotella CM, et al. Role of Insulin in the Type 2 Diabetes Therapy: Past, Present and Future. International Journal Endocrinology and Metabolism. 2013; 11(3): 137-144. 3. Pettus J, et al. The past, present, and future of basal insulins. Diabetes/Metabolism Res Rev 2015. Published online. DOI: 10.1002/dmrr 4. Novo Nordisk A/S. Novo Nordisk History. Available at: http://www.novonordisk.com/aboutnovo-nordisk/novo-nordisk-history/step-by-step.html. Accessed on 4 July 2016. 5. Ashok KD & Ashida TS. Modern Insulins – The Insulin Analogues: A Reappraisal. Medicine Update. (n.d.) Available at: http://www.apiindia.org/pdf/medicine_update_2007/39.pdf. Accessed on 4 July 2016. 6. Novo Nordisk (USA). What is Insulin? Types of Insulin (2015). Available at: https://www. novolog.com/type-2-diabetes/general-type-2/diabetes-and-mealtime-insulin/what-is-insulin. html. Accessed on 4 July 2016. 7. Unnikrishnan AG, et al. Review of clinical profile of IDegAsp. Journal of The Association of Physicians of India. 2015; 63(Suppl): 15-20. 8. Selam JL. Evolution of Diabetes Insulin Delivery Devices. Journal of Diabetes, Science and Technology. 2010; 4(3) 505-513. 9. Hyllested-Winge J et al. NovoPen Echo® insulin delivery device. Med Devices: Evidence and Research (Auckl). 2016; 9: 11-18. 10. Pearson TL. Practical Aspects of Insulin Pen Devices. Journal of Diabetes Science and Technology. 2010;4(3): 522-531.

This message is brought to you by Novo Nordisk, the world’s leaders in diabetes management.

AFRICA

CARDIOVASCULAR JOURNAL OF AFRICA • Volume 27, No 4, July/August 2016

Cardiovascular Topics Management of arterial hypertension in Cotonou city, Benin: general practitioners’ knowledge, attitudes and practice Martin Dèdonougbo Houenassi, Léopold Houétondji Codjo, David Dokoui, Serge Hugues Mahougnon Dohou, Armand Wanvoegbe, Anthelme Agbodande, Angelo Cossi Attinsounon, Adebayo Alassani, Séraphin Ahoui, Albert Comlan Dovonou, Thierry Armel Adoukonou

Abstract Objective: We aimed to assess the management of hypertensive patients by general practitioners in Cotonou city. Methods: This was a cross-sectional study based on a multicentre survey conducted from 1 May to 31 July 2011. We recruited all consenting general practitioners who worked in public and private centres in Cotonou city. We used the 7th report of the Joint National Committee to assess the manage-

Health Unit, Education and Research in Cardiology, Faculty of Health, University of Abomey Calavi, Cotonou, Bénin Martin Dèdonougbo Houenassi, PhD David Dokoui, MD

Department of Medicine and Medical Specialties, Faculty of Medicine, University of Parakou, Parakou, Bénin Léopold Houétondji Codjo, MD, leostelles@yahoo.fr Angelo Cossi Attinsounon, MD Adebayo Alassani, MD Séraphin Ahoui, MD Albert Comlan Dovonou, MD Thierry Armel Adoukonou, MD

Department of Cardiology, Military Teaching Hospital, Parakou, Bénin Serge Hugues Mahougnon Dohou, MD

Department of Internal Medicine, National University Hubert Koutoukou Maga Hospital, Cotonou, Bénin Armand Wanvoegbe, MD Anthelme Agbodande, MD

ment of hypertension by general practitioners. A tested and validated self-questionnaire was used to collect the data on hypertension management by general practitioners. Results: In eight centres that approved the study, 41 general practitioners were included. The definition of hypertension was known by 20 (48.8%) practitioners. Only 25 (61.0%) could describe the conditions for blood pressure measurement. Ten of them were unable to list half of the minimum recommended tests for hypertension, and the majority (92.7%) did not have any idea of global cardiovascular risk. The blood pressure goal was known by only 18 (43.9%) practitioners. Lifestyle (82.9%) and monotherapy (70.7%) were the therapeutic modalities most prescribed. Antihypertensive agents commonly used by practitioners were calcium channel blockers (82.9%), angiotensin converting enzyme inhibitors (53.7%) and diuretics (36.6%). The general practitioners referred their patients to cardiologists mainly for uncontrolled hypertension (63.4%) and the onset of acute complications (56.1%). Conclusion: The general practitioners’ knowledge of hypertension was insufficient and their management did not reflect international guidelines.

Keywords: management, arterial hypertension, general practitioners, knowledge, attitudes and practice, Africa Submitted 29/7/15, accepted 7/12/15 Cardiovasc J Afr 2016; 27: e1–e6 DOI : 10.5830/CVJA-2015-094

www.cvja.co.za

CARDIOVASCULAR JOURNAL OF AFRICA • Volume 27, No 4, July/August 2016

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Prise en charge des hypertendus dans la ville de Cotonou (Bénin) en 2011: connaissances attitudes et pratiques des médecins généralistes Martin Dèdonougbo Houenassi, Léopold Houétondji Codjo, David Dokoui, Serge Hugues Mahougnon Dohou, Armand Wanvoegbe, Anthelme Agbodande, Angelo Cossi Attinsounon, Adebayo Alassani, Séraphin Ahoui, Albert Comlan Dovonou, Thierry Armel Adoukonou

Abstract But: Ce travail vise à évaluer les connaissances, attitudes et pratiques des médecins généralistes sur la prise en charge de l’hypertension artérielle à Cotonou. Méthodes: L’étude était transversale et descriptive basée sur une enquête multicentrique du 1er Mai 2011 au 31 Juillet 2011. Un recrutement de tous les médecins généralistes, volontaires exerçant dans les centres de santé privés, publics et confessionnels de la ville de Cotonou, ayant autorisé l’étude, a été fait. Le 7ème rapport de Joint National Committee (JNC7) a été utilisé comme référentiel pour l’évaluation de la prise en charge des hypertendus. Un auto-questionnaire adressé aux généralistes, testé et validé, a été utilisé pour recueillir ces données. Résultats: Au total, 41 médecins généralistes dans huit établissements sanitaires ont été inclus. Près de la moitié des généralistes (48.8%) ne connaissaient pas la définition de l’HTA. Seulement 25 généralistes (61.0%) pouvaient décrire les conditions de mesure de la pression artérielle. Dix généralistes (24.4%) étaient incapables de lister la moitié des examens du bilan minimum de l’hypertension artérielle (HTA). La majorité (92.7%) ne connaissait pas la notion de risque cardiovasculaire global. L’objectif tensionnel (TA ≤ 140/90 mmHg) n’était connu que par 18 (43.9%) médecins.

Unité de Soins, d’Enseignement et de Recherche en Cardiologie, Faculté des Soins de la Santé, Université d’Abomey Calavi, Cotonou, Bénin Martin Dèdonougbo Houenassi, PhD David Dokoui, MD

Département de Médecine et spécialités médicales, Faculté de Médecine, Université de Parakou, Parakou, Bénin Léopold Houétondji Codjo, MD, leostelles@yahoo.fr Angelo Cossi Attinsounon, MD Adebayo Alassani, MD Séraphin Ahoui, MD Albert Comlan Dovonou, MD Thierry Armel Adoukonou, MD

Service de cardiologie, Hôpital d’Instruction des Armées de Parakou, Bénin Serge Hugues Mahougnon Dohou, MD

Service de Médecine Interne, Centre National Hospitalier Universitaire Hubert Koutoukou Maga, Cotonou Bénin Armand Wanvoegbe, MD Anthelme Agbodande, MD

Les mesures hygiéno-diététiques seules (82.9%) et la monothérapie seule (70.7%) étaient les modalités thérapeutiques les plus prescrites. Les classes pharmacologiques antihypertensives prescrites étaient surtout les inhibiteurs calciques (82.9%), les inhibiteurs de l’enzyme de conversion (53.7%) et les diurétiques (36.6%). Les généralistes référaient les hypertendus aux cardiologues principalement pour non-maitrise du chiffres tensionnels (63,4%) et l’apparition de complications aigues (56.1%). Conclusion: Les connaissances des généralistes sur la prise en charge de l’HTA étaient insuffisantes et leurs attitudes et pratiques ne respectaient pas les recommandations internationales.

Mots de cles: prise en charge, hypertension artérielle, médecins généralistes, connaissances attitudes et pratiques, Afrique Submitted 29/7/15, accepted 7/12/15 Cardiovasc J Afr 2016; 27: e1–e6

www.cvja.co.za

DOI : 10.5830/CVJA-2015-094

L’hypertension artérielle (HTA), grave problème de santé publique, est en forte progression dans toutes les régions de l’Afrique sub-saharienne.1 Elle est un facteur de risque majeur dans la survenue d’accidents vasculaires cérébraux, d’insuffisance cardiaque, d’insuffisance rénale et de maladies coronaires qui représentent les principales causes de décès dans le monde.2,3 En dépit de la disponibilité d’une thérapie médicale efficace, plus de la moitié des hypertendus traités ont une pression artérielle mal contrôlée.4,5 Ce mauvais résultat est la conséquence d’une inefficacité du système de soins à tous les niveaux: dépistage, traitement, et observance thérapeutique.6 La mauvaise connaissance sur la prise en charge de l’HTA a été rapportée par Noubiap et al. au Cameroun en 2014. Peu de praticiens non cardiologues maitrisaient la définition de l’HTA, le bilan minimal, le choix des antihypertenseurs et la notion d’objectif thérapeutique.7 Safar et al. ont rapporté d’autres barrières à la gestion efficiente de l’hypertension artérielle. Il s’agissait entre autres de la faible fréquentation de ces centres par les patients, la mauvaise tenue des dossiers médicaux et le non respect des recommandations locales sur la prise en charge de l’HTA.8 Ce travail a été donc initié pour évaluer les connaissances, attitudes et pratiques des médecins généralistes sur la prise en charge de l’hypertension artérielle à Cotonou en 2011.

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Methodes

Traitement et analyse des données

L’étude était multicentrique et avait inclus huit centres privés, publiques et confessionnels de médecine générale de la ville de Cotonou. Capitale économique du Bénin, Cotonou est une ville cosmopolite qui concentre l’essentiel des activités politiques, administratives, culturelles et sanitaire du Bénin. Il s’agissait d’une étude transversale et descriptive menée du 1er Mai 2011 au 31 Juillet 2011. La population d’étude était composée de médecins généralistes, volontaires exerçant dans les différents centres de santé choisis depuis au moins le mois de janvier 2011. Pour le choix des centres de santé, une demande d’autorisation a été adressée à l’administration de chaque établissement pratiquant la médecine générale. Seuls les centres qui ont donné leur autorisation ont été inclus. Dans ces centres le recrutement des médecins généralistes était systématique. La variable dépendante était la prise en charge de l’hypertension artérielle par les médecins généralistes. Cette prise en charge a été jugée à partir des connaissances, attitudes et pratiques de ces derniers en matière d’hypertension artérielle. Le 7ème rapport de Joint National Committee (JNC7)9 a été utilisé comme référentiel pour l’évaluation de la prise en charge. Les tableaux 1 et 2 présentent les items abordés dans l’évaluation des connaissances, attitudes et pratiques des généralistes. Les médecins généralistes ont été investigués à l’aide d’un auto-questionnaire anonyme. L’enquête a été menée par un étudiant en 7ème année de médecine préalablement formé au questionnaire. Ce questionnaire a été testé sur 10 médecins généralistes et amélioré. L’accord des autorités administratives des différents établissements sanitaire a été obtenu. Le consentement écrit des médecins généralistes enquêtés a été obtenu. La confidentialité des données recueillies sur les participants a été garantie. Des séances de restitution des résultats de cette enquête et une mise au point sur la gestion de l’hypertendu par le généraliste ont été organisés dans chaque structure sanitaire ayant participé à l’enquête.

Les données collectées ont été saisies, traitées et analysées avec le logiciel Epi info version 3.5.3. Les variables qualitatives étaient exprimées en pourcentage avec leur intervalle de confiance à 95% et les variables quantitatives en moyenne ± écart type.

Tableau 2. Critères de jugement l’attitude et pratique des médecins généralistes en matière d’hypertension artérielle (HTA) dans la ville de Cotonou (Bénin) en 2011 Questions

Réponses attendues

Quelle(s) est (sont) la (les) mesure(s) hygiéno-diététique(s) de base que vous recommandez à tout hypertendu?

• • • • •

Restriction sodée; Arrêt du tabac; Modération de la consommation d’alcool; Réduction pondérale; Exercice physique régulier

Quels gestes d’examen physique • Recherche de contact lombaire, de souffle faites-vous devant une HTA de abdominal, de souffle cardiaque, des pouls; découverte récente en absence • Calcul de l’IMC (après mesure de poids et de complications apparentes? de taille) Mesure du tour de taille • Palpation de la thyroïde Faut-il évaluer le risque cardio- Oui vasculaire global avant le traitement antihypertenseur? Quelle(s) classe(s) d’antihypertenseur utilisezvous en première intention en monothérapie?

• Diurétiques thiazidiques ou inhibiteurs calciques (IC) ou inhibiteurs de l’enzyme de conversion (IEC) ou antagonistes des récepteurs de l’angiotensine II (ARA II) ou bétabloquants (BB)

Quelle(s) stratégie(s) thérapeutique(s) adoptez-vous en première intension?

• HTA grade I et risque faible: mesures hygiéno-diététiques (MHD) • HTA et risque moyen: monothérapie ou bithérapie en combinaison fixe et MHD • HTA et risque élevé: Au moins une bithérapie en combinaison fixe

Quelle(s) est (sont) • Deux antihypertenseurs de la même classe le(s) association(s) • IEC et ARA II d’antihypertenseurs à proscrire? Quels sont vos objectifs thérapeutiques au cours d’un traitement antihypertenseur?

• Baisse TAS < 140 et TAD < 90 mmHg • Prévention des accidents cardiovasculaire par la réduction du risque cardiovasculaire absolu

Quelle(s) est (sont) votre (vos) attitude(s) devant un patient hypertendu qui associe des tisanes aux antihypertenseurs?

• Le sensibiliser sur le risque d’insuffisance rénale • Arrêter les tisanes

Tableau 1. Critères de jugement des connaissances des médecins généralistes en matière d’hypertension artérielle (HTA) dans la ville de Cotonou (Bénin) en 2011 Questions

Réponses attendues

Quels sont les critères diagnostiques de l’HTA?

• PAS ≥ 140 mmHg et/ou PAD ≥ 90 mmHg • Mesurée chez un patient assis ou couché depuis au mois 05 minutes, les bras soutenus dans le plan du cœur. La caféine, l’exercice physique et le tabagisme doivent être évités au moins 30 minutes avant la mesure • Valeurs confirmées au cours d’au moins trois consultations différentes (deux mesures à chaque consultation au cours d’une période de 3 à 6 mois) si toutes fois la gravité de l’HTA ne justifie pas un traitement antihypertenseur immédiat.

Quelle est la classification de l’HTA selon JNC 7 (Joint National Comittee 7)?

• • • •

Quel est le bilan minimum de l’HTA?

• Biologie sanguine: glycémie à jeun, créatinine, taux d’hémoglobine, cholestérol (total, HDL), triglycérides, uricémie • Biologie urinaire : protéinurie, hématurie • Electrocardiogramme

TA normale: TAS < 120 mmHg et TAD < 80 mmHg Pré-HTA: TAS entre 120–139 mmHg ou TAD entre 80–89 mmHg HTA grade I: TAS entre 140–159 mmHg ou TAD entre 90–99 mmHg HTA grade II: TAS ≥ 160 mmHg ou TAD ≥ 110 mmHg

Quels sont les autres facteurs de risque cardio- • Age, sexe, antécédents familiaux d’accident vasculaire cérébral (AVC)/d’infarctus du myocarde(IDM) précoce ou de vasculaire à rechercher chez l’hypertendu? mort subite • Tabac, diabète, dyslipidémie, sédentarité, obésité Quels sont les critères de gravité de l’HTA

• TAS ≥ 180 mmHg et/ou TAD ≥ 110 mmHg ou • Diabète associé à l’HTA ou • Syndrome métabolique (association de trois facteurs parmi l’HTA, l’hyperglycémie, l’hypocholestérolémie HDL, l’hypertriglycéridémie et l’obésité abdominale) ou • Accumulation d’au moins trois facteurs de risque cardiovasculaire ou • Existence d’une atteinte d’organe cible (hypertrophie ventriculaire gauche, protéinurie, rétinopathie hypertensive) ou de maladie cardiovasculaire (AVC, IDM, Artériopathie) ou de maladie rénale

Comment évaluer le risque cardiovasculaire absolu

• Avec la table de Framingham ou autres scores • A partir des valeurs tensionnelles, de cholestérols, des antécédents de tabagisme et de diabète.

CARDIOVASCULAR JOURNAL OF AFRICA • Volume 27, No 4, July/August 2016

Resultats L’enquête a été autorisée par huit établissements sanitaires. Il y avait trois centres publiques, trois centres privés et deux centres confessionnels. Les médecins généralistes présents et recensés étaient au nombre de 41. L’âge moyen des médecins était de 35.8 ± 9.6 ans avec les extrêmes de 24 à 66ans. Il y avait 28 (68.3%) hommes. En ce qui concerne l’expérience professionnelle, 20 (48.8%) médecins exerçaient depuis moins de cinq ans; 13 (31.7%) avaient une ancienneté entre 5 et 10 ans et 8 (19.5%) depuis au moins 10 ans.

Connaissances des définition et classification de l’hypertension artérielle Près de la moitié des généralistes (48.8%) ne connaissaient pas la définition exacte de l’HTA et 14 (34.1%) ne pouvaient pas donner les seuils de pression artérielle systolique (PAS) et de pression artérielle diastolique définissant l’HTA. Seulement 25 généralistes (61.0%) pouvaient décrire les conditions de mesure de la pression artérielle. Presque tous les généralistes (95.1%) ignoraient la classification de l’HTA selon JNC 7.

Connaissances des evaluation de l’hypertendu Concernant le bilan minimum de l’organisation mondiale de la santé (OMS), 10 généralistes (24.4%) étaient incapables de lister la moitié des examens constituant ce bilan. Un seul médecin (2.4%) a pu lister tous les examens de ce bilan. Le tableau 3 présente la proportion de médecins connaissant chacun des examens du bilan minimum de l’OMS. Au moins 26.8% des médecins généralistes ne connaissaient aucun critère de gravité de l’HTA. Selon le facteur de gravité énuméré par ces médecins, on pouvait les répartir comme suit: (30) 73.2% pour la sévérité des chiffres tensionnels (TAS ≥ 180 mmHg ou TAD ≥ 110 mmHg), 25 (61%) pour l’existence d’atteinte d’organes cibles, 10 (24.4%) pour l’existence de complication cardiaque, 10 (24.4%) pour l’existence de maladie rénale, six (14.6%) pour l’accumulation d’au moins trois facteurs de risque cardiovasculaire et quatre (9.8%) pour l’existence d’un diabète associé. Aucun généraliste ne reconnaissant le syndrome métabolique comme critère de gravité de l’HTA. La majorité des généralistes (92.7%) ne connaissaient pas la notion de risque cardiovasculaire global.

Tableau 3. Proportion de médecins généralistes connaissant chacun des examens composant bilan minimum de l’Organisation Mondiale de la Santé à Cotonou (Bénin) en 2011 Examens paracliniques

Effectif

Pourcentage

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Connaissances des traitements de l’hypertension artérielle Chez un hypertendu traité, l’objectif tensionnel (TA ≤ 140/90 mmHg) n’était connu que par 18 (43.9%) médecins. Encore moins de généralistes (17.1%) savaient que le traitement de l’HTA visait non seulement la baisse des chiffres tensionnels mais aussi la réduction du risque cardiovasculaire global et la prévention des accidents/maladies cardiovasculaires. Un médecin généraliste sur 10 (9.8%) ne connaissait aucune mesure hygiéno-diététique spécifique à recommander à un hypertendu. La plupart des médecins savaient qu’il fallait recommander un régime pauvre en sel (90.2%), la consommation d’une alimentation pauvre en graisse (85.4%) et la pratique d’une activité physique régulière (85.4%). Par contre peu de médecins savaient que la réduction pondérale (4.9%), l’arrêt du tabac (29.3%) et la réduction de la consommation d’alcool (39%) faisant partie des conseils d’hygiène à donner à un hypertendu. Les éléments de surveillance du traitement antihypertenseur cités par les médecins généralistes étaient les chiffres tensionnels (92.1%), la survenue de complications de l’HTA (34.1%) et les effets secondaires du traitement (15.6%).

Évaluation des attitudes et pratiques des généralistes face à l’HTA A l’examen physique de l’hypertendu, les gestes réalisés systématiquement par les médecins généralistes étaient la recherche d’anomalies auscultatoires cardiaques et vasculaires (90.2%), la palpation des pouls périphériques (39%) et le calcul de l’indice de masse corporelle (7.9%). Aucun médecin ne mesurait le tour de taille des patients. Aucun médecin généraliste n’évaluait le niveau de cardiovasculaire global des patients avant la mise sous traitement antihypertenseur. En ce qui concerne les modalités thérapeutiques fréquemment prescrites par les médecins, on retrouve les mesures hygiénodiététiques (MHD) seules dans 82.9%, la monothérapie seule dans 70.7%, la bithérapie seule dans 14.6%, l’association MHD-monothérapie dans 53.7% et l’association MHD-bithérapie dans 14.6% des cas. Aucun médecin ne prescrivait la trithérapie antihypertensive. Les classes pharmacologiques antihypertensives prescrites étaient les inhibiteurs calciques (82.9%), les inhibiteurs de l’enzyme de conversion (53.7%), les diurétiques (36.6%), les bétabloquants (14.6%) et les antihypertenseurs centraux (7.3%). Lors du suivi des hypertendus, les motifs pour lesquels les généralistes référaient les patients aux cardiologues étaient la non-maitrise des chiffres tensionnels (63.4%), l’apparition de complications aigues (56.1%) et la référence systématique de tous les cas d’HTA (36.6%).

Dans le sang Glycémie à jeun

80.5

Créatinine

85.4

Potassium

56.1

Cholestérol (total, HDL)

90.2

Triglycérides

82.9

Acide urique

29.3

Hémoglobine/hématocrite

21.9

Dans les urines (bandelette urinaire) Protéines, Hématies

07.3

Electrocardiogramme

80.5

Discussion Ce travail nous a permis de décrire les connaissances, attitudes et pratiques des médecins généralistes de la ville de Cotonou sur la prise en charge de l’HTA. La prise en compte des établissements sanitaires aussi bien publiques, confessionnels que privés et le recrutement systématique de tous les médecins généralistes y exerçant nous ont permis de réduire les biais de sélection. Cependant, le choix non aléatoire de ces établissements empêchera la généralisation de nos résultats à toute la ville de

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Cotonou. L’usage d’un auto-questionnaire adressé aux médecins garantit la sincérité des déclarations recueillies. Au terme de cette étude, nous avons observé que les médecins généralistes ne maitrisaient pas la définition de l’HTA ni l’évaluation de l’hypertendu conformément aux recommandations de JNC7. Les objectifs tensionnels n’étaient pas connus de tous les médecins généralistes de même la prescription des MHD. Les médicaments anti hypertenseurs recommandés sont bien connus par les généralistes mais les diurétiques thiazidiques sont peu prescrits par ces derniers. La mauvaise connaissance des généralistes sur l’HTA n’est pas spécifique à la ville de Cotonou. En effet, au Pakistan en 2010, Rehman et al. ont rapporté que 25.5% des médecins généralistes ne connaissaient pas la valeur seuil qui définit l’HTA.10 À l’ouest du Cameroun en 2012, 36.4% des médecins généralistes utilisaient de fausses valeurs pour définir l’HTA.7 La situation est encore pire au Burkina Faso où, en 2002, 65% des médecins avaient basé la définition de l’HTA essentiellement sur la pression artérielle systolique.11 De même, en Chine, Chen et al., en 2011, ont observé une mauvaise définition de l’HTA par les généralistes dans 55.8%.12 Pour l’évaluation de l’hypertendu, un bilan minimum est recommandé en complément de l’examen clinique et a pour but de rechercher les facteurs de risque associés à l’HTA, les atteintes des organes cibles et co-morbidités de l’HTA (insuffisance rénale, insuffisance cardiaque, coronaropathie) et les signes orientant vers une éventuelle hypertension artérielle secondaire (hypokaliémie, insuffisance rénale, anémie).9,13,14 Dans notre série, 80% des médecins généralistes demandaient le bilan minimum mais il n’était à moitié conforme que chez 24.4% des praticiens. Notre observation rejoint celle de Noubiap et al. qui avaient rapporté une demande de bilan minimum par 87% des généralistes avec un taux d’adéquation de faible (10.4%).7 Cette déficience observée dans l’évaluation de l’hypertendu nous permet de comprendre la méconnaissance des critères de gravité de l’HTA par les généralistes. Si la sévérité des chiffres tensionnels (≥ 180/110 mmHg) et les retentissements viscéraux étaient connus comme éléments de gravité de l’HTA par certains généralistes dans notre série, aucun des ces derniers ne considérait le syndrome métabolique, et seulement 16.9% de ces derniers considéraient le diabète comme critères de haut risque cardiovasculaire. De même, aucun généraliste ne connaissait la notion du risque cardiovasculaire global (RCVG). Pourtant, l’évaluation du RCVG est une étape majeure de la prise en charge de l’hypertendu. En effet, le niveau de risque découle du bilan de l’hypertendu et oriente le traitement médicamenteux de même que la définition des objectifs tensionnels.9,13,14 La méconnaissance du RCVG par les praticiens expose les patients à une mauvaise prise en charge avec comme conséquence un mauvais contrôle tensionnel et un risque accru d’accident cardiovasculaire.15 Sur le plan thérapeutique l’objectif tensionnel n’était connu que par 43.9% des généralistes et seulement 14.1% tenaient compte du RCVG pour définir le traitement de leurs patients hypertendus. Au Pakistan en 2005, Jafar et al. avaient observé que 52.3% des généralistes connaissaient les objectifs tensionnels.16 Ce taux était de 44.2% au Cameroun en 2014.7 La place des mesures hygiénodiététiques (MHD) dans le traitement de l’HTA était bien connue par la plus part des médecins généralistes dans notre étude (90.2%) de même que celle de Chen et al. (99.3%).12 Les MHD sont moins pratiquées dans d’autres séries africaines:

77.9% au Cameroun7 et 50% en Afrique du Sud.17 Il est important de reconnaitre cependant que, même si ces MHD ont été souvent prescrites par ces généralistes, les différentes composantes de ces MHD n’étaient pas toujours bien maitrisées par ces praticiens. Par exemple, nous avons observé dans notre enquête que peu des médecins savaient que l’arrêt du tabac (29.3%), la réduction de la consommation d’alcool (39%) et la réduction pondérale (4.9%) faisaient partie des MHD adressées aux hypertendus. Selon les travaux de Chen et al., quatre problèmes principaux entravaient la prescription des MHD par les généralistes: la mauvaise observance des patients, le manque de temps de consultation à consacrer à l’éducation sur les MHD, le manque de compétence du médecin en matière de MHD et le manque de technique pour enseigner ces MHD.12 Les classes pharmacologiques les plus prescrites en monothérapie dans notre étude étaient les inhibiteurs calciques suivis des inhibiteurs de l’enzyme de conversion, les diurétiques thiazidiques et les bétabloquants. Au Cameroun, c’étaient les diurétiques de l’anse qui étaient les plus prescrits (49.3%).7 Au Pakistan, par contre, on a observé curieusement une forte prescription d’anxiolytiques comme médication de première intention contre l’HTA.16 Actuellement les classes d’antihypertenseurs recommandées en première intention par les sociétés savantes restent les diurétiques thiazidiques, les inhibiteurs calciques, les inhibiteurs de l’enzyme de conversion, les antagonistes des récepteurs de l’angiotensine II et les bétabloquants.9,13,14 Il existe une préférence pour les inhibiteurs calciques et les thiazidiques chez les sujets de race noire.18 L’étude ALLAT a montré que les thiazidiques confèrent les mêmes bénéfices que les autres classes thérapeutiques recommandées mais ils sont supérieurs aux bétabloquants, aux inhibiteurs calciques et aux inhibiteurs de l’enzyme de conversion dans la prévention des accidents vasculaires cérébraux et de l’insuffisance cardiaque.19 Ces bénéfices pronostiques s’ajoutent au faible coût des thiazidiques. L’accent devrait donc être mis sur la prescription des thiazidiques dans nos pays à faible revenu. Bien entendu, il faudra tenir compte de leurs effets secondaires métaboliques et les prescrire à faible dose et surtout en association avec d’autres classes d’antihypertenseurs, notamment les inhibiteurs calciques et les IEC. L’ensemble de ces données de la littérature sur la prise en charge de l’HTA par les généralistes doit être considérée comme une alerte. Une riposte à cette situation désastreuse s’impose. Elle doit consister en une formation universitaire et post universitaire. Une priorité doit être donnée aux maladies non transmissibles et particulièrement l’HTA dans les curricula de formation des médecins généralistes dans nos pays. De même les autorités politico-administratives et l’ordre des médecins devraient soutenir les formations post universitaires des médecins praticiens. Ce besoin de formation a été clairement exprimé par les généralistes dans l’enquête de Chen et al. en Chine en 2011.12 La négligence de cette incapacité des généralistes à gérer l’HTA contribuera forcement à un mauvais contrôle de cette pathologie comme l’ont démontré quelques travaux.16,20-22

Conclusion La connaissance des généralistes sur la prise en charge de l’HTA était insuffisante et leur gestion ne respectait pas les recommandations internationales.

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Lafay V, Diarra M, Coulibaly S, Coulibaly S, Desplats D. Mise en place du réseau hypertension du MALI (RHYTM). Med Trop 2006; 66: 437–442.

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Falaschetti E. Blood pressure. In: Sproston K, Primatesta P, eds. Health Dzudie A, Kengne AP, Muna WF, Ba H, Menanga A, Kouam Kouam

Joint National Committee VI. Sixième rapport du Joint National Commitee sur la prévention la détection, l‘évaluation et le traitement de l’hypertension artérielle. Arch Int Med 1997; 157: 2413–2446. Noubiap JJN, Jingi AM, Veigne SW, Onana AE, Yonta EW, Kingue S. Approach to hypertension among primary care physicians in the West Region of Cameroon: substantial room for improvement. Cardiovasc Diagn Ther 2014; 4(5): 357–364.

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G, et al. 2007 Guidelines for the management of arterial hypertension:

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C, et al. Prevalence, awareness, treatment and control of hypertension in

12. Chen Q, Zhang X, Gu J, Wang T, Zhang Y, Zhu S. General practition-

Murray CJ, Lopez AD. Global mortality, disability, and the contribu-

Survey for England. London: TSO, 2003: 181–220. 5.

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of Cardiology (ESC). J Hypertens 2007; 25(6): 1105–1187. 15. Gallois P, Vallée JP, Charpentier JM. Risque cardiovasculaire ‘global’: intérêts et limites pour la décision médicale. Médecine 2006; 2(3): 112–117. 16. Jafar TH, Jessani S, Jafary FH, Ishaq M, Orkazai R, Orkazai S, et al. General practitioners’ approach to hypertension in urban Pakistan: disturbing trends in practice. Circulation 2005; 111: 1278–1283. 17. Parker A, Nagar B, Thomas G, et al. Health practitioners’state of knowledge and challenges to effective management of hypertension at primary level. Cardiovasc J Afr 2011; 22: 186–90. 18. Richardson AD, Piepho RW. Effect of race on hypertension and antihypertensive therapy. Int J Clin Pharmacol Ther 2000; 38(2): 75–79. 19. ALLHAT officers and coordinators for the ALLHAT Collaborative

Safar A. Al-Saleem, Abdullah Al-Shahrani, Yahia M. Al-Khaldi.

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2560–2572. 10. Rehman A, Rehman T, Shaikh MA, Naqvi SA. Awareness of hypertension among the medical students and junior doctors: a multicenter study from Pakistan. J Pak Med Assoc 2011; 61(11): 1153–1157. 11. Zabsonré P, Sanou G, Avanzini F, Tognoni G. Knowledge and perception of cardiovascular risk factors in Africa South of the Sahara. Arch Mal Coeur Vaiss 2002; 95(1): 23–28.

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Case Report A rare giant pericardial cyst mimicking a paracardiac mass Hakan Akbayrak, Serkan Yildirim, Murat Simsek, Mehmet Oc

Abstract Pericardial cysts are rare benign lesions of the thoracic cavity and are mostly congenital anomalies. They are induced by an incomplete coalescence of foetal lacunae during the development of the pericardium. Pericardial cysts are usually unilocular, well marginated spherical or teardrop shaped and may be attached to the pericardium directly or by a pedicle. Of all pericardial cysts, 70 to 75% are located at the right cardiophrenic angle. We report a case that was incidentally diagnosed with only chest magnetic resonance imaging because of a paracardiac mass. In order to prevent complications, the giant pericardial cyst was excised outside of the pericardium with median sternotomy. Keywords: pericardial cyst, surgical treatment, sternotomy Submitted 27/10/15, accepted 1/3/16 Published online 12/7/16 Cardiovasc J Afr 2016; 27: e7–e9

www.cvja.co.za

DOI: 10.5830/CVJA-2016-016

Pericardial cysts occur infrequently, with an incidence of one in 100 000 individuals. Mostly they are congenital anomalies but may also be acquired pericardial anomalies (e.g. postinflammatory, hydatid, neoplastic). Pericardial cysts are induced by an incomplete coalescence of foetal lacunae during the development of the pericardium.1,2 Pericardial cysts are usually unilocular, well marginated spherical or teardrop shaped and may be attached to the pericardium directly or by a pedicle.3 Of all pericardial cysts, 70 to 75% are located at the right cardiophrenic angle, and the rest are on the left side of the mediastinum.4 They contain clear serous fluid that is called ‘spring water’.5 Histologically, these cysts contain a single layer of mesothelial cells, with the remainder of the wall composed of connective tissue with collagen and elastic fibres.6 Department of Cardiovascular Surgery, Faculty of Medicine, Selcuk University, Konya, Turkey Hakan Akbayrak, MD, hakanakbayrak@yahoo.com Serkan Yildirim, MD Murat Simsek, MD Mehmet Oc, MD

Occasionally, pericardial cysts may alter cardiovascular haemodynamics and/or pulmonary expansion, producing signs and symptoms mimicking tricuspid stenosis, pulmonary stenosis or constrictive pericarditis.7 Pericardial cysts occur most frequently in the third or fourth decade of life and the incidence of cases is equal in men and women.8 We report on a patient with an extremely large pericardial cyst that was connected to the right atrium. In order to prevent complications, it was excised outside the pericardium with median sternotomy.

Case report A 48-year-old man presented with a dry cough. He was incidentally diagnosed with a paracardiac mass with chest magnetic resonance imaging and referred to our hospital. In the right thoracic cavity, there was a heterogenous hyper-intense mass, which seemed to be connected with the right atrium (Fig. 1). The mass extended from the superior to the inferior vena cavae, outside the pericardium, in the right thoracic cavity. On examination, his vital signs were as follows: pulse 96 beats/min, blood pressure 140/80 mmHg, body temperature 37.2°C, respiratory rate 18 /min and oxygen saturation on room air was 97%. The white blood cell count was 7 000 cells/µl. His respiratory and heart sounds were normal. The electrocardiogram (ECG) and dobutamine stress echocardiography were normal. Two-dimensional echocardiography from the subcostal view showed a giant homogenous hypo-echoic mass that extended from the superior to the inferior vena cavae. Because of the concern that the mass was connected with the right atrium, we operated on the patient with a median sternotomy and excised the giant paracardiac cystic mass outside the pericardium. The mass measured 27 × 5 × 2 cm (Fig. 2), was well marginated and teardrop shaped, and was attached to the right superior-lateral pericardium by a pedicle. Pathological examination confirmed the diagnosis of a fibrolipomatous cyst wall with no evidence of malignancy or tissue other than pericardium. There were no other complications in the postoperative period. Our patient was discharged on the seventh day postoperatively.

Discussion Pericardial cysts are rare, mostly benign, congenital lesions of the mediastinum, but they may also be acquired pericardial anomalies (e.g. post-inflammatory, hydatid, neoplastic).1,2 They occur most frequently in the third or fourth decade of life, and

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Fig. 1. A xial (A), coronal (B) and sagittal (C) views of T2-weighted images in the paracardiac area showing a heterogeneous hyperintense mass (arrows).

occur equally among men and women. The cysts range in size from 2–3 cm, up to a maximum of 28 cm reported by Braude et al.8 In our case, the pericardial cyst measured 27 × 5 × 2 cm. Most of these cases, including ours, are asymptomatic and are diagnosed incidentally on chest X-ray.4 The absence of symptoms at diagnosis is a good prognostic sign. However, patients may be admitted to a hospital with symptoms of chest discomfort or pain, cough, dyspnoea, or palpitation due to compression of the heart.9,10 Life-threatening complications, including cardiac tamponade, obstruction of the right main stem bronchus, cyst infection with cardiac or large vessel erosion and sudden death may be encountered. Cardiac tamponade generally occurs due to intrapericardial rupture of the cyst. Spontaneous cyst rupture and A

significant haemorrhage into the cysts have been reported, but these have not been linked adversely to cyst size. Asymptomatic cysts of this size are an unusual phenomenon.11 Other reported complications include right ventricular outflow tract obstruction, pulmonary stenosis, atrial fibrillation, congestive heart failure, and even sudden death after a stress test.12-14 Pericardial cysts usually follow a benign course in the majority of cases. There are no reports of malignant transformation. For asymptomatic patients, conservative management with short follow-up periods is recommended.15 Treatment is needed when symptoms or complications occur, and the management of those patients should be performed in the light of clinical characteristics. Indications for surgical resection of pericardial cysts include large size, symptoms, cyst infection, patient request, suspected malignancy, and prevention of complications.10,14,16 Other treatment options for pericardial cysts include simple observation, excision by thoracotomy, thoracoscopic surgical removal, and percutaneous aspiration with injection of a sclerosing agent. Although our patient was asymptomatic, surgical excision was planned due to the large size of the cyst and the concern that the mass was connected to the right atrium.

Conclusion B

Conservative management with short follow-up periods is recommended for asymptomatic patients with pericardial cysts. However, surgery should be considered for patients who become symptomatic and there is doubt about the paracardiac mass. Our patient was unusual because of a rare giant pericardial cyst mimicking a paracardiac mass. This article was presented at the 10th International Congress: Update in Cardiology and Cardiovascular Surgery (UCCVS) in March 2014.

References Fig. 2. Image of the excised cystic mass.

Losanoff JE, Richman BW, Curtis JJ, Jones JW. Cystic lesions of the pericardium. Review of the literature and classification. J Cardiovasc

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Surg 2003; 44: 569–576. 2.

Satur CM, Hsin MK, Dussek JE. Giant pericardial cysts. Ann Thorac 1996; 61: 208–210.

Strollo DC, Rosado-de-Christenson ML, Jett JR. Primary mediastinal tumors: part II. Tumors of the middle and posterior mediastinum. Chest 1997; 112: 1344–1357.

Nina VJ, Manzano NC, Mendes VG, Salgado Filho N. Giant pericardial cyst: case report. Rev Bras Cir Cardiovasc 2007; 22: 349–351.

Kaklikkaya I. A giant pericardial cyst. Cardiovasc J Afr 2011; 22: e1–3.

Abad C, Rey A, Feijóo J, Gonzalez G, Martín-Suarez J. Pericardial cyst. Surgical resection in two symptomatic cases. J Cardiovasc Surg 1996; 37: 199–202.

1418. 10. Moratalla MB, Garcia LG, Salvador RL, Bisquert BC. Giant pericardial cyst located at the left cardiophrenic angle. Eur J Radiol Extra 2008; 68: e111–e112. 11. Modi S, Chenzbraun A, Fewins H, Binukrishnan S, Ramsdale DR. Giant asymptomatic pericardial cyst. J Cardiovasc Med (Hagerstown) 2009; 10(8): 646–648. 12. Celik T, Firtina S, Bugan B, Sahin MA, Ors F, Iyisoy A. A giant pericardial cyst in an unusual localization. Cardiol J 2012; 19: 317–319. 13. Patel J, Park C, Michaels J, Rosen S, Kort S. Pericardial cyst: case reports and a literature review. Echocardiography 2004; 21: 269–272. 14. Simsek H, Gunes Y, Akil MA, Bilsel T. Asymptomatic giant pericar-

al. Task force on the diagnosis and management of pericardial diseases

dial cyst mimicking dextrocardia on chest X-ray. Herz 2013; 39(8):

management of pericardial diseases executive summary. Eur Heart J 2004; 25: 587–610. Braude PD, Falk G, McCaughan BC, Rutland J. Giant pericardial cyst. Aust NZ J Surg 1990; 60: 640–641. 9.

(eds). Cardiovascular Medicine, 3rd edn. Philadelphia: Springer, 2007:

Maisch B, Seferović PM, Ristić AD, Erbel R, Rienmüller R, Adler Y, et of the European society of cardiology. Guidelines on the diagnosis and

McAllister HA, Buja LM, Ferrans VJ. Pericardial disease. Anatomic abnormalities. In: Willerson JT, Cohn JN, Wellens HJJ, Holmes DR Jr

1013–1015. 15. Vlay SC, Hartman AR. Mechanical treatment of atrial fibrillation: removal of pericardial cyst by thoracoscopy. Am Heart J 1995; 129: 616–618. 16. Thanneer L, Saric M, Perk G, Mason D, Kronzon I. A giant pericardial cyst. J Am Coll Cardiol 2011; 57: 1784.

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Case Report Simultaneous presentation of giant aneurysms of the coronary sinus and superior vena cava Yan Cheng, Huanhuan Gao, Zhelan Zheng, Yun Mou Case report

Abstract Aneurysms of the coronary sinus and superior vena cava are rare and their aetiologies remain controversial. Some studies have shown that these acquired venous aneurysms are caused by an increase in right atrial pressure, which may be related to right heart failure. However, few reports have provided direct evidence to support this hypothesis. We present a rare case of combined giant aneurysms of the coronary sinus and vena cava, diagnosed using multiple imaging modalities. This case strongly supports the hypothesis that right heart diastolic failure may be an important mechanism underlying the pathogenesis of combined giant aneurysms. Keywords: aneurysm, coronary sinus, superior vena cava, right heart failure Submitted 2/11/15, accepted 11/3/16 Published online 12/4/16 Cardiovasc J Afr 2016; 27: e10–e13

www.cvja.co.za

DOI: 10.5830/CVJA-2016-031

Venous aneurysms are rare and the simultaneous presentation of aneurysms localised in the coronary sinus (CS) and superior vena cava (SVC) has not been previously reported. Venous aneurysms may be congenital or secondary to anomalous drainage.1,2 Some studies have shown that acquired venous aneurysms are most likely caused by an increase in right atrial pressure and right heart failure.3 However few reports have provided direct evidence supporting this pathogenic mechanism. Here, we present a case of giant aneurysms localised in both the CS and SVC. Our case provides strong evidence for the possible role of longstanding right heart failure during the pathogenesis of combined giant venous aneurysms.

Echocardiography and Vascular Ultrasound Centre, the First Affiliated Hospital, College of Medicine, Zhejiang University, Hangzhou, China Yan Cheng, MD Huanhuan Gao, MD Zhelan Zheng, PhD Yun Mou, PhD, yunmou2015@126.com

A 22-year-old woman was referred to our hospital because of a large mass in the left thorax, detected on chest X-ray. She had a long history of constrictive pericarditis and had undergone pericardiectomy seven years earlier. On presentation, a physical examination revealed severe facial and lower-extremity oedema, hepatomegaly and ascites. She had no cardiac murmur and no cyanosis, with an oxygen saturation of 96.6% on room air. Transthoracic echocardiography showed an enlarged right atrium and a huge cavity behind the left heart (Fig. 1A). Pulsedwave Doppler identified a restrictive transmitral inflow pattern. Tissue Doppler imaging (TDI) velocities at the septal mitral annulus showed that early diastolic myocardial velocity (e′) and systolic myocardial velocity (s′) were 5.85 and 10.4 cm/s, respectively (Fig. 1B). Transoesophageal echocardiography revealed a large cystic cavity with spontaneous echo contrast attached to the posterior wall of the left heart and communication with the right atrium. Colour Doppler flow imaging demonstrated a to-and-fro flow between the cavity and right atrium (Fig. 2A). Contrastenhanced computed tomography (CT) showed a giant cavity (10.4 × 7.3 cm) connected with three cardiac veins (Fig. 3A) and revealed that it was an aneurysm of the CS. Moreover, CT showed no evidence of pericardial calcification or thickening. CT also revealed a dilated inferior vena cava (IVC), aneurysmal dilated SVC (7.4-cm diameter, Fig. 3B), and a thrombus in the SVC and left inferior pulmonary artery. Cardiac catheterisation showed the shape of the CS aneurysm (Fig. 2B). The right heart pressure was recorded, including pulmonary arterial pressure (25/22 mmHg), right ventricular pressure (25/21 mmHg), mean right atrial pressure (23 mmHg) and mean CS aneurysm pressure (22 mmHg). The family declined a request for limited biopsy. No other associated cardiac abnormalities or defects were noted. Based on the TDI evidence of intact active relaxation and the presence of diastolic equalisation of pressures, it was decided that right heart diastolic failure was the dominant factor contributing to the patient’s symptoms. We reviewed the patient’s earlier images. Transthoracic echocardiography performed four years prior to presentation showed that the aneurysmal dilated CS was 5 cm in diameter (Fig. 4B) and the dilated SVC was 3 cm in diameter. Septal mitral annulus e′ and s′ velocity were 6.54 and 19.5 cm/s, respectively (Fig. 4A). These features suggested that the patient had had a long history of right heart diastolic failure, and this may have contributed to the progression of the combined giant aneurysms.

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s′

LV AO LA CSA

e′ a′

Fig. 1. T ransthoracic echocardiography on presentation showed: A. the appearance of the giant aneurysm of the coronary sinus in the parasternal long-axis view; B. tissue Doppler imaging velocities at the septal mitral annulus: early diastolic myocardial velocity (e′) was 5.85 cm/s and systolic myocardial velocity (s′) was 10.4 cm/s. RV, right ventricle; LA, left atrium; LV, left ventricle; CSA, coronary sinus aneurysm; AO, ascending artoa, a′, late diastolic myocardial velocity.

A diagnosis of combined giant aneurysms of the CS and SVC accompanied by right heart failure was made. Due to the two giant venous aneurysms, cardiac thrombosis and severe right heart failure, cardiac transplantation was considered the only viable option. At the time of publication of this report, the patient was being maintained on aspirin and furosemide while awaiting cardiac transplantation.

Discussion Venous aneurysms are uncommon, particularly those involving the CS or SVC. The aetiologies of venous aneurysms are still under debate. The causes of acquired venous aneurysms may A

include trauma, inflammation or pathological processes that affect the vascular wall.4 Longstanding venous hypertension secondary to heart failure, tricuspid valve lesions, cardiomyopathy and constrictive pericarditis have also been considered to cause vascular damage.5 Mahmud and colleagues found that CS size was positively correlated with right atrial size and pressure.6 Dilatation of the vena cava may be observed in right heart failure. Wells et al. reported a massive aneurysm of the IVC, and the patient’s longstanding right heart failure was presumed to be the underlying cause.7 We report a case of combined giant venous aneurysms, diagnosed using multiple imaging modalities, which provided B

IVS

CSA CSA

Fig. 2. A . Transoesophageal echocardiography colour Doppler flow imaging demonstrated flow from the coronary sinus aneurysm to the right atrium. B. Cardiac catheterisation showed the appearance of the giant coronary sinus aneurysm. RA, right atrium; IVC, inferior vena cava; CSA, coronary sinus aneurysm.

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** *

Fig. 3. C ontrast-enhanced CT showed a giant aneurysm of the coronary sinus (*) and a giant aneurysm of the superior vena cava (**).

strong evidence for an aetiology of longstanding right heart diastolic failure. Aneurysm of the CS is a rare abnormality of the intracardiac vein.8 Previous cases in the literature were considered to be congenital or secondary to anomalous drainage.9,10 We excluded the possibility of congenital causes on the basis of previous CT examinations. Echocardiography and contrast-enhanced CT gave a precise anatomical view of the CS and SVC, demonstrating the absence of anomalous drainage. Constrictive pericarditis (CP) is pathologically characterised by scarring and a loss of pericardium elasticity, resulting in an external interruption of cardiac filling.11 Pericardiectomy A

remains the most effective therapy for CP. Diastolic dysfunction and low-output syndrome occur in a considerable number of patients after pericardiectomy, which may be the result of atrophic changes in the myocardium associated with longstanding pericardial restriction.12 Our patient had a long history of CP before pericardiectomy. On the basis of previous echocardiography, we concluded that the patient had had a long history of right heart diastolic failure. Echocardiography also revealed the progression of combined venous aneurysms. Colour Doppler echocardiography revealed to-and-fro flow between the CS aneurysm and right atrium. Catheter examination demonstrated equally increased pressure in B

CSA

s′

e′ a′

Fig. 4. T ransthoracic echocardiography performed four years prior to presentation showed: A. the aneurysmal dilated CS was 5 cm in diameter; B. tissue Doppler imaging velocities at the septal mitral annulus: early diastolic myocardial velocity (e′) was 6.54 cm/s and systolic myocardial velocity (s′) was 19.5 cm/s. RV, right ventricle; RA, right atrium; CSA, coronary sinus aneurysm; a′, late diastolic myocardial velocity.

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the right atrium and CS aneurysm. Therefore, it was reasonable to presume that the main cause of the two venous aneurysms was longstanding right heart diastolic failure. Based on the high pressure of the two aneurysms and complications of severe right heart failure, cardiac transplantation may be the only treatment option for this patient. TDI was helpful in the diagnosis. TDI is a contemporary echocardiographic tool that allows the measurement of intrinsic myocardial velocity. The e′ velocity reflects early diastolic ventricular relaxation in the longitudinal plane, and the s′velocity reflects systolic function in the longitudinal plane. Values of e′ (septal) < 8 cm/s are suggestive of impaired myocardial relaxation.13 In this patient, two TDI echocardiography examinations both showed a reduced e′ velocity and a normal s′ velocity. These results strongly suggested a long history of right heart diastolic failure.

Conclusion

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Thummar AC, Phadke MS, Lanjewar CP, et al. Supracardiac total anomalous pulmonary venous drainage with giant superior vena cava aneurysm: a rare combination. J Am Coll Cardiol 2014; 63: e51.

Sasaki T, Kubo T, Miyamoto T, et al. Clinical significance of measuring inferior vena cava dimension in patients with acute exacerbation of chronic heart failure. J Cardiol 2001; 37: 309–313.

Sullivan VV, Voris TK, Borlaza GS, et al. Incidental discovery of an inferior vena cava aneurysm. Ann Vasc Surg 2002; 16: 513–515.

Elliot A, Henn A, Pamuklar E, et al. Aneurysm of the inferior vena cava: case report. Abdom Imaging 2006; 31: 457–460.

Mahmud E, Raisinghani A, Keramati S, et al. Dilatation of the coronary sinus on echocardiogram: Prevalence and significance in patients with chronic pulmonary hypertension. J Am Soc Echocardiogr 2001; 14: 44–49.

Well IT, Bhatnagar R: Presumed rupture of a massive inferior vena cava aneurysm associated with right heart failure:a unique case. Clin Radiol 2008; 63: 1181–1183.

To our knowledge, this is the first reported case of simultaneous presentation of giant aneurysms of the coronary sinus and superior vena cava. Acquired venous aneurysms may result from longstanding right heart failure. The combination of echocardiography, contrast-enhanced CT and cardiac catheterisation facilitated the diagnosis of combined giant venous aneurysms and provided strong evidence for this aetiology.

Thal S, Thai H, Juneman E, et al. Coronary sinus diverticulum complicating CRT device implantation. Pacing Clin Electrophysiol 2008; 31: 1184–1185.

Gerlis LM, Davies MJ, Boyle R, et al. Pre-excitation due to accessory sinoventricular connexions associated with coronary sinus aneurysms: a report of two cases. Br Heart J 1985; 53: 314–322.

10. D’Cruz IA, Shala MB, Johns C. Echocardiography of the coronary sinus in adults. Clin Cardiol 2000; 23: 149–154. 11. Sengupta PP, Eleid MF, Khandheria BK. Constrictive pericarditis. Circ J 2008; 72: 1555–1562.

This study was supported by grants from the National Natural Science Foundation, People’s Republic of China (No. 81371571).

12. Chowdhury UK, Subramaniam GK, Kumar AS, et al. Pericardiectomy for constrictive pericarditis: A clinical, echocardiographic, and hemodynamic evaluation of two surgical techniques. Ann Thorac Surg 2006;

References 1.

81: 522–529. 13. Nagueh SF, Appleton CP, Gillebert TC, et al. Recommendations for the

Funabashi N, Asano M, Komuro I. Giant coronary sinus diverticu-

evaluation of left ventricular diastolic function by echocardiography. J

lum with persistent left superior vena cava demonstrated by multislice

Am Soc Echocardiogr 2009; 22:107–133.

computed tomography. Int J Cardiol 2006; 111: 468–469.

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Case Report Successful continuous-flow left ventricular assist device implantation with adjuvant tricuspid valve repair for advanced heart failure Chih-Hsien Lee, Jeng Wei

Abstract The prevalence of end-stage heart failure (HF) is on the increase, however, the availability of donor hearts remains limited. Left ventricular assist devices (LVADs) are increasingly being used for treating patients with end-stage HF. LVADs are not only used as a bridge to transplantation but also as a destination therapy. HeartMate II, a new-generation, continuous-flow LVAD (cf-LVAD), is currently an established treatment option for patients with HF. Technological progress and increasing implantation of cf-LVADs have significantly improved survival in patients with end-stage HF. Here we report a case of a patient with end-stage HF who was successfully supported using cf-LVAD implantation with adjuvant tricuspid valve repair in a general district hospital. Keywords: cf-LVAD, end-stage heart failure, HeartMate II Submitted 19/9/15, accepted 16/3/16 Published online 12/4/16 Cardiovasc J Afr 2016; 27: e14–e16

www.cvja.co.za

DOI: 10.5830/CVJA-2016-034

HeartMate II is a new-generation, continuous-flow left ventricular assist device (cf-LVAD, Thoratec, Pleasanton, CA, USA) used as a bridge to transplantation (BTT) and as a destination therapy (DT) in patients with end-stage heart failure (HF).1 Applying continuous-flow technology to mechanical circulatory support systems has revolutionised the treatment of end-stage HF. The use of implantable mechanical circulatory support devices, such as cf-LVAD, has increased in recent years.2 Safer long-term cf-LVAD support has been achieved because of improved outcomes.1 Patients supported for BTT or DT using cf-LVADs have an overall reduction in life-threatening complications and have prolonged survival time, with an active

Department of Cardiac Surgery, Tungs’ Taichung MetroHarbor Hospital; Department of Biological Science and Technology, National Chiao-Tung University; Department of Surgery, National Defense Medical Centre, Taiwan Chih-Hsien Lee, MD, jamesolee@yahoo.com.tw

Heart Centre, Cheng Hsin General Hospital, Taiwan Jeng Wei, MD, MSD

lifestyle and an acceptable quality of life. Cardiac transplant recipients can safely wait for extended periods while their status at transplant is optimised, aiding post-transplant survival. In this case report, we describe a patient with end-stage HF who was successfully supported using cf-LVAD implantation with adjuvant tricuspid valve repair.

Case report A 39-year-old man was admitted to our hospital because of deteriorating heart function despite maximal medical treatment. He had a body surface area of 1.88 m2, had suffered from advanced HF following dilated cardiomyopathy, and had been waiting for a heart transplant for four years. In addition, he had diabetes mellitus requiring insulin control. On examination, he exhibited bilateral grade IV pitting oedema of the lower limbs with bilateral pleural effusion and ascites. Laboratory tests showed a total bilirubin level of 6.2 mg/dl, aspartate aminotransferase level of 109 U/l, and blood creatinine level of 1.8 mg/dl. Echocardiography revealed a left ventricular (LV) diastolic diameter of 67 mm, LV systolic diameter of 58 mm, and ejection fraction of 19% with severe tricuspid regurgitation (TR). A pulmonary artery catheter was inserted to measure pulmonary artery systolic pressure, pulmonary capillary wedge pressure, central venous pressure and cardiac index, which were found to be 34, 50 and 36 mmHg, and 0.5 l/min/m2, respectively. This facilitated optimising the use of dobutamine and dopamine to decrease the right ventricular (RV) afterload while maintaining ventricular contractility. An intra-aortic balloon pump was inserted at the same time because the urine output had decreased. Furosemide was continuously administered to obtain an adequate urine output and to decrease central venous pressure to < 24 mmHg. Right ventricular failure (RVF) is common after cf-LVAD implantation and is a leading cause of morbidity and death after cf-LVAD implantation. Five days after admission, tricuspid valve repair was performed using the de Vega annuloplasty procedure under anaesthesia. HeartMate II was implanted under cardiopulmonary bypass. The alignment of the mitral valve and inflow cannula was checked using transoesophageal echocardiography. Initial pump flow was at 6 000 rpm, and we gently needlepunctured the cf-LVAD outflow tract to assist in de-airing the intracardiac air bubbles. The needle and clamp were removed, cardiopulmonary bypass was terminated, and the pump speed

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was increased from 6 000 to 9 200 rpm; the pump flow was 4.5 l/m, pulse index was 5.7, pump power was 5.5 W, and mean blood pressure was 60 mmHg. Protamine was administered slowly. The intra-aortic balloon pump was removed, and an open repair of the left femoral artery was performed. Dobutamine and dopamine maintained the RV contractility and decreased the RV afterload. The patient’s condition stabilised, and he was transferred to the intensive care unit. The endotracheal tube was removed the next day. More than 12 hours following cf-LVAD implantation, when the chest tube drainage decreased to ≤ 50 ml/h and the coagulation profile returned to normal levels, an intravenous heparin infusion was started to maintain activated partial thromboplastin time between 50 and 70 s. Aspirin (100 mg) was administered once daily after extubation, and warfarin was administered to maintain the international normalised ratio (INR) between 2.0 and 3.0. Heparin infusion was continued until the INR target range was attained. He was discharged one month after the operation and was categorised as New York Heart Association functional class II.

Discussion The prevalence of end-stage HF is on the increase, however, the availability of donor hearts remains limited. Therefore, the number of patients requiring long-term support with cf-LVAD implantation has increased. HeartMate II is a new-generation cf-LVAD used as BTT and DT in patients with end-stage HF.1,3 The waiting time for cardiac transplant recipients has increased, and so has BTT by using the support device in clinical settings. We occasionally encounter patients who require unexpected long-term device support. In addition to being a life-saving treatment, cf-LVADs currently also provide long-term survival with favourable quality of life for patients with severe HF. Consequently, long-term cf-LVAD implantation has become a valuable alternative to cardiac transplantation for treating end-stage HF. Studies have reported that post-transplant survival at one, two, five and 10 years is approximately 90, 80, 70 and 50%, respectively.1 The survival of patients receiving DT with cf-LVADs within this cohort at one, three and five years was 80–83, 75 and 61%, respectively.1,4 Prolonged post-transplant survival in patients receiving BTT can reduce the need for cardiac transplantation as a first-line replacement therapy. However, it is too premature to draw conclusions about survival comparisons because of the lack of head-to-head comparative data. Furthermore, considering the frequent readmissions in this population, patient survival, quality of life, and healthcare costs should be considered before drawing conclusions.1 Current cf-LVADs provide satisfactory long-term survival, but rehospitalisation for specific reasons is common in this population.1 Despite this progress, cf-LVAD implantation is still associated with a risk of complications, which challenges patient selection and adversely impacts on outcomes. The incidence of RVF in this population ranges from 10 to 50% and is a risk factor for peri-operative and postoperative mortality and morbidity in patients undergoing cf-LVAD implantation.2,4,5 Moreover, some reports have suggested that cf-LVAD implantation in patients with pre-operative hepatic failure entails considerable mortality.2 cf-LVAD recipients who develop

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postoperative RVF have poor outcomes, including increased incidences of multi-organ failure, postoperative haemorrhage, pulmonary complications, thromboembolic events, and migration of intracardiac air to the coronary arteries, causing transient myocardial ischaemia.5 In this setting, the function of the right heart becomes critical to patient survival, and RVF remains a considerable postoperative complication that affects mortality. In a previous study, 33.4% of patients experienced RVF postoperatively and 10–15% required RV support.2,5 RVF after cf-LVAD implantation is occasionally unavoidable in a deteriorated heart. Therefore RVF is a contraindication for receiving cf-LVAD implantation because it may require the use of a biventricular assist device. The setting of RVF is associated with a poor prognosis and influences early outcomes.5 The prediction and treatment of RVF are crucial to improve survival after cf-LVAD implantation. The ratio of central venous pressure to pulmonary capillary wedge pressure and secondary pulmonary hypertension is a critical predictor of RVF after cf-LVAD implantation; RVF significantly reduces survival after cf-LVAD implantation.5 Careful evaluation of central venous pressure, pulmonary capillary wedge pressure, and laboratory data may help to predict postoperative RVF. Furthermore, secondary TR is common among patients with RVF who undergo cf-LVAD implantation. Although the repair of TR in combination with cf-LVAD implantation is not an established approach, recent reports have suggested that concomitant tricuspid valve repair may reduce postoperative RVF.5 In our case, tricuspid valve repair was performed by the de Vega annuloplasty procedure to decrease the risk of RVF. To determine whether tricuspid valve repair is useful to rescue patients from possible RVF, a randomised study is required. HeartMate II may transiently worsen the right ventricular function in the initial post-implant period because of a higher cardiac output resulting in increased venous return and right ventricular preload.2 In addition, this initial temporary cholestasis resulting from RV dysfunction has been documented previously.2 Although laboratory parameters tend to normalise with successful cf-LVAD implantation, the effect appears to dissipate over time. A gradual improvement in RVF caused by improved LV unloading is observed in the majority of patients. Survival rates have increased because major adverse events, such as stroke, bleeding and infection, have decreased. The occurrence of thrombosis ranges from annualised rates of 2–4% and that of haemolysis ranges from 2–3%.3 The major highrisk factors, such as female gender, young age, implantation technique, and inflow cannula malposition, are related to the development of pump thrombosis.3 Other risk factors, such as sub-therapeutic INR, non-compliance, hypercoagulable disorder, and infection require pre-operative optimisation, intraoperative techniques, and postoperative management strategies.3 The pre-operative period of haemodynamic and fluid balance is optimised when possible. Some reports have evaluated heparin antibodies, aspirin resistance and baseline platelet function where possible.3 An adequate pump pocket depth is critical to allow favourable positioning of the cf-LVAD and inflow cannula angle, which should lie parallel to the septum and oriented to the central LV and mitral valve. Echocardiography is essential to enable surgeons and anesthaesiologists to make prompt decisions during cf-LVAD implantation and is necessary for detecting cardiac

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structural and functional abnormalities, such as ventricular dysfunction, valvular pathology, mural thrombosis, and atrial septal defect or patent foramen ovale. The pocket is made deep and lateral to allow the pump to be fixed below the diaphragm. The outflow cannula is placed to the right of the sternal midline with enough graft length to avoid any compression of the RV. The management of bleeding is indivisibly linked to the risk of thromboembolic events, and anticoagulation and antiplatelet therapies seem to be the only methods for carefully managing complication and individual risks. Further understanding of the mechanisms underlying bleeding and novel strategies, such as new anticoagulant drugs, are expected to play crucial roles in the long-term management of cf-LVAD therapy.1 Cardiac arrhythmia, such as ventricular arrhythmia, is also a common issue in the early and late periods after cf-LVAD implantation. Although such arrhythmias may not be lethal in the presence of cf-LVAD, it could put patients at a risk of RVF.1 Anti-arrhythmic medication, catheter ablation, intra-operative cryoablation, and implantable cardioverter–defibrillator may be employed to minimise the risk of recurrent arrhythmias.1 End-organ function was restored one month after the initiation of support.1 These improvements persisted throughout the support period; for example, the LV diastolic dimension significantly decreased and the TR ratio reduced from 45 to 22% at one month, except for the creatinine level.1

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Conclusion The implantation of cf-LVAD, either as BTT or DT, remains a critical treatment option for selected patients with end-stage HF.

References 1.

Takeda K, Takayama H, Kalesan B, Uriel N, Colombo PC, Jorde UP, et al. Long-term outcome of patients on continuous-flow left ventricular assist device support. J Thorac Cardiovasc Surg 2014; 148: 1606–1614.

Weymann A1, Patil NP, Sabashnikov A, Mohite PN, Garcia Saez D, Bireta C, et al. Continuous-flow left ventricular assist device therapy in patients with pre-operative hepatic failure: are we pushing the limits too far? Artif Organs 2015; 39: 336–342.

Klodell CT, Massey HT, Adamson RM, Dean DA, Horstmanshof DA, Ransom JM, et al. Factors related to pump thrombosis with the heartmate II left ventricular assist device. J Card Surg 2015; 30: 775–780.

Kalogeropoulos AP, Kelkar A, Weinberger JF, Morris AA, Georgiopoulou VV, Markham DW, et al. Validation of clinical scores for right ventricular failure prediction after implantation of continuousflow left ventricular assist devices. J Heart Lung Transplant 2015; 34: S1053–2498.

Sabashnikov A, Mohite PN, Weymann A, Patil NP, Hedger M, Sáez DG, et al. Outcomes after implantation of 139 full-support continuousflow left ventricular assist devices as a bridge to transplantation. Eur J Cardiothorac Surg 2014; 46: e59–66.

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Case Report An unusual condition during internal jugular vein catheterisation: vertebral artery catheterisation Ozge Korkmaz, Sabahattin Göksel, Burçak Söylemez, Kasım Durmuş, Ahmet Cemil İşbir, Öcal Berkan Case report

Abstract Vertebral artery cannulation is an unusual complication during internal jugular vein cannulation. We report a case of vertebral artery cannulation, which occurred during an attempt to cannulate the right internal jugular vein, and we discuss the management of such a rare complication. Keywords: vertebral artery, cannulation, internal jugular vein, anterior forminectomy, complication Submitted 20/11/15, accepted 30/3/16 Published online 27/5/16 Cardiovasc J Afr 2016; 27: e17–e19

www.cvja.co.za

DOI: 10.5830/CVJA-2016-040

Internal jugular vein catheterisation is an essential procedure, specifically in major operations, such as cardiac surgery. Vertebral artery cannulation is a rare complication during internal jugular vein cannulation, but it may lead to sequelae that could prove fatal. When this condition occurs, it should be diagnosed quickly and treated as soon as possible.1 This case discusses a patient who had inadvertent vertebral artery cannulation during internal jugular vein access in the operating room for coronary bypass surgery. The right vertebral artery was repaired via foraminectomy using an intraoperative anterior approach, and coronary bypass surgery was subsequently continued.

Department of Cardiovascular Surgery, Cumhuriyet University Medical Faculty, Sivas, Turkey Ozge Korkmaz, MD, ozgekorkmaz73@hotmail.com Sabahattin Göksel, MD Öcal Berkan, MD

Department of Neurosurgery, Cumhuriyet University Medical Faculty, Sivas, Turkey Burçak Söylemez, MD

Department of Head and Neck Surgery, Cumhuriyet University Medical Faculty, Sivas, Turkey Kasım Durmuş, MD

Department of Anaesthesilology and Reanimation, Cumhuriyet University Medical Faculty, Sivas, Turkey Ahmet Cemil İşbir, MD

A 65-year-old male patient who, following diagnosis of coronary artery disease, had angiography and was admitted to the cardiovascular surgery unit for bypass grafting on four vessels. Carotid and vertebral artery Doppler ultrasonography, performed during pre-operative preparation of the patient, revealed no pathology in either the carotid or vertebral arteries. After preparation, the patient was taken into the operating room for coronary bypass surgery. Peripheral venous and radial artery catheterisation was completed prior to general anesthaesia. Following the administration of general anaesthesia, the head of the patient was placed 15 degrees below the whole body, which was in the Trendelenburg position, and he was turned to the left. He was stained and covered under sterile conditions. The puncture was made by an anaesthetist, using a 18-G needle, through the apex of the triangle composed of the sternal and clavicular parts of the sternocleidomastoid muscle. During puncture, the blood was established not to be bright in colour or pulsatile, and a 12-F × 15-cm double-lumen catheter was inserted using the Seldinger method. However, the blood from the catheter was bright and pulsatile, and so the catheter tip was attached to a transducer. From the arterial trace and the arterial nature of the blood sample taken, it was assumed that carotid artery catheterisation had been performed. Doppler ultrasonography was carried out in the operating room on the patient under general anaesthesia. It was observed that the catheter was not in the carotid artery, and therefore the carotid artery was accessed by surgical exploration through an incision at the catheter tip. It was established from ultrasonography that the catheter had advanced deeper towards the cervical vertebrae. In the ensuing minutes, brain, and head and neck surgeons were invited to the operating room. With neck dissection, the anterolateral view of the C5–C6–C7 vertebrae was accessed. It was seen that the catheter had entered through the C5–C6 disc space and had gone into the C6 vertebral foramen. The catheter was found to be in the subclavian artery of C7. Foraminectomy was performed anterior to the foramen of the C6 vertebra. This region was cleared and it was established that the catheter had advanced through the vertebral artery and was in the subclavian artery (Fig. 1). The catheter was withdrawn in a controlled manner, the existing vascular damage was primarily repaired, and coronary bypass surgery was then initiated. During the operation, neurological soundness of the patient was followed up by the anaesthetists (pupil reflexes, absence of anisocoria, etc). The operation was completed without any problems and the patient was taken to the intensive care unit. He

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Fig. 1. A fter foraminectomy the catheter was clearly seen advancing through the vertebral artery. The black arrow shows the side of the catheter.

was awake and conscious at about postoperative hour four. A subsequent neurological examination revealed no pathology and the patient was extubated. Tomographic angiography performed postoperatively revealed that the right vertebral artery was present at the outlet but totally occluded from about 0.5 cm (Fig. 2). The patient was again examined neurologically but no neurological deficit was identified. It was therefore recommended that he continue his treatment of low-molecular-weight heparin. His cardiac medication and other medical treatment were planned and he was discharged on condition of follow-up visits.

Discussion Internal jugular vein catheterisation is preferred in patients undergoing open-heart surgery, since it is easy to cannulate and is associated with a reduced risk of complications during cannulation; it is also distant from the surgical site. The prevalence of vertebral artery cannulation during internal jugular vein catheterisation is unknown, probably because there are fewer cases reported than actually occur. When we reviewed the literature, it was found to be reported less often than carotid artery punctures. Carotid artery puncture during internal jugular vein cannulation has been reported at a rate of 0.5 to 11.4%, whereas the rate of vertebral artery cannulation was from 0.099 to 0.775%,2 and fewer than 30 cases were reported on iatrogenic vertebral artery cannulation.3 Dissection, thrombosis, formation of arteriovenous fistulae, and pseudo-aneurysms are complications of vertebral artery injury during vein cannulation.3 Diagnosis of pseudo-aneurysm of the vertebral artery is often delayed because symptoms occur only late after cannulation.4 In particular, some patients may develop fatal vertebrobasilar ischaemia due to vertebral artery cannulation and associated severe and damaging sequelae, such as stroke or visual defects, while others may be asymptomatic, which can be attributed to sufficient extracranial collateral circulation.5 The best way to avoid iatrogenic vertebral artery cannulation is to take the necessary precautions. In other words, the best way to

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Fig. 2. Three-dimensional tomographic angiography showing the right vertebral artery was present at the outlet but totally occluded for about 0.5 cm. The white arrow shows the right vertebral artery and the occluded part.

perform this procedure is under the guidance of ultrasonography, as recommended in many guidelines. However, this is almost impossible in emergent cases and when ultrasonography is not available or is difficult to access. In such cases, intervention may be performed using anatomical reference points. The vertebral artery is classically the first branch of the ipsilateral subclavian artery, and arises from the posteriorâ&#x20AC;&#x201C; superior part of this artery. The vertebral artery, after separating from the subclavian artery, generally passes through the transverse process of the C7 vertebra and superiomedially enters the transverse foramen of the cervical vertebrae (C6 in 95% of cases), extending vertically to the level of the C2 vertebra within the transverse foramen of the vertebrae.6 The extraforaminal region is about 4 cm and located deeper and more medially than the internal jugular vein. It is the region most open to injury, and in this region it is difficult to stop haemostasis with compression,7 because the vertebral artery courses in a deep plane. A safer approach when arterial damage is caused, is with an immediate surgical or endovascular stent application by a wide-scale catheter, as stated by Hong-liang et al.8 These kinds of punctures frequently result from hyperextension of the neck, accompanied by excessive rotation of the head, or failure to adjust the direction and depth of the puncture, or using an excessively long needle to perform the puncture. In addition, the non-pulsatile flow from the first puncture, due to the thinner diameter of the vertebral artery, may be misleading. Both the quantity and colour of the flow are manifested more clearly when the final tip reaches the subclavian artery following catheterisation, as in our case.

Conclusion It should always be considered that during percutaneous interventions, vertebral artery cannulation may occur, even if there is only a slight probability. In the event of such a case, there should be a management plan, previously prepared by the surgeon and anaesthetists. If endovascular repair is not possible in vertebral artery catheterisations, or if the artery is completely

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CARDIOVASCULAR JOURNAL OF AFRICA • Volume 27, No 4, July/August 2016

occluded by the catheter, the catheter should be explored without delay, together with brain and/or head and neck surgeons, and the necessary repair procedure should be competently performed on the vertebral artery.

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of foreseen hemorrhage during catheter withdraw. J Vasc Access 2014; 2.

2(3): 359–369.

caliber cannulas during jugular vein catheterization: case for surgical 3.

Vinchon M, Laurian C, George B, et al. Vertebral arteriovenous fistulas: a study of 49 cases and review of the literature. Cardiovasc Surg 1994;

Shah PM, Babu SC, Goyal A, et al. Arterial misplacement of largemanagment. J Am Coll Surg 2004; 198(6): 939–944.

Duan S, He E, Lv S, Chen L. Three-dimensional CT study on the anatomy of vertebral artery at atlantoaxial and intracranial segment.

central venous catheter in the vertebral artery: endovascular treatment 15(5): 418–423.

Arthur ME, Castresana MR, Paschal JW, et al. Acute cerebellar stroke after inadvertent cannulation and pulmonary artery catheter placement

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Bernik TR, Friedman SG, Scher LA, Safa T. Pseudoaneurysm of the subclavian– vertebral artery junction – case report and review of the

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Momly J,Vasquez J.Iatrogenic vertebral artery pseudoaneurysm due to

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