YL Basic Research and Young Investigator Research Lecture

YL-1

HEME-DERIVED BILINS: NEW THERAPEUTICS FOR INFLAMMATORY DISEASES

JON Y. TAKEMOTO, PH.D.

Professor of Biology; Department of Biology, Utah State University

In mammals, heme-derived bilins are most often associated with red blood cell destruction and hemoglobin degradation in the spleen and skin bruises. Bilins result from cleavage of heme by the enzyme heme oxygenase to generate biliverdin, bilirubin, and other bilins that are eventually eliminated in bile, feces and urine. The overall process reduces levels of heme which is toxic when accumulated. Bilins also occur - often in abundance - in microbes, plants and other animals where they play many roles. Discoveries of new and diverse roles for bilins are emerging such as sensing and and bilins are being explored for beneficial uses in medicine and agriculture. Most prominent is biliverdin which has protective effects against several diseases and inflammatory conditions. Discussed will be the experimental therapeutic effects of biliverdin (including against diabetes) in animal models, the discovery of its bilin analog mesobiliverdin, and cytoprotection by these bilins of pancreatic islets in islet transplantation procedures.

YL-2

INVESTIGATION OF THE CAUSES AND MANAGEMENT OF NONALCOHOLIC FATTY LIVER DISEASES

P-J HSIAO

Division of Endocrinology and Metabolism, Department of Internal Medicine, Kaohsiung Medical University, Taiwan, R.O.C

Non-alcoholic fatty liver disease (NAFLD), first described in 1980, is defined by the accumulation of hepatic fat (triglyceride) more than 5% without other recognized cause of fatty liver, e.g., alcohol, virus, drugs, autoimmunity or inherited disease. It is rapidly becoming the most common cause of chronic liver diseases, and is the second-most common indication for liver transplantation in the world. The global prevalence of NAFLD is estimated 25% and 3-18% in pediatric population. NAFLD is regarded to be the hepatic manifestation of metabolic syndrome with comorbidities with obesity (51.3%), type 2 diabetes (22.5%), hyperlipidemia (69.2%), hypertension (39.3%) and metabolic syndrome (42.5%). From simple steatosis, NAFLD may progress and worsen to nonalcoholic steatohepatitis (NASH, 30-40%) and further progress to cirrhosis (40-50%) and/or to hepatocellular hepatoma (HCC, 7%) within 10 years. The major risk factors for progression are well established, e.g., age > 50 yrs, obesity, insulin resistance, T2DM, increased ferritin, and PNPLA3 (1148M) polymorphism. The growing evidence has demonstrated NAFLD is a multisystem disease with an important burden of extra-hepatic chronic complications, such as metabolic syndrome, atherosclerosis cardiovascular disease (CVD), higher myocardial fat, cardiac arrhythmia, chronic kidney disease (CKD) and colon cancer. Cardiovascular disease is the leading cause of mortality in NAFLD patients, followed by non-hepatic malignancy and liver-related death (liver cirrhosis or hepatocellular carcinoma).

Pathophysiology of the hepatic fat accumulation originates from the “ectopic fat accumulation”, when positive energy imbalance has occurred with lipid accumulation besides adipose tissue. It is associated with increased diabetogenic hepatokines (retinol binding protein (RBP4), fibroblastic growth factor (FGF)-21) or inflammatory biomarkers (e.g., CRP, TNF-a, IL-6) to enhance gluconeogenesis, decrease glycogen synthesis and inhibit insulin signaling. Increasing evidence also suggests that dysbiosis of the gut microbiota plays a key role to regulate the intrahepatic metabolism progression of NAFLD. All the guideline has advocated with lifestyle intervention for NAFLD, such as Mediterranean diet, aerobic exercise, resistance training to target weight loss 7-10%. The potential

drugs are widely discordant by the guidelines, such as metformin, pioglitazone, Vitamin E, silymarin, GLP-1 analogues and statins, etc. Effective weight loss by bariatric surgery has successfully resolved years but with higher mortality. Because co-existing comorbidities is the progression or fatal cause of NAFLD, treatment strategy should focus the co-morbidities and lie on the management of the “insulin resistance” related metabolic syndrome but not NAFLD itself.

YL-3

INVESTIGATING THE ROLE OF GLUCOCORTICOID-HEPASSOCIN PATHWAY IN THE HIGH-FAT DIET-INDUCED OBESITY-RELATED METABOLIC DISORDERS

1H-C HUNG, 2Y-M KUO

1Division of Endocrinology and Metabolism, Department of Internal Medicine, National Cheng Kung University Hospital. 2Department of Cell Biology and Anatomy, College of Medicine, National Cheng Kung University

OBJECTIVE High levels of glucocorticoids, the primary stress hormone, play a critical role in the development of obesity. However, the role of the HFD-induced increment of corticosterone level in the HFD-induced metabolic disorders remains unclear.

METHODS We employed adrenalectomy in male C57BL/6 mice and subjected a transgenic C57BL/6-Cyp11a1tm2Bcc/Narl (L/L) mice which has normal basal corticosterone level but a blunted corticosterone response to stress stimuli to address this question.

RESULTS A 12-week HFD regimen induced obesity, impaired glucose metabolism, and elevated the circulating level of corticosterone in male C57BL/6 mice. Bilateral adrenalectomy attenuated the HFD-induced adipose expansion, hepatic steatosis and insulin resistance without affecting the food intake amounts in mice. The adrenalectomized mice had higher blood triglyceride level than the sham control mice after the 12-week HFD treatment. The preadipocyte marker, preadipocyte factor 1 (Pref-1), decreased in the epididymal white adipose tissue in the HFD mice but was restored by adrenalectomy. Re-supplementing corticosterone to the adrenalectomized mice abolished this adrenalectomy-induced effect on the Pref-1 level in the white adipose tissue. The L/L mice had lower peripheral expression of corticosterone than the wild-type mice after a 12-week HFD regimen. HFDinduced obesity and adipose expansion were milder in the L/L mice than those in the wild-type ones. Compared to the wild-type-HFD mice, the L/L-HFD mice had higher circulating triglyceride level and Pref-1 expression in the white adipose tissue.

CONCLUSIONS HFD-induced increase of corticosterone is essential for the adipogenesis during the process of HFD-induced adipose expansion.

YL-4

THE RELATIONSHIP BETWEEN SERUM ANGPTL6 AND INCIDENT DIABETES IN HUMAN – FOLLOW UP STUDY

1H-Y LI

1Division of Endocrinology and Metabolism, Department of Internal Medicine, National Taiwan University Hospital, Taipei, Taiwan, ROC

Type 2 diabetes mellitus is one of the most prevalent chronic diseases in the world and could lead to multiple complications to the patients such as cardiovascular disease, neuropathy, nephropathy and retinopathy. Recently, liver is found to secrete various hepatokines to regulate energy homeostasis and glucose metabolism, which is similar to the concept of adipokines. Angiopoietin-like 6 (ANGPTL6) is a novel hepatokine which increases energy consumption, lower body weight and improve insulin sensitivity, as shown by animal studies. In 2015, we analyze the relationship between serum ANGPTL6 and the incidence of diabetes in a longitudinal cohort, using the grant from the Diabetes Association of R.O.C. We found that high serum ANGPTL6 is associated with a lower incidence of diabetes, adjusted for age, gender, family history of diabetes, body mass index, and hemoglobin A1c, which is baseline in our cohort reveals that plasma ANGPTL6 is higher in subjects with pre-diabetes, compared with that in normal controls. Similarly, cross-sectional studies in the literature showed that serum ANGPTL6 were higher in subjects with diabetes than in healthy subjects.

We believed that hyperglycemia can stimulate serum ANGPTL6 secretion to maintain glucose homeostasis, which can be used to explain the results from cross-sectional studies. In the present study, we explored the changes of serum ANGPTL6 level after acute hyperglycemia in humans. Besides, we investigated regulation and function of ANGPTL6 in glucose metabolism in hepatic cell and rodent models.

By using the data and materials from the cohort study, we measured serum ANGPTL6 concentrations during 75g oral glucose tolerance tests (OGTT), including samples collected at fasting, 30-min and 120-min after OGTT. The result showed that serum ANGPTL6 increased after glucose challenge. Serum ANGPTL6 concentration at 120-minutes after 75gm OGTT was significantly higher than fasting serum ANGPTL6 concentration (p < 0.001). Besides, fasting serum ANGPTL6 concentration was positively associated with serum ANGPTL6 concentration at 30-minutes after glucose challenge (r = 0.5833, p < 0.0001), serum ANGPTL6 concentration at 120-minutes after glucose challenge (r = 0.05873, p < 0.0001) and area under the curve of serum ANGPTL6 ANGPTL6 is positively associated with the response of serum ANGPTL6 to hyperglycemia.

We also conducted animal and cell studies to explore the regulation and function of ANGPTL6.

Firstly, we analyze the plasma ANGPTL6 protein in mice fed a normal diet (ND) or a high-fat diet (HFD) since 8 weeks for a period of 12 weeks (n = 6) by Western blot and ELISA. Immunoblotting revealed more abundant ANGPTL6 expression in mice fed with a high-fat diet than in a normal diet. < 0.05). Secondly, we validated that if ANGPTL6 could protect from advancer glycated end-products (AGE)-induced hepatocyte death. The HepG2 cells were pre-treated with ANGPTL6 recombinant protein for 3 hours and then treated with 3 mM methylglyoxal for another 18 hours. The cell viability by MTT assay revealed that ANGPTL6 dose-dependently protected from AGE-induced hepatocyte death.

In conclusion, serum ANGPTL6 increased in response to hyperglycemia in humans and mice, which could be useful to maintian glucose homeostasis and to protect hepatocytes from hyperglycemiainduced injury.