Dr Neharika Malhotra Bora & Dr keshav Malhotra Editors’
his year on its own has been out of the box, with remarkable projects that were done like the embryology Consensus, the isar iFs-aCe-FoGsi Consensus, the unique Masterclasses, skill certification programs, online learning amongst others. it is therefore, befitting that the last edition of the isar express be called ‘out of the Box’. We all want to do the best for our patients and sometimes, it may need some out of the box thinking to achieve that one precious pregnancy which we all pray for. it sounds rather perverse and archaic today to call a child born by ivF a “test-tube baby”. the technique of assisted reproduction has become so widespread and normalised, with more than 8 million babies down the road, that there can be nothing as remarkable or stigmatising to have been conceived in a petri dish. in many countries worldwide, 3-6% of all children are born this way. Given how much scepticism and opposition ivF faced when it was still an untried and somewhat speculative field of research in the late 1960s, it’s surprising how quickly it became accepted. Critics of ivF should look at themselves, because pregnancy is a fundamental thing. if we are not here to reproduce what are we here for? so much so that various therapies are now available which supposedly increase the chances of getting pregnant. the emma and alice tests are two new practices that allow uterine mapping for a healthy pregnancy, thus maybe increasing pregnancy success. For successful ivF treatment, a healthy embryo as well as a healthy uterine environment are essential. Until recently, the uterine environment was thought to be completely sterile. But studies have
T
shown the presence of some useful and some harmful germs inside the uterus. the useful germs, called lactobacilli, help the pregnancy to continue in a healthy way. the eMMa test can detect these and other useful germs and provide the most accurate time for embryo transfer. New diagnostic tests are also in the offering which could help in giving that necessary closure to the patient when they ask that dreadful question of why they didn’t conceive. stanford researchers announced last month that they have developed a technique to determine whether an embryo should be implanted in an in vitro fertilization procedure. the trick: Check how squishy it is. the squishiness predicts how well the embryos will undergo cell division and, in theory, how likely they are to thrive. a human trial is underway- but is telling the patients that your embryos aren’t squishy enough advisable? We don’t think so. the rapid evolution of certain technologies specially in the field of genetics is something that we ought to look forward to. PGt-a is used to identify viable embryos, so the transfer or storage of embryos with an incorrect number of chromosomes can be avoided, as those typically lead to failed ivF cycles. traditionally, PGt-a requires a biopsy of a developing embryo by creating an opening in the outer coating prior to removal and testing of a few cells. however, recent studies have shown that an embryo releases small amounts of DNa into the culture media in which it is growing, allowing the surrounding fluid to be genetically tested instead. so while the efficacy of PGt-a is still heavily debated in the ivF community we already have an alternative for it and only time will tell how it gets incorporated in the indian ivF scenario, specially when, maybe, the indian government wants us to do PGt for all??? huntington’s disease, a genetic disorder that ravages the brain and nervous system and invariably ends in an early death, with a 50% chance of passing on the same grim fate to their children, PGt is something which can be a big help. But what if its just edited out of all the
EDITORIAL embryos, the prospect of using CrisPr to alter the gene in human embryos is fascinating but still worrisome. the first known attempt at heritable gene editing in humans was an effort to disable a gene called CCr5, which produces an immunecell receptor that allows hiv to infect humans. Break the gene, and the children should be resistant to the virus, reasoned he Jiankui, then at the southern University of science and technology in shenzhen, China. he attempted to create a CCr5 mutation that is found naturally in some people of european descent and is associated with hiv resistance. But a study published this month using data from the Uk Biobank found that the deletion might also shorten lifespan and now the scientist sits in prison. the long term consequences of technology is something that should not be taken lightly because, as doctors our first aim is always Primum non nocere First do no harm, and harm today, can be classified in so many more ways that we can even imagine. therefore, before offering any therapy the robustness of the evidence is something we must all look at, and always whenever trying anything controversial, discussing the facts with the patient followed by documented consents are life savers. With these thoughts in mind the edition was conceived and we quickly realised that there was so much to cover and it was challenging for us to contain ourselves to the limits of this magazine. it was also a fun challenge to interview one of the pioneers in the field who also happens to be our Dad, thanks to this we got a fun outing together which again is something that all of us should do more often. its also valentine month and our fantastic isar foundation day happens to be in this month as well. our love birds highlight their mantras for long lasting fruitful relationships. thanks to team isar for giving us this opportunity, and with that we hope that you have a fantastic time reading this edition. Neharika & Keshav
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ISAR Express February 2020
Contents PRESIDENT’S MESSAGE SECRETARY GENERAL’S MESSAGE SHORT TAKES ENDOMETRIAL MICROBIOME METAGENOMIC ANALYSIS INTRALIPID INFUSION DURING IVF NON-INVASIVE CHROMOSOMAL SCREENING OF EMBRYOS INVOCELL: INTRAVAGINAL CULTURE LYMPHOCYTE IMMUNIZATION THERAPY FOR RECURRENT MISCARRIAGE EDITING THE GENOME INTERVIEW: DR NARENDRA MALHOTRA ISAR’S VALENTINE COUPLES BEAUTIFUL INSIDE OUT THE BOND OF TRUST VAASTU TIPS FOR A STRESS FREE HOME EMBRYOLOGY ISAR 2019 MASTER CLASS ON MALE INFERTILITY STATE CHAPTER REPORTS
PAGE 6 PAGE 8 PAGE 12 PAGE 18 PAGE 20 PAGE 22 PAGE 26 PAGE 29 PAGE 32 PAGE 34 PAGE 38 PAGE 40 PAGE 42 PAGE 44 PAGE 46 PAGE 50 PAGE 51
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ISAR Express February 2020
FROM THE PRESIDENT’S DESK
“If you want something new, you have to stop doing something old.” –Peter F. Drucker
Dear Friends, loads of love and greetings. the year ran past us and we are now on our last issue of isar express. it has been an exhilarating experience, interacting with all of you, and i feel that together we have achieved all what we had set out to do. i am happy and sad at the same time. happy for this unique opportunity which you all gave me and helped me to be creative and think out of the box on how to work “towards better practices, better outcomes”. and sad because this one year of achievements has passed in a whirl and finally comes to a close. i do believe in Not thinking out of the box but thinking as if there is No box. “Creativity involves breaking out of the established patterns in order to look at things in a different way” edward de Bono. We tried to do exactly that. We focussed on different ways to reach out to the people. apart from public forums, we made a short film “Main Bhanj Nahin Hun” for creating public awareness about infertility and highlighted that reaching out at the right time to the right place for seeking treatment for subfertility will result in the best outcomes. We also made a film on “How to choose the Right IVF Clinic”. as our Prime Minister said “I draw pleasure in governance, in doing new things and bringing people together. That pleasure is all I need from life”. We brought all organisations together - isar, iFs, FoGsi and aCe, to bring out two consensus ; one for Clinicians and one for embryologists. it has been a marathon task, but together we have make it possible. “Keep away from people who try to belittle your ambitions, small people always do that, but the really great make you feel that you too can become great.” - Mark twain. i want to put on record my sincere thanks to all the great people from all our organisations, who had faith in me and positively motivated all of us to achieve whatever we could do. “Leadership means bringing people together in pursuit of a common cause, developing a plan to achieve it, and staying with it until the goal is achieved.” Bill Clinton our young turks, our siGs, our committees, all put in their best foot forward and have come out with many publications, educational pamphlets, CMes, workshops, courses (Fertility Nurses training), webinars and online learning programs together and i am sure these will have a huge impact in the coming years. this issue of isar eXPress is all about ideas which challenge conventional thinking, and i must thank both Neharika and keshav for their untiring efforts to make not only this issue an interesting one but also facilitating me to take on this humongous task. thank you from the bottom of my ‘Box’.
Dr Jaideep Malhotra President ISAR
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ISAR Express February 2020
Dear Friends, it gives me great pleasure to write the secretary’s message for the last newsletter of my term, isar express 4. We have had a series of educative and informative newsletters this year and also the “out of the Box” theme of this newsletter aptly concludes the year long activities which the President Dr Jaideep Malhotra had initiated. With all round progress happening in the field of infertility treatment, few “out of the Box” innovative ideas are always welcome to maintain the trend of progressive thoughts and solutions. i am sure this newsletter will go a long way in informing us of such strategies which ultimately benefit our patients. i sincerely thank the editors for all their efforts towards the newsletter, and am sure, everyone will enjoy contents and become richer in ideas. sure, to see you in large numbers at the silver Jubilee conference of isar at hyderabad between March 6-8, 2020. Dr S Krishnakumar Hony Secretary General, ISAR
SECRETARY GENERAL’S MESSAGE
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ISAR Express February 2020
EXECUTIVE COMMITTEE ISAR 2019-20
Dr. S. Krishnakumar Hon. Secretary General
Dr Jaideep Malhotra President
Dr Prakash Trivedi President Elect
Dr Sujata Kar Hon. Joint Secretary
Dr. Kedar Ganla Hon. Treasurer
Dr Nandita Palshetkar Vice President
Dr. Asha Baxi Hon. Joint Treasurer
Dr. Ameet Patki Second Vice President
Dr. R B Agrawal Vice Chairman for Embryology
Mr. Sudesh Kamat Chairman for Embryology
Dr. Rishma D Pai Immediate Past President
Dr A. Suresh Kumar Dr Poonam Loomba Hon. Clinical Secretary Hon. Librarian
Dr Madhuri Patil Editor: JHRS
PAST PRESIDENTS
Dr Duru Shah
Dr Narendra Malhotra
Dr HD Pai
Dr Manish Banker
Dr Dhiraj Gada
Dr Sadhna Desai
Dr Kamini Rao Dr Firuza Parikh
Late Dr Mehroo Hansotia
Dr Mahendra N Parikh Founder President
ISAR Express February 2020 MEMBERS
Dr. Ashish Kale
Dr. Charudutt Dr. Dharmesh Kapadia Joshi
Dr. Mangla Kawade
CO-OPTED MEMBERS
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Dr. Seema Pandey
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Dr. Gautam Khastgir
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Dr. Rajesh Modi
Dr. Kanthi Bansal
Dr. Priya Dr. Prakash Shivdas Patil Kannan
ISAR STATE CHAPTERS ISAR StAte cHAPteRS
cHAIRPeRSon
Hon. Gen. SecRetARy
ChhatisGarh
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GUJarat
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haryaNa
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karNataka
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orissa
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raJasthaN
Dr. leela vyas
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taMilNaDU &
Dr. kaMala selvaraJ
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aNDhra PraDesh
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ChaNDiGarh
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ISAR Express February 2020 SHORT TAKES
Scientists reverse reproductive clock in mice Researchers have lifted fertility rates in older female mice with small doses of a metabolic compound that reverses the aging process in eggs, offering hope for some women struggling to conceive.
https://ivf.net/ivf/the-aftermath-of-the-hejiankui-ďŹ asco-china-s-res ponse-o11310.html
The University of Queensland study found a noninvasive treatment could maintain or restore the quality and number of eggs and alleviate the biggest barrier to pregnancy for older women. A team led by UQ's Professor Hayden Homer found the loss of egg quality through aging was due to lower levels of a particular molecule in cells critical for generating energy. "Quality eggs are essential for pregnancy success because they provide virtually all the building blocks required by an embryo," Professor Homer said. "We investigated whether the reproductive aging process could be reversed by an oral dose of a 'precursor' compound -- used by cells to create the molecule." The molecule in question is known as NAD (nicotinamide adenine dinucleotide) and the 'precursor' as NMN (nicotinamide mononucleotide). Professor Homer said fertility in mice starts to decline from around one year of age due to defects in egg quality similar to changes observed in human eggs from older women. "We treated the mice with low doses of NMN in their drinking water over four weeks, and we were able to dramatically restore egg quality and increase live births during a breeding trial," Professor Homer said. Professor Homer said poor egg quality had become the single biggest challenge facing human fertility in developed countries. "This is an increasing issue as more women are embarking on pregnancy later in life, and one in four Australian women who undergo IVF treatment are aged 40 or older," he said. "IVF cannot improve egg quality, so the only alternative for older women at present is to use
eggs donated by younger women. "Our findings suggest there is an opportunity to restore egg quality and in turn female reproductive function using oral administration of NADboosting agents -- which would be far less invasive than IVF. It is important to stress, however, that although promising, the potential benefits of these agents remains to be tested in clinical trials." This study was conducted in collaboration with UNSW and published in the journal Cell Reports. Story Source: Materials provided by University of Queensland. www.sciencedaily.com Journal Reference: Michael J. Bertoldo, Dave R. Listijono, Wing-Hong Jonathan Ho, Angelique H. Riepsamen, Dale M. Goss, Dulama Richani, Xing L. Jin, Saabah Mahbub, Jared M. Campbell, Abbas Habibalahi, WeiGuo Nicholas Loh, Neil A. Youngson, Jayanthi Maniam, Ashley S.A. Wong, Kaisa Selesniemi, Sonia Bustamante, Catherine Li, Yiqing Zhao, Maria B. Marinova, Lynn-Jee Kim, Laurin Lau, Rachael M. Wu, A. Stefanie Mikolaizak, Toshiyuki Araki, David G. Le Couteur, Nigel Turner, Margaret J. Morris, Kirsty A. Walters, Ewa Goldys, Christopher O’Neill, Robert B. Gilchrist, David A. Sinclair, Hayden A. Homer, Lindsay E. Wu. NAD Repletion Rescues Female Fertility during Reproductive Aging. Cell Reports, 2020; 30 (6): 1670 DOI: 10.1016/j.celrep.2020.01.058
SHORT TAKES ISAR Express February 2020
DIET HAS RAPID EFFECTS ON SPERM QUALITY Sperm are influenced by diet, and the effects arise rapidly. This is the conclusion of a study by researchers at Linköping University, in which healthy young men were fed a diet rich in sugar. The study, which has been published in PLOS Biology, gives new insight into the function of sperm, and may in the long term contribute to new diagnostic methods to measure sperm quality. "We see that diet influences the motility of the sperm, and we can link the changes to specific molecules in them. Our study has revealed rapid effects that are noticeable after one to two weeks," says Anita Öst, senior lecturer in the Department of Clinical and Experimental Medicine at Linköping University, and head of the study. Sperm quality can be harmed by several environmental and lifestyle factors, of which obesity and related diseases, such as type 2 diabetes, are well-known risk factors for poor sperm quality. The research group that carried out the new study is interested in epigenetic phenomena, which involve physical properties or levels of gene expression changing, even when the genetic material, the DNA sequence, is not changed. In certain cases such epigenetic changes can lead to properties being transferred from a parent to offspring via the sperm or the egg. In a previous study, the scientists showed that male fruit flies which had consumed excess sugar shortly before mating more often produced offspring who became overweight. Similar studies on mice have suggested that small fragments of RNA known as tsRNA play a role in these epigenetic phenomena that appear in the next generation. These RNA fragments are present in unusually large amounts in the sperm of many species, including humans, fruit flies and mice. So far, their function has not been examined in detail. Scientists have speculated that the RNA fragments in sperm may be involved in epigenetic phenomena, but it is too early to say whether this is the case in humans. The new study was initiated by the researchers to investigate whether a high consumption of sugar affects the RNA fragments in human sperm. The study examined 15 normal, non-smoking young men, who followed a diet in which they were given all food from the scientists for two weeks. The diet was based on the Nordic Nutrition Recommendations for healthy eating with one exception: during the second week the researchers added sugar, corresponding to around 3.5 litres of fizzy drinks, or 450 grammes of confectionery, every day. The sperm quality and other indicators of the participants' health were investigated at the start of the study, after the first week (during which they ate a healthy diet), and after the second week (when the participants had additionally consumed large amounts of sugar). At the beginning of the study, one third of the participants
had low sperm motility. Motility is one of several factors that influence sperm quality, and the fraction of people with low sperm motility in the study corresponded to that in the general population. The researchers were surprised to discover that the sperm motility of all participants became normal during the study. The researchers also found that the small RNA fragments, which are linked to sperm motility, also changed. They are now planning to continue the work and investigate whether there is a link between male fertility and the RNA fragments in sperm. They will also determine whether the RNA code can be used for new diagnostic methods to measure sperm quality during in vitro fertilisation. Story Source: Materials provided by Linköping University. www.sciencedaily.com Journal Reference: Daniel Nätt, Unn Kugelberg, Eduard Casas, Elizabeth Nedstrand, Stefan Zalavary, Pontus Henriksson, Carola Nijm, Julia Jäderquist, Johanna Sandborg, Eva Flinke, Rashmi Ramesh, Lovisa Örkenby, Filip Appelkvist, Thomas Lingg, Nicola Guzzi, Cristian Bellodi, Marie Löf, Tanya Vavouri, Anita Öst. Human sperm displays rapid responses to diet. PLOS Biology, 2019; 17 (12): e3000559 DOI: 10.1371/journal.pbio.3000559
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ISAR Express February 2020 SHORT TAKES
Scientists Discover Stem Cells That Can Promote Endometriosis, Endometrial Cancer you cause mutations in these cells, you get endometrial cancer,” Pradeep tanwar, PhD, lead researcher and senior author of the study, said in a news story. “What we now hypothesise is that when women have endometriosis, what they have is an expansion of these mutated cells. these cells end up going into the abdominal cavity. Because they are so highly regenerative — because these are the cells that are repairing the uterus in each cycle — they start making uterine-like tissues in the abdominal cavity — which is what endometriosis is,” said tanwar, also an associate professor at the hunter Medical research institute. tanwar also believes the malstem cells responsible for repairing the womb (endometrium) are responsible function of these stem cells could be womb following menstruation can also for replenishing the endometrial tissue the reason why women undergoing ferpromote endometriosis and endomethat is lost during menstruation. tility treatments fail to conceive. trial cancer if they become dysfuncIn vivo lineage tracing is a technique the hunter researchers spent seven tional, researchers have found. that allows scientists to label and folyears “exhaustively testing” their findsuch malfunctioning stem cells also can low specific cells inside an organism. in ings, tanwar said. he said they colreduce the chances of success for so doing, the team discovered these lected and banked gynecological tissue women undergoing in vitro fertilization stem cells contained high levels of a samples from hundreds of women (ivF), the study shows. gene called axin 2, which has been treated at the center. the researchers’ findings were refound to be active in cell types from Given the wide implications of these ported in “Endometrial Axin2+ Cells other highly regenerative tissues. findings, many scientists around the Drive Epithelial Homeostasis, Regenera- When the researchers specifically deglobe have reached out to congratulate tion, and Cancer following Oncogenic stroyed axin 2-positive stem cells in the the team on their discovery, he said. Transformation,” a study published in wombs of female mice, they found the “there have been so many questions the journal Cell Stem Cell. endometrium was no longer able to reabout these conditions, and this has Using a technique called in vivo lineage pair itself and became highly dysfuncgiven us a framework to start addresstracing in female mice, investigators tional. Moreover, when investigators ing those, and — hopefully — come up from the hunter Medical research inintroduced cancer-associated mutawith some answers,” tanwar said. stitute in australia and their colleagues tions into these stem cells, the cells discovered that stem cells located at started to malfunction and to fuel the the base of special glands found development of endometrial cancer. throughout the inner lining of the “What we are able to show is that if
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ISAR Express February 2020 SHORT TAKES
Chlamydia in testicular tissue linked to male infertility The potential impact of undiagnosed sexually transmitted chlamydia infection on men's fertility has been highlighted in a study led by Queensland University of Technology (QUT), which for the first time found chlamydia in the testicular tissue biopsies of infertile men whose infertility had no identified cause. The researchers also found antibodies specific to the bacteria responsible, Chlamydia trachomatis, in the blood of 12 of 18 donors of the fresh testicular biopsies, indicating the men had been exposed to the bacteria -- yet none of the men reported symptoms of infection or being previously diagnosed with chlamydia or any other sexually-transmitted infection (STI). The study, in collaboration with Monash IVF Group, Hudson Institute of Medical Research, Monash Health, and Queensland Fertility Group, has been published in the journal Human Reproduction. Key findings: • Men whose tissue was tested were moderately to severely infertile, producing no or little sperm, and the majority had no defined cause of their infertility. • Chlamydia was found in 45.3 per cent of fixed testicular biopsies (43 of 95 men), obtained from the Monash Health Anatomical Pathology Department. All men in this group had no defined cause of infertility. • Chlamydia was also found in 16.7 per cent of fresh testicular biopsies (3 of 18 men), obtained during patient sperm recovery procedures by the Monash IVF Group and Queensland Fertility Group. These 3 men, and another 10 in the group, had no identified cause for their infertility. • In 12 of the 18 men providing the fresh biopsies (66.7 per cent) Chlamydia trachomatis-specific antibodies were found in serum, indicating the men had been exposed to the bacteria -- but all were asymptomatic and said they'd not been diagnosed with any STI. • Research leader QUT Professor of Immunology Ken Beagley, from the Insti-
tute of Health and Biomedical Innovation, said chlamydia infection in men has not been as widely studied as it has in women, despite similar infection rates. "Chlamydia infection has been associated with women's infertility but much less is known about its impact on male infertility, particularly if men do not experience symptoms, which is estimated to be in about 50 per cent of cases," he said. "When people have no symptoms they can unknowingly pass on the infection to sexual partners. "This is the first reported evidence of chlamydia infection in human testicular tissue, and while it can't be said that chlamydia was the cause of the infertility of the men, it is a significant finding. "We believe future studies with male patients should look at how chlamydia infection might cause damage to the male reproductive system and contribute to infertility." Professor Beagley said testing testicular tissue could also be a useful future screening and diagnostic tool for clinicians and help inform them about treatments to improve reproductive outcomes. Story Source: www.sciencedaily.com Materials provided by Queensland University of Technology. Journal Reference: Emily R Bryan, Robert I McLachlan, Luk Rombauts, Darren J Katz, Anusch Yazdani, Kristofor Bogoevski, Crystal Chang, Michelle L Giles, Alison J Carey, Charles W Armitage, Logan K Trim, Eileen A McLaughlin, Kenneth W Beagley. Detection of chlamydia infection within human testicular biopsies. Human Reproduction, 2019; DOI: 10.1093/humrep/dez169
Choosing the best Embryos Healthy cells usually have one nucleus, where DNA containing our genetic information is stored. Embryos that are created in vitro in fertility clinics to enable women to have a child, often have cells with two nuclei. As of today, many fertility clinics still transfer these socalled "binucleated embryos" back to the patient's uterus. "In our study, we showed in mouse embryos that binucleation has profound consequences. Basically, having two nuclei is bad news for the embryo. We found that binucleation increases the chances of the embryo developing a condition called aneuploidy, which reduces embryo health and could contribute to pregnancy failures," explained Lia Paim, first author and PhD student in Dr. Greg FitzHarris lab. "We hope our results will help fertility clinics to select the best embryos to be transferred back to the patients. This step is one of the keys to success when it comes to in vitro fertilization. Ultimately it could increase some couples' chances of giving birth," said Mrs. Paim. This study is at the basic research stage and was carried out in the laboratory on mice. "Basic science experiments such as Lia's allow us to understand how embryos develop, to help inform our clinical colleagues how to select the best embryos in the clinic," added Dr. Fitzharris, CRCHUM researcher and professor at the Université de Montréal. Story Source: Materials provided by University of Montreal Hospital Research Centre (CRCHUM). www.sciencedaily.com Journal Reference: Lia Mara Gomes Paim, Greg FitzHarris. Tetraploidy causes chromosomal instability in acentriolar mouse embryos. Nature Communications, 2019; 10 (1) DOI: 10.1038/s41467-019-12772-8
Heartiest Congratulations to Dr Prakash Trivedi for Taking Over as President ISAR 2020-2021
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ISAR Express February 2020 CLINICAL ART
ENDOMETRIAL MICROBIOME METAGENOMIC ANALYSIS DR VANIE SARDA THAPAR MBBS, MD - Obstetrics & Gynaecology
DR VIDHU MODGIL MBBS, MD - Obstetrics & Gynaecology
t
he term human microbiome represents the totality of microorganisms and their collective genetic material present in or on the human body. it is quite clear that the vaginal and uterine microbiomes play a role in the physiology and pathophysiology of human reproduction. the normal vaginal microbiome in healthy women is generally dominated by lactobacilli species, in an effort to categorize the vaginal flora, it has been further classified into five community state types (Csts). More than 70% of women demonstrated vaginal microbiota dominated by l. crispatus, l. gasseri, l. iners, or l. jensenii, corresponding to Cst-i, -ii, -iii, and -v. a smaller proportion of women exhibit Cst-iv, characterized by lower percentage of lactobacilli and dominance of anaerobic bacteria including aerococcus, atopobium, Dialister, Gardnerella, Megasphaera, Prevotella, and sneathia.1 Uterine microbiota may also contribute to healthy endometrium physiology. Upon pattern recognition receptor stimulation by these bacteria, epithelial cells release soluble factors, such as cytokines, affecting local lymphocyte populations e.g. antigen presenting cells, thereby initiating a signaling cascade and altering the mucosal t cell balance. Cells important for healthy implantation and placentation, such as uNk cells, are potentially affected.2 aPC, antigen presenting cell; il-10, interleukin 10; Psa, polysaccharide a; Prr, pattern recognition receptor; th17, t helper 17 cells; treg, regulatory t cell. since a successful embryo implantation requires both a synchronous development and an intricate interplay between the hatched blastocyst and endometrium. this depends on a receptive state of the endometrium. During the female cycle, if the endometrium fails to undergo the proper adaptations to acquire this receptive state, infertility and impaired placentation may be the consequence. since bacteria can play a role in morphological changes of endometrium, the maternal microbial colonization contributes to development of the unborn child by micronutrient provision, xenobiotic metabolism and enhancing maternal energy conversion.3 the most prominent example of a pathology caused by an altered endometrial microbiota is chronic endometritis. it is characterized
by the continuing inflammation of the endometrial mucosa produced by the colonization of the uterus by common bacteria, namely, enterococcus faecalis, escherichia coli, Gardnerella vaginalis, klebsiella pneumoniae, Proteus spp., Pseudomonas aeruginosa, staphylococcus spp., streptococcus spp., genital pathogens as Mycoplasma and Ureaplasma spp., and yeasts like saccharomyces cerevisiae and Candida spp.4,5 the prevalence of chronic endometritis in the general population has been estimated to be 19%, and it is estimated to be 45% in infertile patients. however, this high prevalence seems to be mainly associated to recurrent implantation failure (riF) and recurrent pregnancy loss (rPl) rather than other causes of infertility.6,7 the endometrial microbiome of infertile patients and its functional impact on reproductive outcome have been recently assessed in two different studies using sequencing technology. in the first study, 33 patients of different ethnicities (26 white, five asian, one african-american, and one hispanic) had the endometrial microbiome characterized at the time of embryo transfer of a single euploid embryo. analysis showed the microbiome was composed of 278 genera, with Flavobacterium and lactobacillus being the most abundant genera in patients with ongoing and non-ongoing pregnancies. the most recent study analyzed the impact of endometrial microbiome on reproductive outcome in endometrial fluid from 35 infertile white patients with recurrent implantation failure. the endometrial microbiota was made of 108 components, with lactobacillus spp. being the most abundant bacteria detected. the results of this study show that the endometrial microbiota profile can be classified as lactobacillus dominated or non–lactobacillus dominated with a cut off value of lactobacillus relative abundance r90%; this cut off served as a significant variable able to predict reproductive success. thus, a non– lactobacillus dominated (<90%) endometrial microbiota significantly correlates with adverse reproductive outcomes—measured as implantation, pregnancy, ongoing pregnancy, and miscarriage rates—when compared with subjects presenting a lactobacillus-dominated (r90%) endometrial microbiota.8 also, Moreno et al. investi-
CLINICAL ART ISAR Express February 2020 gation of the presence of a uterine microbiome. Negative association of non- lactobacillus dominated subjects with pregnancy outcome (decreased implantation of 23.1% versus 60.7%; pregnancy rates 33.3% versus 70.6%; ongoing pregnancy rates 13.3% versus 58.8%, and live birth rates 6.7% versus 58.8%). an especially negative impact on reproductive outcome was observed when G. vaginalis and streptococcus species were present in abundance. these results were independent of ph of the sample, known to be affected by lactobacillus species.9 With culture-based approaches, it has been shown that certain bacteria, such as enterobacteriaceae and staphylococcus, found at the time of embryo transfer on the transfer catheter were associated with poorer outcomes.10 the vaginal microbiome as it pertains to art has been analyzed with both cultivative and sequencing approaches. in culture-based approaches, rapidly growing, aerobic species dominate, leaving rare species that demand specific culture conditions undetected.11 Molecular approaches allow detection of species that will not be revealed by culture-dependent techniques. New investigative techniques including DNa fingerprinting, microarrays, and targeted or whole genome sequencing have empowered the study of metagenomics by analyzing the bacterial communities contained in samples based on their genetic information. these techniques have revealed that sites in the body historically thoughts to be sterile, such as the uterine cavity and the placenta, are in fact colonized with their own unique microbiome.12 these molecular techniques take advantage of the 16s rrNa gene that is unique to bacteria and contains a number of hypervariable regions that serve as unique identifiers for a genus or species of bacterium. Metagenomic sample sequencing data is processed and organized into clusters termed operational taxonomic units (otUs) by mapping the 16s sequence to publicly available taxonomic databases. otUs are then used to determine sample composition and diversity. this read count clustering, also known as ‘‘binning,’’ can only be performed when known sequences exist; it becomes much more challenging and less accurate when analyzing novel species.13 Further limitations of microbiome sequencing are related to the clinical utility of the results as it does not give information about its biologic function, such as antibiotic susceptibility testing. We conclude that the concept of the sterile endometrium, and the uterine compartment in general, is outworn, although the true core uterine microbiome still needs to be assessed. Functional studies are needed to elucidate the physiological importance of the microbiome in fertility. the challenge of studying reproductive immunology and the microbiota involved is that research on all of the different aspects is still in its infancy. Microbiome, immunity, endocrinology in preg-
nancy, and placental and fetal development need to be studied together to obtain a more comprehensive overview. References 1. Ravel J, Gajer P, Abdo Z, Schneider GM, Koenig SSK, McCulle SL, et al. Vaginal microbiome of reproductive-age women. Proc Nat Acad Sci 2011; 108(Suppl 1):4680–7. 2. Espinoza J, Erez O, Romero R. Preconceptional antibiotic treatment to prevent preterm birth in women with a previous preterm delivery. Am J Obstet Gynecol 2006;194:630–637. 3. Macpherson AJ, de Aguero MG, Ganal-Vonarburg SC. How nutrition and the maternal microbiota shape the neonatal immune system. Nat Rev Immunol 2017; 10:508–517. 4. Greenwood SM, Moran JJ. Chronic endometritis: morphologic and clinical observations. Obstet Gynecol 1981;58:176–84. 5. Cicinelli E, De Ziegler D, Nicoletti R, Tinelli R, Saliani N, Resta L, et al. Poor reliability of vaginal and endocervical cultures for evaluating microbiology of endometrial cavity in women with chronic endometritis. Gynecol Obstet Invest 2009;68:108–15. 6. Yoshii N, Hamatani T, Inagaki N, Hosaka T, Inoue O, Yamada M, et al. Successful implantation after reducing matrix metalloproteinase activity in the uterine cavity. Reprod Biol Endocrinol 2013;11:37. 7. Kushnir VA, Solouki S, Sarig-Meth T, Vega MG, Albertini DF, Darmon SK, et al. Systemic inflammation and autoimmunity in women with chronic endometritis. Am J Reprod Immunol 2016;75:672–7. 8. Moreno I, Codo~ner FM, Vilella F, Valbuena D, MartinezBlanch JF, Jimenez- Almaz_an J, et al. Evidence that the endometrial microbiota has an effect on implantation success or failure. Am J Obstet Gynecol 2016;215:684–703. 9. Tachedjian G, Aldunate M, Bradshaw CS, Cone RA. The role of lactic acid production by probiotic Lactobacillus species in vaginal health. Res Microbiol 2017;168:782–792. 10. Selman H, Mariani M, Barnocchi N, Mencacci A, Bistoni F, Arena S, et al. Examination of bacterial contamination at the time of embryo transfer, and its impact on the IVF/pregnancy outcome. J Assist Reprod Genet 2007;24:395–9. 11. Verstraelen H, Verhelst R, Claeys G, Temmerman M, Vaneechoutte M. Cultureindependent analysis of vaginal microflora: the unrecognized association of Atopobium vaginae with bacterial vaginosis. Am J Obstet Gynecol 2004;191: 1130–1132. 12. Handelsman J. Metagenomics: application of genomics to uncultured microorganisms. Microbiol Mol Biol Rev 2004;68:669–85. 13. Alneberg J, Bjarnason BS, de Bruijn I, Schirmer M, Quick J, Ijaz UZ, et al. Binning metagenomic contigs by coverage and composition. Nat Meth 2014;11:1144–6.
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INTRALIPID INFUSION DURING IVF DR SHARDA JAIN MD. ( PGIMER), MANMS, FICOG, FIMSA, DHM, QM & AHD
DR JYOTI BALI MBBS, MS (Obstetrics & Gynaecology), Fellow Reproductive Medicine
Intralipid is a 20% intravenous fat emulsion that has been used as a source of fat and calories for patients requiring parental nutrition. It consists of soya bean, Egg yolk, phospholipids, glycerine and water.
the intravenous fat emulsion intralipid has been hypothesised to be an effective and safe treatment for repeated ivF implantation failure and pregnancy loss.1 there is evidence that altered maternal immune function as a factor in recurrent implantation loss and miscarriage (sacks, 2015a) 1.1 although it is difficult to identify and define with current clinically available tests. the clinical practice of reproductive immunology was dominated by Peter Medawar’s seminal paper, describing the need for maternal immune suppression to accommodate an invading placenta (sacks, 2015b; Medawar,1953) 1.2. however, it is now clear that some elements of the maternal immune system are activated and essential for successful implantation (Moffett and Colucci, 2014). intralipid is a 20% intravenous fat emulsion that has been used as a source of fat and calories for patients
requiring parental nutrition. it consists of soya bean, egg yolk, phospholipids, glycerine and water. in a small and still unpublished non randomised trial, presented at a scientific meeting in the Uk 2. a 50% pregnancy rate and 46% clinical pregnancy rate were achieved in patients with riF who had an elevated th1 cytokine response. MecHAnISM oF ActIon alteration of th1 to th2 cytokine activity ratio which decrease in all cases appear to corelate with successful outcome that resulted. the mechanism by which intralipid modulates the immune system still unclear it has been postulated that fatty acids within the emulsions serve as ligands to activate peroxisome proliferator activated receptors expressed by the Nk cells. activation of such nuclear receptors has been shown to decrease Nk cytotoxic
CLINICAL ART ISAR Express February 2020 activity, enhancing implantation. IntRALIPID ReGIMe intralipid infusion is administered once between days 4 and 9 of ovarian stimulation, and again within 7 days of a positive pregnancy test, then every 4 weeks until 12-13 weeks of pregnancy .intralipid 20% (intralipid, Fresenius kabi, hamburg, Germany) was given as an intravenous infusion made up from 100ml of intralipid diluted in 500ml of normal saline over 3 to 4 hours in a clean, sterile inpatient hospital environment. Women at ivFa received an intralipid infusion on day 5-9 of their cycle, and again following detection of a positive Beta-hCG. Fertility sa women received intralipid infusion at oocyte retrieval in a fresh embryo transfer cycle or at embryo transfer when in a frozen embryo transfer cycle, and then again at detection of a positive Beta- hCG. a positive Beta-hCG was defined as a serum estimate >25iU. PRecAUtIonS BeFoRe USInG IntRALIPID InFUSIon to rule out the history of• severe liver disease • kidney disease • lung disease • anaemia • Blood clotting disorder
10 maternal cycle because of no clinical pregnancies achieved in those who received intralipid versus 40% clinical and a 30% live birth rate in untreated controls. this study had some limitations because they included only advanced maternal age group (40-42 years), perhaps he founds this therapy detrimental to this age group. in the recent clinical guidelines, rCoG states that there is a paucity of available data and evidence to recommend routine measurement of cytokine level in patients with recurrent implantation failure. there is no agreed protocol for time and duration of treatment. We need to have strong clinical evidence before subjecting our patients to this treatment option. it should be noted that the largest study Bombell and McGuire examining the relationship between uNk cell numbers and future pregnancy outcome reported that raised uNk cell numbers in women with recurrent miscarriage was not associated with an increased risk of miscarriage. recent green top guideline for recurrent miscarriage said testing for uNk cells should not be offered routinely in the investigation of recurrent miscarriage.
REFERENCES 1. Romy Ehrlich, M.Louise Hull, Jane Walkley, Gavin Sacks. Intralipid Immunotherapy for Repeated IVF failure, Nov 2019. 1.1 Sacks G. Enough! Stop the arguments and get on PoSSIBLe SIDe eFFectS oF IntRALIPID InFUSIon with the science of natural killer cell testing. Hum Re• headache prod.2015q;30:1526-31. • Dizziness 1.2 Sacks G. Reproductive immunology: the relevance of • Flushing laboratory research to clinical practice (and vice versa). • Nausea Hum Reprod. 2015b;30:253-5. • vomiting 2. Simon A, Laufer N. Assessment and treatment of re• sweating peated implantation failure (RIF). J Assist Reprod Genet • Drowsiness 2012; 29:1227-39. • Fever 3. Chazara O, Xiong S, Moffett A. Maternal KIR and Fetal • Persistent sore throat HLA-C: A fine balance. J Leukoc Biol 2011; 90:703-16. • Pain/swelling/redness at injection site 4. Shreeve N, Sadek K. Intralipid therapy for recurrent im• Chest pain plantation failure: New hope or false dawn? J Report Im• sudden weight gain munoi 2012; 93:38-40. • shortness of breath as far as intralipid administration is concerned, there are 5. Dakhly DM, Bayoumi YA, Sharkawy M, Gad Allah SH, conflicting evidences available regarding the efficacy and Hassan MA, Gouda HM, et al. Intralipid supplementation in women with recurrent spontaneous abortion and eleproven benefit in women with recurrent implantation failure, while some states that the activation of decidual vated levels of natural killer cells. Int J Gynaecol obstet Nk cell by MhC ligands on trophoblasts has beneficial ef- 2016; 135:324-7. 6. Check JH, Check DL. Intravenous intralipid therapy is fect on pregnancy outcome 3. however shreeve and not beneficial in having a live delivery in women aged sadek 4 found that large scale confirmatory studies are still necessary to prove the efficacy of intralipid before it 40-42 years with a previous history of miscarriage or failure to conceive despite embryo transfer undergoing in should be recommended for routine use. Dakhly et al 5 performed an rtC as well as to investigate vitro fertilization embryo transfer. ClinExp obstet Gynecol 2016; 43:14-5. the efficacy of intralipid administration in patient with recurrent implantation failure and elevated Nk cell activity and found out no increase in chemical pregnancy rates. Check and Check 6 stopped his study prematurely after
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NON-INVASIVE CHROMOSOMAL SCREENING OF EMBRYOS DR SAYALI KANDARI B.Tech (Biotechnology), M.S (Biotechnology, USA), PhD (Stem Cell Biology) Scientist & a Research scholar in the field of Fertility & Reproductive Biology, Senior Consultant Embryologist & a Faculty /Trainer.
Preimplantation genetic testing (PGt) encompasses methods that allow embryos to be tested for severe inherited conditions or for chromosome abnormalities, relevant to embryo health and viability. in order to obtain embryonic genetic material for analysis, a biopsy is required, involving the removal of one or more cells. Universally considered the gold standard technique for isolated embryo’s genetic material, the procedure in undoubtedly technically challenging and calls for substantial change in workflow and dedicated experts to support a unit for clinical adoption the recent discovery of DNa within the blastocoele fluid (BF) of blastocysts and in spent embryo culture media (sCM) has led to interest in the development of non-invasive methods of PGt (niPGt). embryonic DNa is frequently detectable in BF and sCM of embryos produced during ivF treatment. initial studies have achieved some success when performing cytogenetic and molecular genetic analysis. however, in many cases, the efficiency has been restricted by technical complications associated with the low quantity and quality of the DNa. it is fair to say that PGt methods based upon the biopsy of polar bod-ies, blastomeres or trophectoderm cells have been highly successful, helping thousands of families achieve healthy pregnancies (De Ryckeet al., 2017.) reports that blastocyst biopsy poses less risk to the embryo than removal of blastomeres resulted in a dramatic uptake of the procedure, and today the vast majority of embryo biopsies for PGt are conducted using this method (McArthur et al., 2008; Scott et al., 2013a; Glujovsky et al., 2016). Pre and post biopsy preparations are another challenging aspects of the quality of biopsy performed. it is presumed and we can only surmise that the costs of PGt would likely be reduced and bottlenecks associated with logistics, training and financing of equipment could be eliminated, thus improving access to patients. Whether this is true or different challenges are faced by NiPGt practitioners is yet to be evaluated. BLAStocoeLIc FLUID the discovery of DNa suitable for amplification and genetic testing in the BF (BF-DNa) was first reported by Palini et al. (2013)During the BF sampling proce-
dure, sometimes termed ‘blastocente-sis’, an iCsi pipette pierces through the trophectoderm layer on the opposing side of the embryo from the iCM, and the fluid is carefully aspirated, leaving the embryo fully collapsed (Poli et al., 2015; Magli et al., 2016). Figure 1 Blastocentesis. Blastocoele fluid is isolated by blastocentesis. an iCsi injection pipette is used to
(a) puncture though the mural trophectoderm (B) opposite the inner cell mass. the blastocoele fluid is aspirated until the blastocyst collapses around the pipette (Hammond,et al. . “Nuclear and mitochondrial DNA in blastocoele fluid and embryo culture medium: evidence and potential clinical use.” Human reproduction 31 8 (2016): 1653-61 .) there are two major requirements for clinical application of PGt using minimally invasive sources of DNa. Firstly, it must be possible to consistently isolate and analyse the DNa, to avoid the possibility of failing to obtain a clinical diagnosis. secondly, the DNa must be shown to be representative of the developing embryo
EMBRYOLOGY ISAR Express February 2020 to ensure that the diagnosis is accurate. PGt have encountered significant difficulties in isolating and amplifying DNa retrieved using blastocentesis. this is thought to relate to the low quantity and poor quality of BF-DNa, which is believed to be degraded in nature (Palini et al., 2013)., studies attempting to evaluate the suitability of BF-DNa for PGt-M have not yet been able to demonstrate sufficient amplification rates in an adequate number of samples for clinical application to be considered. Risks of Misdiagnosis PCr amplification efficiencies following WGa are consistently lower than that reliably achieved following cellular biopsy and consequently, the risk that an embryo will remain undiagnosed following PGt-M is elevated for these BF strategies. additionally, as aDo rates are increased when DNa is degraded, there may be a greater risk that diagnoses using BF-DNa could be compromised (Piyamongkol et al., 2003; Huang et al., 2015).
the embryo secretome. studies have focused on metabolites,proteins, interleukins and micro-rNas and have considered whether measurement of certain secreted molecules may have value for the prediction of embryonic reproductive competence (katz-Jaffe et al., 2009; Capalbo et al., 2016; huang et al., 2017; sánchezribas et al., 2018).the reported PCr amplification rates achieved from sCM are inferior to those using cellular biopsy, indicating that as in the blastocoel, the gDNa in the sCM is likely of very low abundance and/or degraded and may be a product of cell death processes (Capalbo et al., 2013; Galluzzi et al., 2015).
PGt-M wItH ScM Following culture, day-1-5 media was collected from two blastocyst stage embryos and eight fragmented or arrested embryos, and the embryos underwent a biopsy. For the samples in which MalBaC-WGa (Multiple annealing and looping Based amplification Cycles, yikon Genomics) was successful (80% of sCM and 100% of embryo biopsies), PCr was used to further amplify the hemoglobin beta subunit gene (hBB) and 10 sNPs in close PGt-A BLAStocentenSIS proximity. allele dropout was found to be frequent, with Despite adopting the same blastocentesis, WGa (surePlex, illumina) and aCGh (24-sure, illumina) methodolo- multiple loci affected, and the detection of anomalous sNP alleles raised concern over genetic contamination. gies as Gianaroli, Magli and colleagues, extreme similar observations were reported by Capalbo et al. inconsistencies in concor- dance between BF-DNa and biopsy samples were observed betweenthe groups. it is (2018), amplification occurred in 89.7% of the 378 loci tested in sCM: rates inferior to that achieved using troreported degraded DNa is excluded from mosaic cleavphectoderm biopsies (100%). there was evidence to inage stage embryos, either through active or passive dicate that the accuracy of the diagnosis was mechanisms, expelling some into the blastocoel during blastulation, and consequently compromising the clinical compromised by DNa contamination of maternal origin. only 20.8% of sCM samples displayed full haplotype conutility of BF-DNa for PGt-a cordance with paired trophectoderm biopsies and aDo rates were higher for alleles of paternal origin compared Spent embryo culture medium to those of maternal origin (14.2% vs 8.2%), a potential alternative for the non-invasive genetic assessment of preimplantation embryos involves the Risk of Misdiagnosis analysis of spent embryo culture media (sCM) (Fig. 2). the past decade has seen several publications exploring in conclusion, whilst DNa in the sCM appears to be of
Figure 2 (Hammond,et al. . “Nuclear and mitochondrial DNA in blastocoele fluid and embryo culture medium: evidence and potential clinical use.” Human reproduction 31 8 (2016): 1653-61 .)
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ISAR Express February 2020 EMBRYOLOGY superior integrity and greater quantity compared with that found in the blastocoel, the amplification rates are still lower and aDo rates are higher than those observed using conventional trophectoderm biopsy.. the clinical consequences of this could be severe; potentially leading to misdiagnosis of embryos and the birth of affected offspring. PGt-A wItH ScM as with BF analysis, concordance rates between sCM and whole embryos/biopsy specimens have varied considerably between studies.mUsing MalBaC (yikon Genomics), successful WGa was achieved in 90.9% of samples, rates identical to those achieved using counterpart trophectoderm biopsies. it should be noted, however, that a 90% amplification rate for te specimens is significantly lower than observed for this sample type in most published studies. Mean DNa concentration was lower than that typically obtained from te samples (26.2 ng/μl vs 60.2 ng/μl). Unfortunately, the rates of full chromosome concordance between the samples (including mosaic and segmental aneuploidies) were relatively low: 64.5% when sCM samples were compared with 31 blastocysts; 44.0% when compared with the 25 arrested or degenerated embryos. Risk of Misdiagnosis the magnitude of the problem that maternal contamination poses for accurate PGt-a analysis using sCM was further demonstrated by vera-rodriguez et al. (2018), in one of the most comprehensive studies on the topic to date. Following the sequential culture of 60 embryos, day-3-5 sCM samples were subjected to a double WGa amplification due to the expected low DNa concentration; an initial surePlex (illumina) amplification was followed by a second round using the ion reproseq PGs kit (thermoFisher scientific). in contrast, paired trophectoderm biopsies were amplified using only the ion reproseq PGs kit. samples which were assigned a euploid female karyotype (46, XX) following trophectoderm analysis were excluded from analysis,due to the inability to differentiate from maternal contamination. of these informative paired samples, only 33.3% were concordant in their aneuploidy diagnosis. if DNa is preferentially released from aneuploid cells, perhaps as part of a process of euploidisation in mosaic embryos, there could be increased risks for false positive diagnosis of aneuploidy; the DNa ejected into the BF/sCM would be aneuploid while the cells remaining in the iCM are euploid. hopefully, an insight into the origin of extra-embryonic DNa should be obtained from an ongoing multicentre study, which will include cytogenetic analysis of iCM as well as te and sCM (ClinicalTrials.gov ID:
NCT03520933). the possibility of correction of mosaic embryos via apoptotic or other processes of cell loss/exclusion warrants further investigation in human embryos. the limited information available from published studies on extra-embryonic DNa suggests that there may indeed be a higher number of aneuploid events in BF and sCM samples in comparison with paired te biopsies (Magli et al., 2018; Tsuiko et al., 2018). if confirmed, this could call into question the premise upon which analysis of extra-embryonic DNa for PGt-a is based. GenetIc contAMInAtIon Concerningly, Vera-Ro-driguez et al. (2018) estimated the median percentage of embryonic DNa in sCM to be only 8%, with the rest presumably being non embryonic contaminants. Contamination may originate from five principal sources: (i) it may enter the medium (or individual media components) during the manufacturing processes; (ii) it could be maternally derived from cumulus cells or polar bodies (iii) it could be paternal, originating from sperm cells; (iv) it could arise from exogenous sources such as from laboratory personnel; or (v) it could result from microbial contamination of culture dishes during ivF (although unlikely to cause a misdiagnosis, the microbial DNa can compete with human genetic material during WGa, leading to poorer amplification (Kastrop et al., 2007) low levels of contaminating DNa were recently reported in commercial media commonly used in niPGt studies (Hammond et al., 2017). Further analysis confirmed supplementary human serum albumin (hsa) as a predominant source of contamination. hsa is frequently added to media as it contains a variety of components thought to be beneficial for embryo viability and development (Meintjes et al., 2009; Otsuki et al., 2013). PRotocoL oPtIMISAtIon & StAnDARDISAtIon a further requirement if niPGt is to be used clinically involves the optimisation and standardisation of culture conditions and medium retrieval protocols. initial studies indicate that DNa accumulates in the medium throughout preimplantation development and that the highest rates of amplification and diagnostic accuracy for sCM samples are achieved following extended embryo culture to day 6/7 (Galluzzi et al., 2015; Wu et al., 2015; Hammond et al., 2017; Ho et al., 2018; Rubio et al., 2019; Yeung et al., 2019). a prolonged culture period shouldvbe validated and any potential risks need to be characterised prior to clinical application.
EMBRYOLOGY ISAR Express February 2020 concLUSIon the recent discovery of DNa in the BF and sCM has generated a surge of interest in the potential for minimallyinvasive PGt. Possible benefit cited is avoiding expenses in the gold standard trophectoderm biopsy technique and invasive procedure. however, the reliability of niPGt strategies, with respect to the proportion of samples yielding data and the concordance of genetic results relative to those obtained from whole embryos or biopsy specimens, has varied widely between published studies. the DNa found in BF and sCM is of relatively low abundance and poor integrity, presenting technical challenges for genetic analysis. it is not clear at present which laboratory methods are the most appropriate for the investigation of extra-embryonic DNa.bUncertainty over the optimal method for niPGt and questions concerning the reliability and clinical utility of the data produced suggest that PGt based upon blastocentesis or sCM samples should, at present, only be carried out in the context of pre-clinical studies and carefully designed clinica pilot investigations. Crucially, for accurate data interpretation, and to avoid clinical misdiagnosis, the origin of extra-embryonic DNa must be confirmed and the causes underlying discordance with results rom embryo biopsy specimens should be investigated. Further studies should be encouraged, and we follow this novel field with fascination for progress. References • Cell free DNA and trophectoderm biopsy strategy. J Reprod Infertil 2019;20:57–62. Bodri D, Sugimoto T, Yao Serna J, Kawachiya S, Kato R, Matsumoto T. • Blastocyst collapse is not an independent predictor of reduced live birth: a time-lapse study. Fertil Steril 2016;105:1476–1483.Bolton H, Graham SJL, van der Aa N, Kumar P, Theunis K, Gallardo EF, Voet T, ZernickaGoetz M. • Mouse model of chromosome mosaicism reveals lineage-specific depletion of aneuploid cells and normal developmental potential. Obstet Gynecol Surv 2016;71:665–666. • Brison DR. Apoptosis in mammalian preimplantation embryos: regulation by survival factors. Hum Fertil (Camb) 2000;3:36–47. • Cao Y, Xu J, Zhang Z, Huang X, Zhang A, Wang J, Zheng Q, Fu L, Du J. Association study between methylenetetrahydrofolate reductase polymorphisms and unexplained recurrent pregnancy loss: a meta analysis. Gene 2013;514:105–111. • Capalbo A, Bono S, Spizzichino L, Biricik A, Baldi M, Colamaria S, Ubaldi FM, Rienzi L, Fiorentino F. Sequential comprehensive chromosome analysis on polar bodies, blastomeres and trophoblast: insights into female meiotic errors and chromosomal segregation in the preim-
plantation window of embryo development. Hum Reprod 2013;28:509–518. • Capalbo A, Romanelli V, Patassini C, Poli M, Girardi L, Giancani A, Stoppa M, Cimadomo D, Ubaldi FM, Rienzi L. Diagnostic efficacy of blastocoel fluid and spent media as sources of DNA for preimplantation genetic testing in standard clinical conditions. Fertil Steril 2018;110:870– 879. • Capalbo A, Ubaldi FM, Cimadomo D, Noli L, Khalaf Y, Farcomeni A, Ilic D, Rienzi L. MicroRNAs in spent blastocyst culture medium are derived from trophectoderm cells and can be explored for human embryo reproductive competence assessment. Fertil Steril 2016;105:225– 235e3. • Chang LJ, Huang CC, Tsai YY, Hung CC, Fang MY, Lin YC, Su YN, Chen SU, Yang YS. Blastocyst biopsy and vitrification are effective for preimplantation genetic diagnosis of monogenic diseases. Hum Reprod 2013;28:1435– 1444. • Chi HJ, Koo JJ, Choi SY, Jeong HJ, Roh SI. Fragmentation of embryos is associated with both necrosis and apoptosis. Fertil Steril 2011;96:187–192. • Cohen J, Elsner C, Kort H, Malter H, Massey J, Mayer MP, Wiemer K. Impairment of the hatching process following IVF in the human and improvement of implantation by assisting hatching using micro-manipulation. Hum Reprod 1990;5:7–13. • Cohen J, Wells D, Munné S. Removal of 2 cells from cleavage stage embryos is likely to reduce the efficacy of chromosomal tests that are used to enhance implantation rates. Fertil Steril 2007;87: 496–503. • De Hertogh R, Vanderheyden I, Pampfer S, Robin D, Dufrasne E, Delcourt J. Stimulatory and inhibitory effects of glucose and insulin on rat blastocyst development in vitro. Diabetes 1991;40:641–647. • De Rycke M, Goossens V, Kokkali G, Meijer-Hoogeveen M, Coonen E, Moutou C. ESHRE PGD Consortium data collection XIV– XV: cycles from January 2011 to December 2012 with pregnancy follow-up to October 2013. Hum Reprod 2017;32:1974–1
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INVOCELL INTRAVAGINAL CULTURE DR ANUPAM GUPTA MBBS,MS DIRECTOR & CONSULTANT REPRODUCTIVE MEDICINE, AKANSHA TESTTUBE BABY CENTRE, AGRA
DR DEBASHISH SARKAR MS, FICMCH, FIAOG, DIP End(Germany), FRM (Singapore) DIRECTOR & CONSULTANT REPRODUCTIVE MEDICINE, MORPHEUS UMA INTERNATIONAL IVF CENTRE, AGRA
IntRoDUctIon in vitro fertilization (ivF) is now an established treatment for infertility. ivF and variations such as intra cytoplasmic sperm injection (iCsi) requires complex laboratory equipment and very experienced laboratory personnel. Consequently, these treatments are expensive and are only available for a small fraction of the infertile population. the iNvo technique, ivC (ranoux et al., 1988), is a simple alternative to ivF, where the vaginal cavity of the patient substitutes the complex ivF laboratory. this study will describe a new device, the iNvocell, specially designed for the vaginal incubation. the iNvocell prevents the technical problems that were described during the use of a prototype. iNvo could transform the treatment of infertility. InVo PRoceDURe the iNvo procedure is a major simplification for fertilization and embryo development. the principle is simple. the vagina of the future mother replaces the complex laboratory as the site of incubation. i. the procedure: the sperm preparation is generally
performed 1 h prior to the oocyte retrieval to allow the biologist to perform the insemination immediately after oocyte retrieval. Gradients of density are used to wash the sperm and select the most motile spermatozoa. the device should be filled with medium without interposition of air. air bubbles could be trapped in the cumulus of the mature oocytes and bring them to the surface and therefore prevent fertilization by spermatozoa. a small fraction of the motile spermatozoa (30 000) is used to inseminate the oocytes in the device. after follicle aspiration, oocyte(s) are identified in the follicular fluid. as no pre-incubation of the oocytes is necessary, oocytes are immediately placed into the device, the fraction of motile sperm is added before or after transfer of the oocytes. the device is closed, placed into a protective outer rigid shell and then positioned into the vaginal cavity for 2 or 3 days. a retention system is used to secure the device in the vagina during incubation. No activity restriction is required for the patient, but baths are not recommended as they may modify the temperature of incubation. after incubation the retention system and the device are re-
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moved from the vagina in the physician's office. the outer rigid shell is removed, the device is opened and the contents observed under microscope to find the embryos. the two best ones are loaded into a catheter and transferred immediately into the uterine cavity. Ultrasound guidance may be used in order to improve the quality of the transfer. ii. the Device: a. Prototype. a prototype was used during the first development and in the publications concerning the iNvo procedure. the prototype was chosen because it showed the best sealed closure among the prototypes that were initially tested. to avoid air bubbles the corpus and the plug of the prototype tube were filled with culture medium, then the gametes were introduced and the tube was closed. several disadvantages were described such as: 1. only one sterile opening and closing were possible. 2. During the closing of the prototype oocytes could be lost in the overflow of medium and the culture medium could be easily contaminated by errors in manipulation. 3. a large opening of the tube could rapidly modify the ph of
the medium. 4. a plastic envelope was sealed thermally around the prototype to protect the contents of the tube from the vaginal secretions. During the sealing of the envelope with heat, the increase of temperature could affect the gametes. 5. openings of prototypes have been reported during its placement or removal from the vaginal cavity. 6. to prevent expulsion of the device, a diaphragm was used to maintain the device in the vagina during the period of incubation. a risk of bacterial vaginosis was increased as vaginal secretions were retained behind the membrane of the diaphragm. 7. one major advantage of the iNvo technique is that corona cells surrounding the zona pellucida and the cumulus cells are spontaneously removed after 2 or 3 days of vaginal incubation. the disadvantage of these spontaneously denuded embryos was the difficulty to find them rapidly under microscope in the large volume of medium even for a trained embryologist (chamber volume of the prototype equals 3 ml). b. INVOcell. the new device has been specially designed for
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ISAR Express February 2020 CLINICAL ART tHe RetentIon DeVIce the retention device along with the culture device is placed in the vagina of the patient.
the iNvo procedure which addresses the disadvantages described above. iii. Disposable equipment: other disposable equipment than the iNvocell device are necessary to perform the iNvo procedure such as: a. Needle for oocyte retrieval b. Cover for the vaginal probe c. Petri dishes, tubes, pipettes, container to collect the sperm, slides and gloves used to prepare the gametes d. embryo transfer catheter e. Culture media: saline solution to wash the vaginal cavity, medium to flush the follicle if used, gradient density for sperm preparation, culture medium to wash the sperm and used in the iNvocell. iv. capital equipment: a. rapid hormonal assay instrument. hormonal assays are important to evaluate the follicle growth and the timing of ovulation. instrumentation for rapid hormonal assays is generally available in clinical laboratories. small instruments with individual disposable test units are also available. leasing of the instrument is generally included in the cost of the disposable test units. b. Ultrasound machine with vaginal probe and vacuum pump. Ultrasound is also valuable in the monitoring of ovulation and is necessary for the follicle retrieval. the ultrasound machine with vaginal probe is generally available in the office of a physician who treats infertility. the investment to get the ultrasound machine with the vaginal probe, its needle guides and the vacuum pump to aspirate the follicular fluid may vary from $7000 to $30 000, depending of the age of the instrument and where it is purchased. c. laminar flow including warm bench, small incubator and stereomicroscope with video system and bench centrifuge. this capital equipment is desired to perform the biologic steps of the iNvo procedure. identification of oocytes in the follicular fluid, sperm preparation and embryo observation can be performed successfully without clean environment. however,
tHe cULtURe DeVIce this holds the eggs and the sperm. it is about 1.5 inches in height and just over 1 inch wide
we recommend the use of a laminar flow hood for the iNvo procedure to decrease the risk of contamination and major variations in temperature of the culture media and gametes. REFERENCES1.Paulson RJ, Fauser BC, Vuong LT, Doody K. Can we modify assisted reproductive technology practice to broaden reproductive care access? Fertility and Sterility. 2016;105(5):1138-1143. 2Lucena E, Moreno-Ortiz H. Minimally Invasive IVF as an Alternative Treatment of Option for Infertility Couples: (INVO) Procedure. Development of In Vitro Maturation for Human Oocytes. 2017:267-279. 3.Mitri F, Esfandiari N, Coogan-Prewer J, et al. A pilot study to evaluate a device for the intravaginal culture of embryos. Reproductive BioMedicine Online. 2015;31(6):732-738. 4.Doody KJ, Broome EJ, Doody KM. Comparing blastocyst quality and live birth rates of intravaginal culture using INVOcell™ to traditional in vitro incubation in a randomized open-label prospective controlled trial. Journal of Assisted Reproduction and Genetics. 2016;33(4):495-500.
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LYMPHOCYTE IMMUNIZATION THERAPY FOR RECURRENT MISCARRIAGE DR POONAM LOOMBA MBBS,MD Gynecologist and Infertility Specialist, Loomba Maternity and ENT Hospital, Ambala
DR KABERI BANERJEE MBBS, MD (AIIMS), FRCOG (UK), Commonwealth Fellow IVF (UK) Medical Director of Advance Fertility & Gynecological Center, New Delhi
Fig.1 Maternal-fetal immune tolerance IntRoDUctIon recurrent miscarriage i.e recurrent abortion or habitual abortion is post implantation failure in natural conception is defined as 3 consecutive pregnancy losses prior to 20 weeks from the last menstrual period. approximately 1–5% of women of reproductive age suffer from recurrent miscarriage (rM) (1). the etiological causes include chromosomal anomalies, anatomical disorders, endocrine factors, thrombophilic factors, au-
toimmune abnormalities, and reproductive tract infections (2,3). in 50% cases of rM, the causes are not clear which is called unexplained recurrent miscarriage (urM). it is largely associated with the failure of fetomaternal immunologic tolerance, and it usually occurs in the first trimester of pregnancy (4). IMMUnoLoGy In PReGnAncy in a normal pregnancy, the mother accepts the fetus
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ISAR Express February 2020 CLINICAL ART and maintains its development as an allograft that benefits from maternal-fetal immune tolerance (Fig 1). the homeostasis of th1/th2 cytokines regulates maternal-fetal immune tolerance during pregnancy. During pregnancy, the increase in progesterone levels induces th2-dominant state, increases il-4, il-6 and il-10 to protect the embryo from attack by the immune system, inhibits secretion of th1 cytokines and decreases il-2, il-12 and iFN-γ which enhance the immune cytotoxicity of Nk cells, thus inhibiting embryonic implantation, trophoblast growth, and embryonic development. embryo directly contributes to th2 dominance by secreting il-10 & tGF β. the th2-type cytokines lead to the induction and maintenance of allograft tolerance and th1- dependent effectors play important roles in acute allograft rejection in the urM patients (4,5,6). Maternal and fetal CD4+CD25+FoXP3+ regulatory t cells (tregs) also contribute to the acquisition and maintenance of tolerance during pregnancy by suppressing maternal allogeneic immune responses in peripheral tissues. the percentages of CD4+CD25+ cells in peripheral blood are lower in urM patients as compared with normal early pregnant women (7). so, the th1/th2/ treg paradigm disorders may be a key target in urM.
and if not, they received another PBMCs intradermal administration. after conception, the urM group of patients received lit twice every 8 weeks to strengthen their active immunity (11).
DIScUSSIon lymphocyte immunotherapy is a therapeutic option for couples with urM as it is beneficial for restoring balance in the th1/th2/treg paradigm and improving pregnancy outcome in urM patients. it has been used for urM since the 1980s, and several studies have showed an increase in positive pregnancy outcomes after lymphocyte immunotherapy in urM patients (10,11). there were various factors that predicted the success of lit. it offered a significantly higher success rate in patients with immune abnormalities compared to patients without any such abnormalities including autoantibodies (12,13). reported dose of lit showed that immunotherapy performed with high-dose lymphocytes (more than 1× 108 lymphocytes) exerted less effect on improving the live birth rate, while lit performed with lowdose lymphocytes (less than 1 × 108 lymphocytes) improved the success rate significantly for patients with urM. there were better results when administered intradermally then subcutaneous injection (14). the use of stored fresh and not refrigerated lymphocytes showed dePRIncIPLe oF LyMPHocyte IMMUnotHeRAPy (LIt) the deficiency in proper recognition of fetal alloantigens by creased miscarriage rates and improved the live birth rate and lit performed before and during pregnancy produced the maternal immune system is associated with urM and a better outcome compared with that performed only bemay be prevented by boosting the maternal immune response with paternal or third party lymphocyte immuniza- fore pregnancy (15,16). the risk of miscarriage due to adtion. the low-dose lymphocyte immunotherapy (lit) could vanced maternal age is associated with a higher incidence effectively decrease the abnormally high levels of th1 cells of embryonic chromosomal abnormalities. the increasing number of previous miscarriages negatively affected lit and increase the abnormally low levels of th2 and treg success among patients with primary rM, but among pacells and also increases the concentration of tGF-β1 in serum which is known to promote FoXP3 expression by in- tients with secondary rM, the opposite occurred: lit outducing treg differentiation from CD4+CD25+ t cells and in- comes tended to be better with increasing number of previous miscarriages. the limitations in these studies were hibit the activity of many cytokines, including iFN-γ and tumor necrosis factor-alpha (tNF-α). this can then lead to a that they had not performed genetic testings of products of conception to rule out chromosomal abnormalities and restoration in the balance of the th1/ th2/treg paradigm also not reported the side effects of lit. the most common and therefore, paternal or third party lymphocyte immuside-effect of lit is reaction at the site of intra- dermal innization has been used treatment for alloimmune-medijection (13). ated miscarriages (8,9,10). there was strong evidence that locally secreted cytokines PRePARAtIon control the implantation process and can cause implantaabout 15 ml of peripheral venous blood was drawn from tion failure. Women with recurrent spontaneous abortions patients’ husband in the immunotherapy group and PBMCs and infertility of multiple implantation failures after ivF– embryo transfer have significantly higher th1/th2 ratios as were isolated by Ficoll-hypaque centrifugation (tianjin compared with normal fertile women. some non-randomhaoyang Biological Co. ltd., China). after centrifugation, ized studies shown a beneficial effect of allogenic leukocyte PBMCs were collected from the interphase layer and washed 3 times with sterile sa- line and resuspended in 0.7 alloimunization in women with recurrent implantation failml of sterile saline at a concentration of 1.6 × 107 cells/ml. ure (17). But there were no randomized trials to conclude that the allogenic leukocyte immunization treatment was the prepared PBMCs (1× 107 cells) were administered 3 times intradermally every 3 weeks. the patients with urM beneficial for repeated ivF failures (18). the therapy is still controversial in terms of effectiveness. were then examined again with respect to the th1/ th2/treg paradigm to determine whether they conceived, there may be side effects to mother and fetus due to un-
CLINICAL ART ISAR Express February 2020 predicted immune response to either autologic or allogenic blood components. By analyzing 12 reliable studies, the Cochrane review by Wong lF et al. reached the conclusion that lymphocyte immunization therapy provided no significant beneficial effect over placebo in preventing further miscarriages (19). there may be better and safer options like intravenous immunoglobulin therapy for women with urM that are associated with high clinical pregnancy and live birth rates. But it needs for randomized trials to proven the other newer options. therefore, lymphocyte immunotherapy is one of the possible treatment of unexplained recurrent miscarriages but the success rates depends on the patient’s age, previous obstetrical history and associated autoimmune and thrombophilic factors. More randomized clinical trials including group of patients without autoantibodies are needed to determine efficacious immunotherapies.
Ding L, Sun H. Low-dose lymphocyte immunotherapy rebalances the peripheral blood Th1/Th2/Treg paradigm in patients with unexplained recurrent miscarriage. Reprod Biol Endocrinol. 2017;15(1):95. 11. Liu Z, Xu H, Kang X, Wang T, He L, Zhao A. Allogenic lymphocyte immunotherapy for unexplained recurrent spontaneous abortion: a meta- analysis. Am J Reprod Immunol. 2016;76(6):443–53. 12. Prins JR, Kieffer TEC, Scherjon SA. Immunomodulators to treat recurrent miscarriage. Eur J Obstet Gynecol Reprod Biol. 2014;181:334–7. 13. Cavalcante MB, Sarno M, Niag M, Pimentel K, Luz I, Figueiredo B, Michelon T, Lima S, Machado IN, Araujo E Junior, Barini R, Neumann J. Lymphocyte immunotherapy for recurrent miscarriages: Predictors of therapeutic success. Am J Reprod Immunol. 2018;79(6):e12833. 14. Cavalcante MB, Sarno M, Araujo Júnior E, Da Silva Costa F, Barini R. Lymphocyte immunotherapy in the treatReferences: ment of recurrent miscarriage: systematic review and 1. Toth B, Wuerfel W, Bohlmann MK, Gillessen-Kaesbach G, meta-analysis. Arch Gynecol Obstet. 2017;295(2):511–518. Nawroth F, Rogenhofer N, Tempfer C, Wischmann T, von 15. Clark DA. Immunological factors in pregnancy wastage: Wolff M. Recurrent miscarriage: diagnostic and therapeutic fact or fiction. Am J Reprod Immunol. 2008;59(4):277–300. procedures. Guideline of the DGGG (S1-level, AWMF reg16. Sarno M, Cavalcante MB, Niag M, Pimentel K, Luz I, istry no. 015/050). Geburtshilfe Frauenheilkd. 2013;75(11): Figueiredo B, Michelon T, Neumann J, Lima S, Machado IN, 1117–28. Araujo E Júnior, Barini R. Gestational and perinatal out2. Larsen EC, Christiansen OB, Kolte AM, Macklon N. New comes in recurrent miscarriages couples treated with lyminsights into mechanisms behind miscarriage. BMC Med. phocyte immunotherapy. Eur J Obs Gynecol and Reprod Bio: 2013;11:154. X 3 (2019) 100036 3. Ford HB, Schust DJ. Recurrent pregnancy loss: etiology, 17. Kling C, Magez-Zunker J, Jenisch S, Kabelitz D 2002 Exdiagnosis, and therapy. Rev Obstet Gynecol. 2009;2(2):76– perience with allogenic leukocyte immunization (AI) for im83. plantation failure in the in vitro fertilization program. Am J 4. Del Prete G, Maggi E, Romagnani S. Human TH1 and TH2 Reprod Immunol. 2002; 48, 147–150. cells: functional properties, mechanisms of regulation, and 18. Urman B, Yakin K, Balaban B. Recurrent implantation role in disease. Lab Investig J Tech Methods Pathol failure in assisted reproduction: how to counsel and man1994;70:299–306. age. A. General considerations and treatment options that 5. Christiansen OB, Nybo Andersen AM, Bosch E, Daya S, may benefit the couple. Reprod Biomed Online. Delves PJ, Hviid TV, et al. Evidence-based investigations and 2005;11(3):371–381. treatments of recurrent pregnancy loss. Fertil Steril 19. Wong LF, Porter TF, Scott JR. Immunotherapy for recur2005;83:821–39. rent miscarriage. Cochrane Database Syst Rev. 6. Franasiak JM, Scott RT. Contribution of immunology to 2014;(10):CD000112. implantation failure of euploid embryos. Fertil Steril. 2017;107(6):1279–1283. 7. Cohen JL, Trenado A, Vasey D, Klatzmann D, Salomon BL. CD4(þ)CD25(þ) immunoregulatory T cells: new therapeutics for graft-versus-host disease. J Exp Med 2002;196:401–6. 8. Zheng SG, Gray JD, Ohtsuka K, Yamagiwa S, Horwitz DA. Generation ex vivo of TGF-beta-producing regulatory T cells from CD4+CD25- precursors. J Immunol. 2002;169(8):4183– 9. 9. Yang H, Qiu L, Chen G, Ye Z, Lü C, Lin Q. Proportional change of CD4+CD25+ regulatory T cells in decidua and peripheral blood in unexplained recurrent spontaneous abortion patients. Fertil Steril. 2008;89(3):656–661. 10. Liu M, Zhen X, Song H, Chen J, Sun X, Li X, Zhou J, Yan G,
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ISAR Express February 2020 GENETICS
EDITING THE
GENOME DR DEEPAK MODI Scientist E and Head Molecular and Cellular Biology laboratory ICMR-National Institute for Research in Reproductive Health Indian Council of Medical Research, Parel, Mumbai 12
note: the views expressed in the article are of the author and not necessary of those of iCMr or Nirrh or isar.
Classical textbooks of genetics teach us that genetic diseases cannot be treated and management is the only option. however, advances in genome research and technologies have provided us with the options of editing the DNa and provide means to cure genetic diseases. this commentary is aimed to provide a basic understanding of this technology and the current status of its use in assisted reproduction. Finally, the article will summarize the way forward on the use of genome editing in reproductive medicine. What is genome editing? like in a conventional text editor, one would need tools to identify the errors, cut at the right places and either delete or replace the inappropriate texts in the genome. the molecular tools that are used in genome editor are CrisPr and Cas9. CrisPr stands for Clustered regularly interspaced short Palindromic repeats. the first requirement is a rNa strand that contains the genome alphabets that we may want to edit. this rNa sequence has to be incorporated in a protein called Cas9. the rNa sequence guides the Cas9 and takes it to the specific locations where Cas9 (the molecular scissors) cuts at the DNa. as a result, it will either delete the target sequences or allow insertion and replacement of the genome alphabets. see the video here (https://www.broadinstitute.org/research-highlights-crispr) for an animated version of how CrisPr works. applications of genome editing in reproductive medicine since its original discovery CrisPr, the technology has undergone several modifications with each newer version better over another. the present day CrisPr technology is very precise, robust and highly reproducible. it is now adaptable in most laboratories with relative ease and researchers have used this tool to inactivate genes, edit different parts of genomes and even repair faulty genes to correct genetic diseases in cells gown in lab dishes, worms, flies and even mice. in assisted reproduction, Preimplantation Genetic testing (PGt) was traditionally introduced to screen out genetically defective embryos which are discarded and normal embryos are offered for transfer. however, it it not more logical and perhaps ethical, to “fix” the abnormal gene rather than allowing an
affected embryo to be generated only to be discarded later ? . For this we need to move from Preimplantation Genetic testing (PGt) to Preimplantation Genetic Correction (PGC). in the last five years researchers have successfully applied CrisPr technology and demonstrated precise genome editing in human embryos (reFs). By injecting the CrisPr, it has been possible to achieve correction of the beta thalassemia mutations, a mutation causing cardiomyopathy and also delete the gene causing hiv entry in to the cells (reFs). two live births have also been reported of babies who have had their CCr5 gene deleted in the embonic stages and are now supposedly free from any possibility of hiv infection. these are unprecedented feats and we have witnessed a history in making. all these novel studies aimed to have largely address if CrisPr technique can “repair” every single cell (preventing “mosaicism”) and avoid gene editing at other sites in the genome (rates of “off target effects”) in human embryos. While the technology is evolving and methods being optimized, the results of four major studies published in scientific journals have been quite assuring. With the experience from four independent studies, the following points need to be borne in mind while considering CrisPr technology in assisted reproduction in the present scenario. 1) human embryonic genomes could be successfully edited with minimal to almost no off-target effects. 2) We yet don’t understand the precise mechanism by which the editing has been achieved inside the embryonic nuclei 3) the technology is successful (and reliable) only when applied in oocytes just prior to fertilization; trying to edit after fusion of the 2PN is unreliable and leads to mosaic embryos. 4) Before editing, one needs to diagnose which embryos are carrying the mutation. thus, a PGt will need to be applied before PGC. Unfortunately, PGt is presently most reliable at the blastocyst stage when PGC can not be applied. thus, at best, present day CrisPr technology can be applied to genetic conditions that are maternally inherited. thus, the scope of application is very limited. More research is required to improve our CrisPr tools and methodology of delivery to minimize mosaicism and maximise efficiency. Despite these limitations, i see the glass as half full.
GENETICS ISAR Express February 2020
We have overcome many hurdles and progressed with the the technical dos and don’ts. it’s a matter of time that they will be optimized and the powerful tools of genome edited embryos will be made available in the clinics across the world. soon we may have clinics that may not just make embryos, but diagnose them and even correct them before initiating the pregnancy. Definitely the realms of an art clinic will expand beyond fertility treatment. however, concerns have been raised regarding the abuse of CrisPr technology. should editing of mutations for treatable conditions allowed? should we offer gene editing to families that carry mutations of late onset disease or those that increase the cancer risk but not always cause it (e.g., BrCa1 and breast cancer). the worry of “positive eugenics” of “desired traits” (e.g intelligence, physical appearance) and “social genetics” (hla matched embryos) is growing amongst the circles of social scientists and bioethicists. Not surprisingly, with the announcement of the Chinese scientists reporting birth of lulu and Nana (the CCr5 gene edited babies), a section of the scientific and the bioethics fraternity have displayed a knee jerk reaction and called for a total ban on human embryo editing (reFs). in india too, iCMr has imposed a ban on embryonic genome editing until proven safe (reF), however the system is open to discussion when needed. however, the “slippery slope” and “designer baby” concerns
that have resurfaced are not unfamiliar to the art fraternity. these dilemmas and knee jerk reactions have already encountered by us when ivF was first demonstrated and PGt was first applied clinically. asn this is a matter of concern. the horrifying stories of Bob edward and siddharth Mukherjee are the witness to the hostility the system. instead of reinventing the wheel each time, lets learn from these examples and not let history repeat itself. let us accept that germ line/embryonic genome editing is a reality and beneficial to the society at large. Banning and pushing it under the carpet with the hope it will die or improve by itself is being immature and definitely not scientific. it will only lead to its further abuse and cause more harm than good. let’s not the ethics lag the scientific progress. instead, it’s time to work together and set up the guidelines to define steps that will be required to appropriately use CrisPr technology for clinical use in assisted reproduction. it should be the responsibility of the medical and scientific fraternity to take the appropriate steps and establish progressive guidelines to make sure that the human embryo genome editing is placed in morally responsible hands. let us not leave it only to the policy makers and a social scientist to make the regulatory guidelines for us and our patients. We are and should be the torch bearers to develop the guidelines and make a positive impact of the genome editing technology for the benefit of mankind.
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INTERVIEW
Master of All Dr Narendra Malhotra in conversation with Dr Neharika Malhotra & Dr Keshav Malhotra Q. Jack of all master of none. you are one person who has contradicted this statement! Well, if you have a will, if you have a drive and if you have a temperament and if you had a mother like late Prabha Malhotra, you are going to do everything and do it well. i love the challenges life throws and take him head on. after 20 years of hard-core obstetrics, delivering over 20,000 babies then ultrasound, then laparoscopy, then art and now Genetics, aesthetic regenerative medicine and anti-ageing that has been my developing interests. academics, lectures, teaching, organising CMes, conferences are hobby with social service, clubs, travelling, sports etc thrown in as fillers. Basically, i am a personality who can handle many things at the sometime; a ‘Multiplug’.
Q
what made you choose this profession? Family profession. My grandfather late r B & s N Malhotra Was a leading practitioner of india and a great academician of the 60’s. he was the inspiring force and of course parents were the motivators. i never looked at any other profession except the armed forces briefly and luckily did not get into NDa.
Q. Beti Bachao Beti Padhao – is it true this slogan was coined by you – what’s the story behind this? Well social obstetrics was always my passion and going to villages to help was always a dream. so when i got elected as FoGsi president in 2008 i chose Chalo Gaon Chale… theme with the slogan of Beti Bachao Beti Padhao. those days sex determination & selective abortions were rampant and we (our family) was of the strong opinion that this is a social problem and only girl education and empowerment could combat this social evil. hence this slogan and to this was added let’s eradicate WeeeP (Women education, employment, environment & empowerment problem). the motive was to educate & empower girls which unfortunately did not succeed, as today, we see that girls may be educated may be more than boys even in the villages, but are still not empowered and safe – so we have changed our slogan to Beta samjhao Beti Bacho. it’s time now to educate the boys to empower the girls and to accept empowered girls. Q. you have been the president of ISAR, ISPAt, IFUMB, FoGSI, etc. what’s your take on organization politics? to become the FoGsi president was a dream of my mother which i had to fulfil. so, from the very first
ISAR Express February 2020 appearance in FoGsi in 1986, Durgapur our (family’s) aim was that one of us to become President of FoGsi. We did not belong to any political groups and it was my love for teaching ultrasound which took me to all corners of india and made all my elections and posts come easily. side by side we were all active in other organisations and rose up the ranks of these to reach presidentship. Jaideep & i are the only couple in the world to be president of 3 professional organisation and also FrCoG. Q. what does ‘good life’ mean to you? Money is important in life to survive and to work on what you want to but our dictum has been do not run after money- just work hard & honestly and let money run after you. keeping in shape, healthy and fit is a necessity which i realised after 40. though an active sportsman in college days, work schedule kept me away from the gym etc. But since last 20 years it’s been regular yoga, walks, meditation and relaxations. When you are relaxed and at peace, creativity comes naturally. i am a little trigger happy with ideas and can churn up creative ideas very quickly. Q. How do you see yourself in the next ten years? Working even harder, being a little more healthier & fitter, indulging in creativity and writing (maybe 2-3 fiction novels and a biography). and of course, bring up grandchildren with good values and morals. Zinda Rahe to Phir Milenge Milte Rahe Tabhi to Zinda Rahenge… and i remember a song by tom Jones which i have modified two words I am rich in friends A millionaire I have so many It seems unfair So why don’t you Take a share Just help yourself To my friends And say the world’s Good to me. with love and smiles to all!
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ISAR Express February 2020 CLINICAL ART ART & LuTeAL phAse suppoRT
DR KABERI BANERJEE MBBS, MD (AIIMS), FRCOG (UK), Commonwealth Fellow IVF (UK) Medical Director of Advance Fertility & Gynecological Center, New Delhi
Revisiting the role of Vaginal progesterone IntRoDUctIon the first pregnancy after in vitro fertilization (ivF) of a human egg and the first birth of an ivF baby were reported in 1976 and 1978, respectively. since then, an estimated seven million pregnancies have been achieved worldwide by ivF and its modifications. as experience has accumulated, art success rates have increased and now accounts for 1 to 3 percent of live births in the United states and europe. however, assisted reproductive technology cycles are known to have an insufficient luteal phase, probably due to the supra-physiologic oestrogen levels in ivF and iCsi in the follicular phase, as a result of ovarian stimulation used to prepare for oocyte retrieval . therefore, sufficient luteal phase support (lPs) is essential during these cycles to improve implantation and pregnancy rates. LUteAL PHASe SUPPoRt luteal support (ls) is considered important in preventing luteal insufficiency and its negative impact on early pregnancy. it is well known that luteal function is compromised in stimulated ivF cycles. in ivF cycles that did not have luteal support, luteal phase length was shortened and early bleeding became more frequent. the meta-analysis and two Cochrane systematic reviews confirmed that luteal support promotes the successful outcome of the ivF procedure. Progesterone plays a crucial role in ls in ivF/ iCsi cycles.
latation & uterine musculature quiescence by inducing No synthesis . although the need of luteal phase support in ivF/iCsi cycles is well known the optimal start, dosage, route and the duration of the lP support is still subject of a debate. VAGInAL PRoGeSteRone amongst the various routes of drug delivery, vaginal route is preferred due to the "first uterine pass" effect . vaginal route circumvents the variable absorption and high first-pass hepatic metabolism after oral ingestion and also prevents the uncomfortable and sometimes painful iM injection. vaginal administration ensures a stable progesterone concentration in the endometrium even when serum levels are low, thus reducing the risk of systemic effects. it is the most used route of progesterone in luteal phase support of induced cycles. the survey from 2014 of 284 600 ivF cycles in 82 centres worldwide, stated that 77% of the cycles were performed with vaginal progesterone only and an additional 17% used vaginal progesterone in combination with oral or intramuscular progesterone (iMP)4. a study by heine P, et.al published in 2017 overall experience and patient convenience of vaginal progesterone tablets used for luteal phase support (lPs) during in vitro fertilization (ivF) treatment . it was a questionnaire-based audit which included responses from 100 patients undergoing ivF treatment.
PRoGeSteRone AS LUteAL PHASe SUPPoRt to maximise pregnancy rates following assisted reproductive techniques (art), luteal phase support (lPs) is now routinely provided by either progesterone, using various routes of administration, or human chorionic gonadotrophin (hCG), although hCG has been associated with higher rates of ovarian hyperstimulation syndrome (ohss) . Progesterone supplementation is generally initiated on the day of oocyte retrieval or at the time of embryo transfer. Progesterone is involved in the induction of secretory transformation of endometrium in luteal phase. it is also involved in the improvement of endometrial receptivity and implantation rates in in vitro fertilization (ivF) cycles. Progesterone also promotes local vasodi-
Figure 1: Patient preference among progesterone formulations
CLINICAL ART ISAR Express February 2020 going frozen-thawed embryo transfer cycles. the study showed no significant differences between the four preparations in terms of clinical pregnancy rate, fetal heartbeat rate, and miscarriage rate. a study by Jiang lei published in 2019 compared vaginal progesterone vs. intramuscular progesterone was done in 3013 patients undergoing artificial cycle for frozen-thawed embryo transfer. results showed that vaginal progesterone supplementation had significantly (P < 0.05) greater implantation (37.0% vs 34.4%), delivery (45.1% vs. 41.0%) and live birth (45.0% vs. 40.8%) rate than intramuscular progesterone injection. another study by hongbin Chi, et.al published in 2019 cLInIcAL eVIDence – also showed that vaginal gel preparation was non-infeReVIew oF Recent eVIDenceS rior to intramuscular progesterone and therefore can a systemic review and meta-analysis of rCts by abdelbe an alternative method of luteal phase support in ivF hakim, et.al published in 2020, showed that vaginal progesterone is similar to intramuscular Progesterone in patients . clinical pregnancy, ongoing pregnancy, miscarriage, and to date, several studies have reported transvaginal progesterone doses in the range of 200-1200 mg. live birth rates. it was also associated with more satisenatsu, et.al evaluated the effectiveness of high-dose faction compared to intramuscular Progesterone4. it also mentions that vaginal progesterone has similar effi- progesterone supplementation for women undergoing cacy even in frozen embryo transfer cycles too, which is a frozen-thawed embryo transfer (Fet). 1188 were important as frozen cycle lacks the endogenous proges- 1200 mg/day of vaginal progesterone, while 822 were 900 mg/day. the results showed that clinical pregnancy terone secretion due to the absence of functional corrate was higher in the 1200 mg group than in the 900 pus luteum, and it almost depends on the externally mg group in both younger and older cohorts. these reprovided hormones for endometrial preparation. sults indicate that the factor that resulted in better outa study by saunders et.al published in 2020, evaluated comes in the 1200 mg group was simply higher the non-inferiority of vaginal pessaries (400mg) to vagiendometrium progesterone concentrations. nal gel (90mg) in 769 women undergoing in vitro fertiliProgesterone supplementation in the luteal phase is assation (ivF)5. in full analysis, clinical pregnancy rates on D38 were 38.3% for progesterone vaginal pessaries and sociated with significant improvement in pregnancy rate and reduction in miscarriage rates. Further, vaginal 39.9% for progesterone vaginal gel. Further they concluded that Progesterone 400 mg pessaries bid for luteal route of administration is found to be most beneficial without any major side effects. it is a time-tested drug phase support is an effective, safe and tolerable treatfor luteal phase support. ment option for women undergoing ivF during art. results showed that overall, the majority of patients found progesterone tablets “very easy” to use. Most patients (90.7%) described their overall experience with the vaginal progesterone tablets as “very comfortable” or “neither comfortable or uncomfortable,” and 87.5% found them to be “very convenient” or “neither convenient or inconvenient” to administer. Most patients found treatment with progesterone tablets to be more convenient and more comfortable, and 60.6% indicated that the progesterone tablet was their preferred progesterone formulation.
eVIDence In FRozen eMBRyo tRAnSFeR cycLeS: in contrast to Fresh-et, the absence of endogenous serum progesterone (P4) secretion before frozen embryo transfer in hormone replacement therapy cycles (hrt-Fet) results in the need for exogenous progesterone formulations at approximately pregnancy weeks 8 to 10. recently published study by Bu, et.al (2019), evaluated the impact of endometrial thickness change after progesterone administration on pregnancy outcome in patients transferred with single frozen-thawed blastocyst. the results showed that dynamic change of endometrial thickness after progesterone administration in Fet cycles. an increased endometrium after progesterone administration was associated with better pregnancy outcome. interestingly, clinical pregnancy outcomes and the increasing rate of endometrium were positively correlated . shiba, et.al. evaluated efficacy of four vaginal progesterones for luteal phase support in 259 patients under-
https://www.uptodate.com/contents/pregnancy-outcome-after-assisted-reproductive-technology https://www.uptodate.com/contents/in-vitro-fertilization?sectionName=Pre-cycle%20factors%20associated%20with%20success&topicref=6788&anchor=h4& source=see_link#h789331013 reproductive biomedicine online, 2018; 36: 630-645. Gynecol endocrinol. 2019 apr 29:1-6. saunders, et.al. human reproduction, 2020: 1–9. Fatemi et al F, v & v in oBGyN 2009, 1(1): 30-46. ismail aM, et al. J Matern Fetal Neonatal Med. 2017 Feb 15:1-7. Patient related outcome Measures 2017:8 169–179. reprod Biol endocrinol. 2019 Nov 25;17(1):99 reprod Med Biol. 2020;19:42–49. scientific reports, 2019; 9:15264 european Journal of obstetrics & Gynecology and reproductive Biology, 2019; 237: 100–105 reprod Med Biol. 2018;17:242–248.
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“Love is in the Air” “Only love let’s us see normal things in an extraordinary way.” Saroj & Dr Rajeev Agarwal
“Unconditional love never fails any test.” Jaideep & Narendra Malhotra
“Every smile is for you.” Monica & Randhir Singh
Sushma & Anupam Gupta
" Every journey is possible and enjoyable with you by my side including the road less travelled." Leena & Amit Patankar
ISAR’s
Valentine Couples Endless Love - For Each Other Endless passion - For IVF
“We feel newly married all over again” Rishma & Hrishikesh Pai
A successful marriage requires falling in love many times, always with the same person. Anshu & Sunil Jindal
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ISAR Express February 2020 TIPS FOR DOCTORS
SMALL STEPS TO BIGGER OUTCOMES
EXERCISE Exercise is a great way to fight stress. It not only improves metabolism and energy. It also slows down the process of aging.
LOOK AFTER YOUR SKIN Hydrate yourself well and take care of your diet. Use moisturizers according to your skin type for optimal benefits.
KEEP UP THE ANTIOXIDANTS we all know that oxidative stress and reactive oxygen species are responsible for ageing. To fight this, make sure you take antioxidants like vit C vit E, CoQ10 beta carotene, superoxide dismutase etc. Include brightly coloured vegetables and fruits in your diet. Organic Farming and pesticide free food to be incorporated. MEDITATE DAILY A 10 min meditation daily would do wonders in your health both inside and outside.
GET YOURSELF INTO A ROUTINE Incorporating yourself into a proper routine will help your balance life both professionally & decrease your stress levels.
IN THE END ALWAYS BE GRATEFUL. GRATITUDE IS THE BIGGEST QUALITY TO CALM THE MIND.
ISAR Express February 2020
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DR. NIDHI SINGH TANDON
Beautiful
MBBS, MD (Dermatology, Venereology & Leprosy) The Skin Art Clinic, Lucknow
ins ide OUT
Doctors should look and feel beautiful too
How important is keeping yourself fit and fab?? Especially for people who work day in and day out to take care of others as a part of their daily duties. Itâ&#x20AC;&#x2122;s absolutely necessary to keep yourself fit and looking your best at any age.
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ISAR Express February 2020 wHAt MAtteRS
SIMRAN BINDRA Head Dept of Sociology and Anthropology The Shri Ram School, Moulsari Campus.
DR. NEHARIKA MALHOTRA BORA MBBS, MD (Obstetrics & Gynaecology) Global Rainbow Hospitals, Agra
coMMUnIcAtIon Patient satisfaction is achieved through the communication behaviours of doctors; their dominance, concern about the psycho-social issues of the patient, illness perceptions, concordance between the doctor’s communication style, the patient’s need for attachment, and the patient’s attitude. as the doctor’s perception is what leads to patient satisfaction and clear evaluation during clinical judgement, there must be no ‘negative space’ for communication. the Doctor must clearly lay out the ambiguities and uncertainties in the process of diagnosis and treatment, and the patient must be able to accept the disease analysis. there should be a two way communication on the doctor’s expectations from the patients on coping mechanism and vice versa. Communication therefore should be strong, clear and appropriate. the healthiest relationship requires honesty, trust and mutual respect. eMPoweRMent the doctor-patient consultation previously had a set format, ‘in assuming the sick role, the patient expects to offer cooperation and compliance to the doctor, in return for some assurance that attempts will be made to relieve the distress, the patient now understands both the benefits and the risks of modern medicine, desiring all information before placing their trust in the doctor. this implies that the patient is now the ‘service user’ and ‘the client’, where there is supposedly a more ‘equal relationship between
PoIntS to ReMeMBeR 1. DoCtors are Not sUPerhUMaN or MaGiCiaNs, they are FoCUsseD aND MotivateD at healiNG as hUMaNBeiNGs. 2. DoCtor – PatieNt relatioNshiP has to Be syMBiotiC iN NatUre so that it is MUtUally MotivatiNG. 3. there shoUlD Be No CoNFliCt oF iNterest.
the professional and the purchaser of a service. the information exchange and the health consumer movement have led to the current communication model of shared decision making and patient-centred communication. StReSS the Doctor’s role in managing stress of the patients super cedes his ability of medical know how and involves intensive psychological counselling of the patients in establishing trust and providing reassurance. however Doctors can have good days and bad days, and with cuts in funding, low staff numbers and huge pressures on time, it’s almost necessary to not treat patients as such. it’s the most sought after skill to be able to make patients feel like you have all the time in the world when you’ve got a target of six minute appointments. the hospital environment is demanding and Competitive. Patients have unrealistic expectations of what to expect in a public sector, you can’t offer hotel service. the expectation of the patient should be centred on the diagnosis and the consecutive treatment. A SPecIAL ReLAtIonSHIP the doctor - patient relationship is a special one and remains a keystone of care, concern and community service. it is critical for vulnerable patients as they experience a heightened reliance on the physician's competence, skills, and good will. the doctor–patient relationship is remarkable for its centrality during life-altering and meaningful times in a persons' lives, at times of birth, death, severe illness, and healing. thus, providing health care, and being a doctor, is a moral enterprise.
4. iNForMatioN FroM the iNterNet shoUlD Not aloNe Be a soUrCe oF iNQUiry For the PatieNt. 5. the eleMeNt oF trUst is the Core iNGreDieNt For healiNG. AnD ReMeMBeR A DoctoR-PAtIent ReLAtIonSHIP IS SAcReD.
DOCTOR-PATIENT RELATIONSHIP In today’s economic climate there is much discussion around lack of resources, as healthcare represents one of the largest expenditures in the world. the doctor-patient relationship is a social system in which roles are defined. Social pathogenesis highlights the need for patient empowerment where people care about the medical experience that their doctor has, but do they realise the impact that the doctor’s actual experience might have on the doctor themselves and hence the patient? A Positive Doctor-Patient relationship is essential for good outcome to any treatment.
THE BOND
OF TRUST
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ISAR Express
Vaastu Tips for a Stress Free Home h ome is said to be a place where you most relaxed and calm as possible, especially in today’s fast paced and hectic and life, this is the place where you prefer going and spending quality time but sometimes our homes too are the source of stress and anxiety. sometimes i get clients saying they are not happy when they are at home or some say they get frustrated when they are alone, well in that case my explanation is, how we do yoga and meditation to keep ourselves healthy and positive, the same way our space also needs balancing with vaastu principles to create peace and happiness. as it is correctly said ‘’ your space is a reflection of your life, Body and Mind’’, here are few simple tips to make your house stress-free • Mirrors should not be in bedroom and should never be in the position that will reflect the bed. if there are any then make sure to cover them in the night. • Most important point to remember is the time should never stop i.e., the clock. if there are any watches which are broken or stopped, remove them from the house along with unwanted, broken or junk items, never keep un-repaired electronic items at home. Make sure your house should be clutter free. • Never put violent, dark, war or unhappy pictures in your house as it creates a negative environment.
• We should avoid sharp edge furniture as they cut the energy flow and changes its direction to go in the wrong way, if you already have them then try to keep it covered nicely. • Do not block North - east corner and central area of your house. it is very important to have these areas clean and uncluttered. • your sleeping position is also very important i.e., your head should never be in north direction when you sleep, best is south, east is good and west being average. • the kitchen should be in south east direction and while cooking you should face east. • if you hang wind chimes or bells in northwest corner, it brings enjoyment and joy in relationships • though black color makes the bedroom look tasteful, but according to vaastu it should be avoided in bedrooms. • the most important point i would like to highlight here is, if you have a habit of complaining and grumbling all the time, it creates negativity in your space and should be stopped, you should feel blessed because there are so many people who cannot even dream about having their own house. By following the principle of vaastu which is based on balancing the Five elements, we can have peace and happiness in our house.
Mirrors should not be in bedroom and should never be in the position that will reflect the bed.
ISAR Express February 2020
DR. SWATI JAGGI is a Vastu Consultant, Numerologist and Astrologer by Profession and have been Practicing Vastu for the last 15 Years.
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EMBRYOLOGY ISAR 2019
EVENTS ISAR Express February 2020 5th Embryology ISAR Conference was organized by KISAR on 30th November and 1st December 2019 at Bengaluru. Dr.HemaDivakar was the Chief Guest Dr.Jaideep Malhotra, President ISAR was the Guest of Honor for Inauguration Programme. Dr. S. Krishnakumar, Secretary ISAR, Dr.SudeshKamat, President Embryology ISAR were present during the programme. 3 International Faculty and more than 120 Faculty are invited for the 5th Embryology ISAR Conference. 330 delegates were attended. 3 Workshops were conducted at the Conference.
isar Presidents oration at 5th embryology isar Conference 2019
Dr. virendra shah oration at 5th embryology isar Conference 2019
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ISAR Express February 2020 EVENTS
EVENTS ISAR Express February 2020
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ISAR Express February 2020 EVENTS Ranthambore
ISAR MASTER CLASS ON MALE INFERTILITY Fertility is not solely a female issue — men are half of the fertility equation too, but are often ignored. The contribution of sperm quantity and quality to successful conception was described back in 1662 and by 1687 varicocele was being identified as a root cause of male infertility. In the 1860s, American gynaecologist James Marion Sims while investigating ‘sterile marriages’ decides to have a quick look at a semen sample under the microscope. And, wait for it… he can now see ACTUAL sperm with his own eyes! Voila, the semen analysis is born. Examination of sperm count, motility and morphology is now possible, and now with automation coming in so many wonderful and fascinating things can be done to diagnose a man as fertile or subfertile. But what to do after - most of us don’t struggle with a diagnosis but we struggle in giving a clear path to our male patients in their quest for having a child, and this is what ISAR in this 2 day masterclass wanted to demystify . Set in the backdrop of the jungles of Ranthambore where sighting tigers that are now endangered is a one in a million opportunity, ISAR took the opportunity to focus on that one in a million sperm which also is becoming somewhat endangered in our current environment. With masters of the field like Prof Ranjit Ramasamy, Dr. Rupin Shah, Dr Jaideep Malhotra Dr Narendra Malhotra and Dr Vineet Malhotra expertly navigating through the challenges in dealing with male fertility, it was an absolutely amazing academic bonanza that was offered to the delegates. Sessions ranged from initial evaluation to interpretation of the tests, chalking out treatment options, when to offer surgery, how to do the surgeries and touched upon male sexual health also. The early morning safaris were the cherries to top this cake as tigers, cubs and loads of wildlife was spotted, an humbling experience to the beauty that mother nature has offered us and how mankind can still change and conserve the wonders that nature has offered us. Like preserving male fertility, preserving nature was also a message we all took back with us.
STATE CHAPTER ACTIVITIES ISAR Express February 2020
andhra pradesh
Dr Padmaja V Chairperson
Dr Chandana V Secretary
VOGS AND AP CHAPTER OF ISAR conducted a CME on Nuances in Fertility and Pregnancy Care on 12the January 2020 at Hotel Novotel, Vijayawada. The meeting was well attended with active interaction by delegates. The Program highlights are: • Executive Body meeting, AP Chapter of ISAR: 9 am • Registration: 9.00am – 9.30 am • Inauguration : 9.30 am • General Body Meeting: 9.20 am • Session 1 : 9.30 – 10.30 am Establishing Fertility Practice: The Personnel By : Dr. Padmalatha Yarasi, Proddutur • Where the ICMR categorised infertility clinics into four levels. • The gynaecologist or physian is in charge of level 1A clinic. • Level 1A clinic does not need an accreditation under these guidelines. • Gynaecologists should have a post graduate degree or diploma in level 1B, 2, and 3 clinics. • Level 1B, 2, and 3 need an accreditation under these guidelines. • An embryologist must not be associated with more than two centres at any given time. • The counsellors and andrologists can work for more than one clinic ESTABLISHING FERTILITY PRACTICE: THE FA-
CILITY was discussed by : Dr. B. Sireesha Rani, Visakhapatnam • Where the Planning, Budgetary allocations, Expertise needed, Design of facility and Execution • And emphasis on the Team and team work QUALITY CONTROL AND MAINTENANCE OF IVF LAB was discussed by Dr. P. Sudhapadmasri M.S, Visakhapatnam • Quality control is a continues process though it may appear cumbersome initially once implemented properly, it becomes a habit. • Laboratories that succeed to implement QC system are the ones to achieve consistent and dependable results. Session 2 : 10.30 – 11.30 am Ovulation induction : Dr.Nalini, Guntur TEN POINTS OF SUCCESS • thorough evaluation to select the suitable candidate for ovualtion induction • choose the better treatment modality as per the casue • individualise the treatment regime • change the treatment modality as and when required • periodical monitoring by TVS • trigger ovulation at optimum time • adequate leuteal support • combine with IUI where ever necessary
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ISAR Express February 2020 STATE CHAPTER ACTIVITIES andhra pradesh
Dr Padmaja V Chairperson
Dr Chandana V Secretary
• treat the associated endocrine disorders • rule out the tubal and uterine causes Intrauterine insemination improving outcome: Dr Kavitha Chalasani, Vijayawada • Where IUI was discussed including Role of Sperm DNA fragmentation index 10 steps for successful IVF: Dr. Sajja Padma MD, Rajahmundry • counselling • lifestyle modification to optimise BMI and diet • local factors • right ovarian stimulation • good oocyte retreival technique • avoiding OHSS & multiple pregnancy • good culture conditions • good embryo tranfer technique • good counselling for failures • self audit Tea Break : 11.30 am-12.00 Session : 3: 12.00- 12.45 pm: AVOIDING COMPLICATIONS: Ovulation induction: Dr Chandana VMS, OBG, Vijayawada • Avoiding complications – 3P’SPrediction • Prevention • Primary (Before starting HMG/FSH) • Secondary (After starting HMG/FSH and before HCG administration) • Post - OHSS Management Avoiding Complications: Early miscarriage Dr.V.Padmaja, Chairperson AP chapter Isar, President Vijayawada OBG society Where the following was discussed: • Chance of miscarriage in general population versus IVF population • Causes of miscarriage • Miscarriage rate following different IVF treatment strategies • Cumulative live birth rate after miscarriage in the first complete cycle of IVF • Prevention of miscarriage
• Effect of miscarriage on patients Epigenetics, Role of Luteal Support in ART/IVF, Antiphospholipid Syndrome: Aspirin + unfractionated heparin Session 4: 12.45 – 1.00Pm Pregnancy following Infertility therapy: Dr. ArunaKumari, Rajamundry • Emphasis was placed on maternal & perinatal risks associated with any ART and ovulation induction pregnancy which remain high • Need to promote optimal outcomes a thorough medical evaluation and maternal health problems or health conditions should be optimised before initiating ART • make every appropriate effort to achieve a singleton gestation • appropriate counselling about the risks associated with ART • Frequent follow-ups, additional tests, & timely intervention Followed by Vote of thanks And Lunch : 1.00 Pm Program was sponsored by ABBOTT
STATE CHAPTER ACTIVITIES ISAR Express February 2020
Chandigarh
Dr Gulpreet Kaur Bedi Chairperson
Dr Pooja Mehta Secretary
Indian Society for Assisted Reproduction (ISAR) – Chandigarh chapter held its Annual Summit for the second time on 09th February 2020 at hotel Hyatt Regency, which was attended by more than 150 delegates from northern India. The objective of the event was to discuss & work on the preservation of the uterus to maintain reproductive health of women. Doctors shared their experiences in the field of gynecology & reproductive medicine, where the role of environmental changes affecting fertility was the highlight of the summit where Dr. Jaideep Malhotra – national president ISAR delivered a thought provoking lecture on the role of environment affecting fertility. Dr. G K Bedi – Chairperson ISAR Chandigarh spoke on Endometritis & infertility & how tuberculosis can be fought to achieve a pregnancy. Dr. Akshay Nadkarni the guest speaker from Surat enlightened the audience with his laproscopic skills for fertility enhancement. All in all it was an academic feast for the doctors to implement the knowledge in day to day practice. A hands on hysteroscopy workshop was conducted simulators. E-posters were introduced and the summit had 15 submissions for the competition from Rohtak, Amritsar, PGI & Govt medical college Chandigarh. The decision was tough to annonce the prizes. The Guest of honour was Mrs. Rajbala Malikthe mayor of Chandigarh, who appreciated the doctors for their unconditional service to mankind.
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ISAR Express February 2020
STATE CHAPTER ACTIVITIES
madhya Pradesh ISAR FOUNDATION DAY 16.02.2020 A free camp organized by Dr. Yatindra Singh Verma for Infertile couple at Elixir Health care polyclinic, Dal Bazar , Gwalior in collaboration with Dr. Verma hospital and Fertility centre Gwalior. 50 patients were examined & councilled. Free Ultrasound and semen examination done, to find out cause of Infertility. A walkathon and free camp organized in Bhopal by Dr. Monika Singh. Camp was organized at Dr. Mangla Kawade Sujalpur where 280 people were benefitted; was surprised to see such high incidence of Male infertility amongst farmers.organized by Chirayu Fertility Center.
Dr Randheer Singh Chairperson
Dr Anju Verma Secretary
Rajasthan Dr Sanjay Makwana Chairperson
Dr Renu Makwana Secretary
REPROMED UPDATE 2019 Date : 21st to 22nd December 2019 Venue : AIIMS Jodhpur (Raj.) Conducted by : ISAR Rajasthan Chapter, Dept. of OBGY AIIMS, Jodhpur Attended by : 150 Delegates Over the two days of 21st and 22nd of December 2019, ReproMed Update 2019 brought together practitioners and specialists from the field of reproductive medicine for a critical examination of evidence behind the prevalent practices. The two days under the flagship of Dr. Sanjay Makwana (President, ISAR Rajasthan Chapter), Dr. Renu Makwana (Secretary, ISAR Rajasthan Chapter), Dr. Pratibha Singh (Organizing Chairpers) and Dr. Navdeep Kaur Guman (Organizing Secretary) and it comprised of various sessions and workshop that was very much appreciated by the Delegates. The conference was structured to foster discussion between participants around the core themes of the infertility. This was achieved by hosting carefully structured panel discussions. The high spot of the programme was Key-note address by Dr. Sudha Prasad on Red Flags in Infertility
STATE CHAPTER ACTIVITIES
treatment. The cherry on cake were ‘3rd Jodhpur Oration‘ presented to Dr. Abha Majumdar and ‘ISAR Rajasthan Chapter Oration’ presented to the Incoming President of ISAR - Dr. Prakash Trivedi. The conference also endorsed by some of eminent Faculty Dr. Ratna Agarwal – Vice President ISAR Embryology, Dr. Ruma Satwik, Dr. Sheetal Punjabi, Dr. Sangeeta Sharma, Dr. Tanu Batra. The best free paper prize was won by Dr. Ritika Gupta. RAJASTHAN CHAPTER INSTALLATION
On the occasion of ISAR Foundation Day, the CME was organized on February 15, 2020. The session was chaired by Dr. Indra Bhati and Prof. Dr. Ran-
ISAR Express February 2020
jana Desai and further it was followed by valuable and informative session on Advances in A.R.T. by Dr. Sanjay Makwana and Luteal Phase Defects by Dr. Leela Vyas. It was well attended by 40 gynecologists & doctors then followed by interactive session and program was ended at very positive note & felicitations! In the presence of Prof. Dr. Ranjana Desai - Superintendent Umed Hospital and Prof. Dr. Mahesh Bhati – Superintendent Mahatma Gandhi Hospital, the installation of new ISAR Rajasthan Executive Team was taken place. Following are the New Team Members that has taken the charge with effective from February 15, 2020. Chairperson – Dr. Leela Vyas, Jaipur Secretary - Dr. B. S. Jodha, Jodhpur Joint. Secretary - Dr. Suchika Mangal, Jaipur and Dr. Arshi Iqbal, Kota Vice Chairperson - Dr. Desh Deepak, Kota Executive - Dr. Alka Gehlot - Jaipur , Dr. Vikram S. Bhati (Embryologist)- Rawatsar, Dr. Tarun Agarwal - Udaipur, Dr. Kirti Gaur - Jodhpur, Dr. Renu makwana – Jodhpur, Dr. Kusum Meena - Jaipur Treasure - Dr. Rahul Sen - Jodhpur Dr. Sanjay Makwana Dr. Renu Makwana President, ISAR Rajasthan Chapter Secretary – ISAR Rajasthan Chapter.
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ISAR Express February 2020 EVENTS INDORE ISAR MASTERCLASS
ART PREGNANCIES - ARE THEY DIFFERENT?
ISAR Masterclass on ART Pregnancies: Are They Different, was organised on 21,22 December 2019. It was a well attended well appreciated topic . The scientific deliberations were excellent. Prof Dov Feldberg from Israel was the international faculty. Day 1: Ultrasound Workshop Speakers were Dr Mumtaz, Dr Wadekar, Dr Korbu, Dr Anita Kulkarni and Dr Archana Baser spoke on Twin Pregnancy. Day 2: There were good interactions and robust presentations from Dr Anita Soni, Dr Mridubhashini, Dr Mumtaz, Dr Prgya Mishra and Prof Dov Feldberg. Dr Asha Baxi Moderated an excellent Panel. Take home message from all talks was ‘YES’, ART pregnancy is ‘NOT’ just another pregnancy and need special attention and closer surveillance. Dr Asha Baxi was the organising chairperson and Dr Archana Baser was the organising secretary. Dr Dov Feldberg from Israel gave us valuable insights about patients with poor ovarian reserve. Dhaval Baxi talked on early pregnancy problems in women who conceive through IVF/ICSI. ISAR’s Indore workshop was a great combination of clinical excellence, team work and organisation.