Role of Preimplantation genetic testing in Indian women with advanced maternal age to optimize Reproductive Outcomes

Role of Preimplantation genetic testing in Indian women with advanced maternal age to optimize Reproductive Outcomes | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Help Center Sign In Submit a Preprint Cite Share Download PDF Article Role of Preimplantation genetic testing in Indian women with advanced maternal age to optimize Reproductive Outcomes Krishna chaitanya mantravadi, Durga Gedela Rao, Beena Rawat, Pooja Chauhan This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-2124932/v1 This work is licensed under a CC BY 4.0 License Status: Published Journal Publication published 07 Feb, 2024 Read the published version in Fertility & Reproduction → Version 1 posted You are reading this latest preprint version Abstract PURPOSE Does preimplantation genetic testing for aneuploidy (PGT-A) in embryos help women of advanced maternal age (AMA) achieve better reproductive outcomes? METHODS Multicenter, controlled retrospective study, a total of 267 patients (n = 267) were recruited, of which 53 patients (PGT-A group) consented to PGT-A, followed by euploid embryo transfer, whereas the remaining 214 patients (non-PGT-A group) underwent embryo transfer of un-screened morphologically graded blastocysts. RESULTS A significant increase in the clinical pregnancy rate was observed in the PGT-A group when compared to the non-PGT-A group (71.6% vs. 51%, p = 0.007), while the miscarriage rate was found to be lower in the PGT-A group compared to the non-PGT-A group (11% vs. 25%, p = 0.02). The live birth rates observed in either group were statistically nonsignificant (62.2% vs 51%, p = 0.14). In the PGT-A group, similarly, the implantation rate was found to be significantly higher than in the non-PGT-A group (53% vs. 33%, p = 0.007). CONCLUSION: The data suggest that PGT-A testing in women of advanced maternal age can improve their reproductive outcomes. Advanced Maternal Age PGT-A Blastocyst Live Birth Embryo Aneuploidy Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Introduction Women’s reproductive potential decreases significantly with advancing age ( 1 ), at the age of 35–37 a women’s cumulative pregnancy rate starts to drop and by the time she is 45 years of age, it is essentially zero ( 2 , 3 ). Many women today are delaying their motherhood into their late thirties due to their changing lifestyles and career aspirations. This emerging reproductive trend is posing a huge challenge for assisted reproduction centers as many women strongly believe that assisted reproduction techniques (ART) can truly reverse maternal aging ( 4 , 5 ). It is well established that advanced maternal age (AMA; defined as ≥ 35 years) is an important variable that greatly reduces the chances of having a healthy live birth even with in vitro fertilization (IVF) ( 6 ). Women who delay childbirth into their late thirties not only have a lower likelihood of having a live birth but also have impaired embryo development in vitro ( 7 ). The ≥ 35-year cut-off is set mainly due to high embryo aneuploidy rates which drastically increase from a baseline of 30–90% in women in their late 30s to 40s ( 8 , 9 ). Numerous studies have shown that infertility, miscarriages, and chromosomally abnormal pregnancies are more common among women of advanced maternal age ( 10 , 11 ). Errors in chromosomal segregation resulting in trisomic pregnancies occur much more frequently in women who are approaching the end of their childbearing years ( 12 ). The higher chances of embryo aneuploidy in AMA women may be due to dwindling ovarian reserve (OR) ( 13 ), cohesin dysfunction ( 14 ), reduced stringency of spindle assembly checkpoints ( 15 ), impaired mitochondrial metabolic activity ( 16 ), and shortening of telomeres ( 17 ). Initially, PGT-A was only performed to rule out hereditary genetic disorders & sex chromosome abnormalities in the suspected couples, so as to prevent the condition from passing down to the next generation. However, recently PGT-A is being done to identify euploid embryos for transfer even in couples with no known genetic disorders, to improve their pregnancy outcomes. Therefore, selecting and transferring a single chromosomally normal embryo can help in achieving higher pregnancy outcomes in a shorter duration with the increase in live birth rate, lower miscarriage rate & can limit multiple pregnancies. In the present controlled retrospective study, we intend to look for the efficacy of PGT-A in Indian women aged ≥ 35years undergoing ART cycles to optimize reproductive outcomes. Hypothesis- Can PGT-A help optimize Reproductive outcomes in indian women above the age of 35yrs undergoing ART cycles? Materials & Methods Study design It is a controlled retrospective study conducted in a private fertility center from the 2014 to 2020. In this study, 267 patients (AMA women) were included following the inclusion criteria. The PGT-A group (n = 53, study group), consisted of 53 patients who had undergone biopsy followed by euploid blastocyst transfer. Whereas the non-PGT-A group (n = 214, control group), comprised of 214 patients who had blastocyst transfer merely based on the morphological study of the embryo alone. Inclusion Criteria Women in the AMA group (≥ 35) were included in this study. Further, only frozen embryo transfer (FET) cycles were included. All the PGT-A cases where at least one or more euploid embryos were available for transfer were also included in this study. Informed consent was obtained from the patients to evaluate the data from the study. Exclusion Criteria All the PGT-A cases where no euploid embryos were available for transfer and with male factor were excluded. Stimulation & Patient Preparation All the patients were stimulated by following a down regulation stimulation protocol with GnRH antagonist (Gonal F, Merck Global, USA) for 10–12 days from D2/D3 of menstruation cycles to stimulate the ovaries to produce enough follicles. The growth of the Antral follicles was continuously monitored by Ultrasound guidance & blood estradiol (E2) levels. The dosage of the stimulation drug was individualized based on patient characteristics. Once the size of the follicles reaches 18–20 mm, a trigger injection was administrated (HCG 10,000, Serum Institute, India) and Oocyte retrieval was done 35–36 hours post trigger. All interventions performed to the women during the infertility treatments were in accordance with relevant guidelines and regulations. All couples that opted for PGT underwent counseling session and an informed written consent was obtained about the benefits, challenges and pitfalls of the embryos biopsy technique and the results of PGT. Only couples that gave informed consent were recruited in this retrospective study. Since the data of this study was retrospective in nature and a prior ethical clearance was not applied. Nevertheless we applied for a waiver of ethical clearance to the institutional review board and obtained a waiver for this study in order to publish our findings in women with advanced maternal age. Oocyte Collection, Insemination & Embryo Development The ovum pick up (OPU) was done post 35–36 h of HCG trigger under trans vaginal ultrasonography (TVS) with the help of suction pressure. The follicular fluid was screened under a stereozoom microscope in the IVF laboratory by the embryologist into a 60mm petri dishes. The cumulus oocyte complexes (COCs) were separated from the follicular fluid and cultured in a fertilization media (Quinn’s advantage protein plus, Cooper surgical inc., USA) for 2–3 hours. The COCs was then denuded with enzymatic (Hyaluronidase 80U/ml, Cooper surgical, USA) & mechanical process. All the metaphase (MII) oocytes were used for intra cytoplasmic sperm injection (ICSI). The semen sample were obtained from their partners by masturbation and analysed for count & motility. Double density gradient was performed as per the count & motility of the sperm. Morphologically normal sperms were used for injection during ICSI. All MII oocytes were injected and cultured in a Single step culture medium till Day5 (Sage 1 step with HSA, Origio, Denmark) at 37 degrees in a humidified incubator with hypoxic culture conditions (5% oxygen). Normal fertilization was confirmed by observing two distinct pronuclei & two polar bodies 16–18 hours post-ICSI. The fertilized zygotes were cultured till Day5/6 using standard incubation protocol. The embryo development assessment was done on Day5/6 and grading of the blastocyst was done as per the standard grading system (The Istanbul consensus workshop on embryo assessment). Biopsy, Tubing & Vitrification Only grade 1& 2 fully expanded blastocysts were selected for biopsies. Number of embryos to be biopsied and consent forms were obtained from the patients. LASER assisted hatching was performed to facilitate the herniation of enough number of trophectoderm (TE) cells and once the appropriate number of cells were herniated, the biopsy was performed by aspirating six to eight TE cells by using a biopsy needle (Blastomere aspiration needle, Cook, USA). Aspirated TE cells were then transferred into a PCR tube that contained Phosphate buffer saline (PBS) provided by the genetic lab (Igenomix, India) carefully in the presence of an eyewitness The biopsied cells were then stored in a deep freezer at -21 degrees Celsius until they were shipped to the genetic lab. The collapsed blastocysts were further cultured for 1–2 hours post biopsies for their expansion and then vitrified using a kitazato vitrification kit (Kitazato BioPharma, Tokyo, Japan) and stored on a cryotop in liquid nitrogen (LN2) Frozen Embryo Transfer (Fet) For subsequent FET, the endometrium was prepared by giving estrogen support. Once the endometrium attains adequate thickness (> 8mm), luteal phase support was started and, FET was planned. On the day of transfer, the thawing of one/two PGT-A euploid embryos was done by using the Kitazato Thawing kit (Kitazato BioPharma, Tokyo, Japan). The thawed blastocyst was cultured for 2 hours for survival confirmation in one step culture medium at 37 degrees Celsius. One or two thawed blastocyst was transferred into the uterus by using an ET catheter (Emtrac set, Gynetic, Belgium). Fourteen days after ET, the patient underwent a urine pregnancy test and pregnancy was confirmed by Beta Hcg levels in the blood (> 50mIU/ml). Statistics The variables were calculated and presented in absolute numbers and percentages. The chi-Square test was performed to analyze the difference between the variables. Differences were considered significant at P < 0.05. Result In this study, a total of 267 patients were included as per the inclusion criteria. Out of the 267 patients, 53 patients who had undergone biopsy followed by euploid blastocyst transfer were grouped as PGT-A (n = 53, Study group) and the rest of the patients where the blastocyst transfer was done entirely based on the morphological analysis of the embryo alone were grouped as Non-PGT-A (n = 214, Control group). The study group includes patients with AMA + PGT-A and the control group refers to patients with AMA + Non-PGT-A (Fig. 1 ). The clinical pregnancy rate was found to be considerably higher in the PGT-A group (71.6% vs 51%, p = 0.007) (Fig. 2 ) which indicates that transferring a PGT-A selected euploid embryo can increase the overall pregnancy outcome in the IVF cycle. Further, the miscarriage rate was found significantly lower in the PGT-A group as compared to the Non-PGT-A group (11% vs 25%, p = 0.02) (Fig. 3 ), which demonstrates that PGT-A may detect most of the chromosomal anomalies responsible for early miscarriages in the embryonic stage. There is no significant difference observed in terms of live birth rate in both groups (62.2% vs 51%, p = 0.14) (Fig. 4 ). The PGT-A group also had a significantly higher overall implantation rate (53% vs 33%, p = 0.007) (Fig. 5 ), which demonstrates that transferring a PGT-A selected euploid embryo increases its chances of implantation due to its normal genetic makeover. S.No. Variables Non-PGT-A PGT-A P-Value 1. No. of FET 214 53 3. Clinical Pregnancy Rate (CPR) 51% 71.6% 0.007 4. Miscarriage Rate (MR) 25.4% 11.3% 0.02 5. Implantation Rate (IR) 33% 53% 0.007 6. Live Birth Rate (LBR) 51% 62.2% 0.14 Table No.1: Comparison of different clinical outcomes between the Non- PGT-A and PGT-A groups. Discussion The primary objective of pre-implantation genetic testing for aneuploidy (PGT-A) in ART is to select the most competent embryo for transfer, after analyzing its genetic makeover or genotype, thus improving the reproductive outcome, as studies have shown that aneuploidy rates in IVF embryos are high, especially in AMA patients ( 9 ). Our study showed a significant increase in overall pregnancy rate (PR), implantation rate (IR), and lower miscarriage rate (MR) in AMA women who had undergone PGT-A as when compared to AMA women without PGT-A (Table 1). There was not much difference observed in the live birth rate in both groups, even though the PGT-A group had slightly increased live birth rates as compared to Non PGT-A group (62.2% vs 51%, p < 0.14), but this was statistically not significant. A lot of studies have emphasized the use of PGT-A in the AMA group of patients and elaborated on the benefits of selecting a euploid embryo for transfer. One such study by Hsiao-Ling Lee and his team, has demonstrated that blastocyst biopsy with array comparative genomic hybridization (aCGH) is a reliable method for detecting euploid embryos for transfer and supported the use of PGT-A in AMA patients ( 18 ). In Another study, it was concluded that testing embryos for common chromosomal aberrations using PGT-A resulted in fewer embryo transfers, lower miscarriages, and increased pregnancy rates in AMA patients ( 19 ). In his recent study, Ubaldi et al. have emphasized the significance of TE biopsy, vitrification, and CCS for single embryo transfer in AMA patients ( 20 , 21 ). Recurrent miscarriage (RM) is a multifactorial disorder defined by two or more losses. Hodes-Wertz et al. found that idiopathic RM is mostly caused by aneuploid embryos and that PGT-A with aCGH could decrease MR and improve PR ( 22 ). IVF/PGT-A appears to lower the miscarriage risk when compared with natural conception. However, there are few studies comparing the live birth rate in AMA patients followed by TE biopsy & euploid embryo transfer. In this regard, Lee et al. have supported that the application of TE biopsy and PGTA could improve the live birth rate in women aged 40–43 years (18). The ultimate goal of ART treatment is to achieve a healthy live birth. To accomplish this goal, we need a tool to select the euploid embryos from the cohort of same to minimize the risk of transferring aneuploid embryos. PGT-A can be a helpful tool for selecting chromosomally competent embryos for transfer in the ART cycle especially in the AMA group of patients where the rate of chromosomal anomalies is higher. Also, transferring one or two PGT-A-selected euploid embryos can lower the multiple gestational pregnancies and can avoid the health risk associated with it to the mother (23). The use of PGT-A in AMA patients can also reduce the number of IVF cycles to achieve a healthy live birth and the time taken for conceiving as well. However, PGT-A has its own limitations such as, it is an invasive technique as it requires TE biopsies to be performed on day5 blastocysts, also the cost of testing each blastocyst is expensive. Moreover, many AMA women have a poor ovarian reserve, which limits the number of blastocysts available for testing, thereby further reducing the chances of having at least one euploid embryo for transfer from the cohort. In such cases, the patient may end up with no blastocyst for transfer in the IVF cycle or needs to go for another cycle. Due to these factors, the use of PGT-A in the ART cycle on a routine basis is debatable. The assessment of necessity and counseling of patients is of utmost priority. Conclusion We found PGT-A to be useful in women of advanced maternal age, where transferring a single euploid embryo revealed better reproductive outcomes when compared to, transferring a blastocyst based on its morphological characteristic alone. As mentioned, PGT-A has its own limitations but at present, it is the only efficient strategy available to minimize the age-related reproductive risk in AMA women. Therefore, a greater number of studies with larger sample size is required to support the live birth data. Declarations Acknowledgment The authors would like to thank the staff and laboratory personnel at Oasis Fertility for their generous support and assistance throughout this study. Authors would also want to thank all the health workers who were involved in the care for the patients. Funding This research did not receive any specific grant from funding agencies in the public, commercial or not-for-profit sectors. Disclosure The authors have no relevant conflicts of interest to declare. Interventions performed as a part of this retrospective study have relevant informed patient consents duly signed. Author’s contribution Krishna Mantravadi and Durga G Rao conceived the study. Pooja Chauhan collected and analyzed the data. Beena Rawat helped preparing the manuscript and statistics. All authors were involved in writing the manuscript. References Devesa M, Tur R, Rodríguez I, Coroleu B, Martínez F, Polyzos NP. Cumulative live birth rates and number of oocytes retrieved in women of advanced age. A single centre analysis including 4500 women ≥ 38 years old. Hum Reprod. 2018;33(11):2010–7. Cetinkaya MB, Siano LJ, Benadiva C, Sakkas D, Patrizio P. Reproductive outcome of women 43 years and beyond undergoing ART treatment with their own oocytes in two Connecticut university programs. J Assist Reprod Genet. 2013;30(5):673–8. Tsafrir A, Simon A, Revel A, Reubinoff B, Lewin A, Laufer N. Retrospective analysis of 1217 IVF cycles in women aged 40 years and older. Reprod Biomed Online. 2007;14(3):348–55. Leridon H. Can assisted reproduction technology compensate for the natural decline in fertility with age? A model assessment. Hum Reprod. 2004;19(7):1548–53. Mills M, Rindfuss RR, McDonald P, te Velde E. Why do people postpone parenthood? Reasons and social policy incentives. Hum Reprod Update. 2011;17(6):848–60. Cimadomo D, Fabozzi G, Vaiarelli A, Ubaldi N, Ubaldi FM, Rienzi L. Impact of Maternal Age on Oocyte and Embryo Competence. Front Endocrinol (Lausanne). 2018;9:327. Janny L, Menezo YJ. Maternal age effect on early human embryonic development and blastocyst formation. Mol Reprod Dev. 1996;45(1):31–7. Capalbo A, Hoffmann ER, Cimadomo D, Ubaldi FM, Rienzi L. Human female meiosis revised: new insights into the mechanisms of chromosome segregation and aneuploidies from advanced genomics and time-lapse imaging. Hum Reprod Update. 2017;23(6):706–22. Franasiak JM, Forman EJ, Hong KH, Werner MD, Upham KM, Treff NR, et al. The nature of aneuploidy with increasing age of the female partner: a review of 15,169 consecutive trophectoderm biopsies evaluated with comprehensive chromosomal screening. Fertil Steril. 2014;101(3):656 – 63.e1. Magnus MC, Wilcox AJ, Morken NH, Weinberg CR, Håberg SE. Role of maternal age and pregnancy history in risk of miscarriage: prospective register based study. Bmj. 2019;364:l869. Nybo Andersen AM, Wohlfahrt J, Christens P, Olsen J, Melbye M. Maternal age and fetal loss: population based register linkage study. Bmj. 2000;320(7251):1708–12. Grande M, Borrell A, Garcia-Posada R, Borobio V, Muñoz M, Creus M, et al. The effect of maternal age on chromosomal anomaly rate and spectrum in recurrent miscarriage. Hum Reprod. 2012;27(10):3109–17. Faddy MJ, Gosden RG, Gougeon A, Richardson SJ, Nelson JF. Accelerated disappearance of ovarian follicles in mid-life: implications for forecasting menopause. Hum Reprod. 1992;7(10):1342–6. Cheng JM, Liu YX. Age-Related Loss of Cohesion: Causes and Effects. Int J Mol Sci. 2017;18(7). Steuerwald N, Cohen J, Herrera RJ, Sandalinas M, Brenner CA. Association between spindle assembly checkpoint expression and maternal age in human oocytes. Mol Hum Reprod. 2001;7(1):49–55. Van Blerkom J. Mitochondrial function in the human oocyte and embryo and their role in developmental competence. Mitochondrion. 2011;11(5):797–813. de Lange T. How telomeres solve the end-protection problem. Science. 2009;326(5955):948–52. Lee HL, McCulloh DH, Hodes-Wertz B, Adler A, McCaffrey C, Grifo JA. In vitro fertilization with preimplantation genetic screening improves implantation and live birth in women age 40 through 43. J Assist Reprod Genet. 2015;32(3):435–44. Sacchi L, Albani E, Cesana A, Smeraldi A, Parini V, Fabiani M, et al. Preimplantation Genetic Testing for Aneuploidy Improves Clinical, Gestational, and Neonatal Outcomes in Advanced Maternal Age Patients Without Compromising Cumulative Live-Birth Rate. J Assist Reprod Genet. 2019;36(12):2493–504. Rubio C, Bellver J, Rodrigo L, Castillón G, Guillén A, Vidal C, et al. In vitro fertilization with preimplantation genetic diagnosis for aneuploidies in advanced maternal age: a randomized, controlled study. Fertil Steril. 2017;107(5):1122–9. Ubaldi FM, Cimadomo D, Vaiarelli A, Fabozzi G, Venturella R, Maggiulli R, et al. Advanced Maternal Age in IVF: Still a Challenge? The Present and the Future of Its Treatment. Front Endocrinol (Lausanne). 2019;10:94. Hodes-Wertz B, Grifo J, Ghadir S, Kaplan B, Laskin CA, Glassner M, et al. Idiopathic recurrent miscarriage is caused mostly by aneuploid embryos. Fertil Steril. 2012;98(3):675–80. Meldrum DR. Introduction: Preimplantation genetic screening is alive and very well. Fertil Steril. 2013;100(3):593–4. Additional Declarations No competing interests reported. Cite Share Download PDF Status: Published Journal Publication published 07 Feb, 2024 Read the published version in Fertility & Reproduction → Version 1 posted You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. Also discoverable on Platform About Our Team In Review Editorial Policies Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-2124932","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Article","associatedPublications":[],"authors":[{"id":142336317,"identity":"44dd6574-41e9-49b0-b05a-3cb025f7f52e","order_by":0,"name":"Krishna chaitanya mantravadi","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAA6klEQVRIiWNgGAWjYBACCQbGBmYgLQfiHGAwYGBsAzOI0GJMihYGBpCWxAaoAGMDTrVQIDn7cPPnwh126f3sxx8e/FFwR7aP/QDj4QI8WqT5EtukZ55Jzp3Zk2NwmMfgmXEbTwLD4Rl4tMjxMLYx87Yx5244kMNwmMHgcGIbA1ALD34tzZ952+rTDc4/f3DwB0gL/wP8WqR5GBukedsOJxjcSDA4wAPSIkHAFskexjagluOGM2e8AfnlsHGbxMMGvFokzrA/BjqsWp6fP/3xxx9/DsvO708+/BmfFmyAcNSMglEwCkbBKCAAALvvTv6fkEy9AAAAAElFTkSuQmCC","orcid":"","institution":"Oasis Fertility, India","correspondingAuthor":true,"submittingAuthor":false,"prefix":"","firstName":"Krishna","middleName":"chaitanya","lastName":"mantravadi","suffix":""},{"id":142336318,"identity":"c20f99c1-0692-43ad-80e7-99ce2aee23c3","order_by":1,"name":"Durga Gedela Rao","email":"","orcid":"","institution":"Oasis Fertility, India","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Durga","middleName":"Gedela","lastName":"Rao","suffix":""},{"id":142336319,"identity":"423cdc95-ba15-4a8e-bffd-ad7b1c542e8d","order_by":2,"name":"Beena Rawat","email":"","orcid":"","institution":"Oasis Fertility, India","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Beena","middleName":"","lastName":"Rawat","suffix":""},{"id":142336320,"identity":"a52e49d3-6731-4d5f-a979-9e5ccdb85553","order_by":3,"name":"Pooja Chauhan","email":"","orcid":"","institution":"Oasis Fertility, India","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Pooja","middleName":"","lastName":"Chauhan","suffix":""}],"badges":[],"createdAt":"2022-10-02 06:44:18","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-2124932/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-2124932/v1","draftVersion":[],"editorialEvents":[{"content":"https://doi.org/10.1142/S2661318224500014","type":"published","date":"2024-02-07T09:51:51+00:00"}],"editorialNote":"","failedWorkflow":false,"files":[{"id":27485861,"identity":"a4804b29-bdbe-45fe-a72e-00ed74716589","added_by":"auto","created_at":"2022-10-07 20:20:13","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":15940,"visible":true,"origin":"","legend":"\u003cp\u003eFlow chart representing the patient recruitment into the groups.\u003c/p\u003e","description":"","filename":"1.png","url":"https://assets-eu.researchsquare.com/files/rs-2124932/v1/3fe6346320b363ed55942056.png"},{"id":27485860,"identity":"1cb1b0f8-2049-4549-9a6d-1667d72b7a93","added_by":"auto","created_at":"2022-10-07 20:20:13","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":31479,"visible":true,"origin":"","legend":"\u003cp\u003eComparison of Clinical Pregnancy Rate (CPR) between the control group \u0026amp; test group patients. The clinical pregnancy rate between the two groups was statistically significant (P = 0.007).\u003c/p\u003e","description":"","filename":"2.png","url":"https://assets-eu.researchsquare.com/files/rs-2124932/v1/8083a401d826687ec772dde0.png"},{"id":27485858,"identity":"92324d82-f535-407e-a7f0-ac0534325767","added_by":"auto","created_at":"2022-10-07 20:20:13","extension":"png","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":20499,"visible":true,"origin":"","legend":"\u003cp\u003eComparison of miscarriage rate (MR) between the control group \u0026amp; test group patients. The Miscarriage rate between the two groups showed statistical significance (P = 0.02).\u003c/p\u003e","description":"","filename":"3.png","url":"https://assets-eu.researchsquare.com/files/rs-2124932/v1/06f259e4c2a1d543fd7b4bd6.png"},{"id":27485936,"identity":"e1e926cb-8161-4626-a6d4-2243b8ae52e7","added_by":"auto","created_at":"2022-10-07 20:25:13","extension":"png","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":16726,"visible":true,"origin":"","legend":"\u003cp\u003eComparison of Live Birth Rate (LBR) between control group \u0026amp; test group patients. The live birth rate between the two groups was not statistically significant (P = 0.14).\u003c/p\u003e","description":"","filename":"4.png","url":"https://assets-eu.researchsquare.com/files/rs-2124932/v1/5eaec79cf6a3f134609a5423.png"},{"id":27485862,"identity":"c9467b4f-80fa-4f02-9b0f-51f810c1e073","added_by":"auto","created_at":"2022-10-07 20:20:13","extension":"png","order_by":5,"title":"Figure 5","display":"","copyAsset":false,"role":"figure","size":20817,"visible":true,"origin":"","legend":"\u003cp\u003eComparison of Implantation rate (IR) between the control group \u0026amp; test group patients. The implantation rate between the two groups was statistically significant (P = 0.007).\u003c/p\u003e","description":"","filename":"5.png","url":"https://assets-eu.researchsquare.com/files/rs-2124932/v1/559e1bc5e36b50de6aaf7a55.png"},{"id":53380099,"identity":"113b2610-5e18-40e1-966f-6dc1663e744b","added_by":"auto","created_at":"2024-03-25 09:51:56","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":356136,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-2124932/v1/70ed99c6-a646-42b4-8faf-c0543c9a0015.pdf"}],"financialInterests":"No competing interests reported.","formattedTitle":"Role of Preimplantation genetic testing in Indian women with advanced maternal age to optimize Reproductive Outcomes","fulltext":[{"header":"Introduction","content":"\u003cp\u003eWomen\u0026rsquo;s reproductive potential decreases significantly with advancing age (\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e), at the age of 35\u0026ndash;37 a women\u0026rsquo;s cumulative pregnancy rate starts to drop and by the time she is 45 years of age, it is essentially zero (\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e, \u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e). Many women today are delaying their motherhood into their late thirties due to their changing lifestyles and career aspirations. This emerging reproductive trend is posing a huge challenge for assisted reproduction centers as many women strongly believe that assisted reproduction techniques (ART) can truly reverse maternal aging (\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e, \u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e). It is well established that advanced maternal age (AMA; defined as \u0026ge;\u0026thinsp;35 years) is an important variable that greatly reduces the chances of having a healthy live birth even with in vitro fertilization (IVF) (\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e). Women who delay childbirth into their late thirties not only have a lower likelihood of having a live birth but also have impaired embryo development in vitro (\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e). The \u0026ge;\u0026thinsp;35-year cut-off is set mainly due to high embryo aneuploidy rates which drastically increase from a baseline of 30\u0026ndash;90% in women in their late 30s to 40s (\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e, \u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eNumerous studies have shown that infertility, miscarriages, and chromosomally abnormal pregnancies are more common among women of advanced maternal age (\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e, \u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e). Errors in chromosomal segregation resulting in trisomic pregnancies occur much more frequently in women who are approaching the end of their childbearing years (\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e). The higher chances of embryo aneuploidy in AMA women may be due to dwindling ovarian reserve (OR) (\u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e), cohesin dysfunction (\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e), reduced stringency of spindle assembly checkpoints (\u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e), impaired mitochondrial metabolic activity (\u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e), and shortening of telomeres (\u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eInitially, PGT-A was only performed to rule out hereditary genetic disorders \u0026amp; sex chromosome abnormalities in the suspected couples, so as to prevent the condition from passing down to the next generation. However, recently PGT-A is being done to identify euploid embryos for transfer even in couples with no known genetic disorders, to improve their pregnancy outcomes.\u003c/p\u003e \u003cp\u003eTherefore, selecting and transferring a single chromosomally normal embryo can help in achieving higher pregnancy outcomes in a shorter duration with the increase in live birth rate, lower miscarriage rate \u0026amp; can limit multiple pregnancies.\u003c/p\u003e \u003cp\u003eIn the present controlled retrospective study, we intend to look for the efficacy of PGT-A in Indian women aged\u0026thinsp;\u0026ge;\u0026thinsp;35years undergoing ART cycles to optimize reproductive outcomes.\u003c/p\u003e \u003cp\u003eHypothesis- Can PGT-A help optimize Reproductive outcomes in indian women above the age of 35yrs undergoing ART cycles?\u003c/p\u003e"},{"header":"Materials \u0026 Methods","content":"\u003cdiv id=\"Sec3\" class=\"Section2\"\u003e \u003ch2\u003eStudy design\u003c/h2\u003e \u003cp\u003eIt is a controlled retrospective study conducted in a private fertility center from the 2014 to 2020. In this study, 267 patients (AMA women) were included following the inclusion criteria. The PGT-A group (n\u0026thinsp;=\u0026thinsp;53, study group), consisted of 53 patients who had undergone biopsy followed by euploid blastocyst transfer. Whereas the non-PGT-A group (n\u0026thinsp;=\u0026thinsp;214, control group), comprised of 214 patients who had blastocyst transfer merely based on the morphological study of the embryo alone.\u003c/p\u003e \u003c/div\u003e\n\u003ch3\u003eInclusion Criteria\u003c/h3\u003e\n\u003cp\u003eWomen in the AMA group (\u0026ge;\u0026thinsp;35) were included in this study. Further, only frozen embryo transfer (FET) cycles were included. All the PGT-A cases where at least one or more euploid embryos were available for transfer were also included in this study. Informed consent was obtained from the patients to evaluate the data from the study.\u003c/p\u003e\n\u003ch3\u003eExclusion Criteria\u003c/h3\u003e\n\u003cp\u003eAll the PGT-A cases where no euploid embryos were available for transfer and with male factor were excluded.\u003c/p\u003e\n\u003ch3\u003eStimulation \u0026 Patient Preparation\u003c/h3\u003e\n\u003cp\u003eAll the patients were stimulated by following a down regulation stimulation protocol with GnRH antagonist (Gonal F, Merck Global, USA) for 10\u0026ndash;12 days from D2/D3 of menstruation cycles to stimulate the ovaries to produce enough follicles. The growth of the Antral follicles was continuously monitored by Ultrasound guidance \u0026amp; blood estradiol (E2) levels. The dosage of the stimulation drug was individualized based on patient characteristics. Once the size of the follicles reaches 18\u0026ndash;20 mm, a trigger injection was administrated (HCG 10,000, Serum Institute, India) and Oocyte retrieval was done 35\u0026ndash;36 hours post trigger.\u003c/p\u003e \u003cp\u003e All interventions performed to the women during the infertility treatments were in accordance with relevant guidelines and regulations. All couples that opted for PGT underwent counseling session and an informed written consent was obtained about the benefits, challenges and pitfalls of the embryos biopsy technique and the results of PGT. Only couples that gave informed consent were recruited in this retrospective study. Since the data of this study was retrospective in nature and a prior ethical clearance was not applied. Nevertheless we applied for a waiver of ethical clearance to the institutional review board and obtained a waiver for this study in order to publish our findings in women with advanced maternal age.\u003c/p\u003e\n\u003ch3\u003eOocyte Collection, Insemination \u0026 Embryo Development\u003c/h3\u003e\n\u003cp\u003eThe ovum pick up (OPU) was done post 35\u0026ndash;36 h of HCG trigger under trans vaginal ultrasonography (TVS) with the help of suction pressure. The follicular fluid was screened under a stereozoom microscope in the IVF laboratory by the embryologist into a 60mm petri dishes. The cumulus oocyte complexes (COCs) were separated from the follicular fluid and cultured in a fertilization media (Quinn\u0026rsquo;s advantage protein plus, Cooper surgical inc., USA) for 2\u0026ndash;3 hours. The COCs was then denuded with enzymatic (Hyaluronidase 80U/ml, Cooper surgical, USA) \u0026amp; mechanical process. All the metaphase (MII) oocytes were used for intra cytoplasmic sperm injection (ICSI). The semen sample were obtained from their partners by masturbation and analysed for count \u0026amp; motility. Double density gradient was performed as per the count \u0026amp; motility of the sperm. Morphologically normal sperms were used for injection during ICSI. All MII oocytes were injected and cultured in a Single step culture medium till Day5 (Sage 1 step with HSA, Origio, Denmark) at 37 degrees in a humidified incubator with hypoxic culture conditions (5% oxygen). Normal fertilization was confirmed by observing two distinct pronuclei \u0026amp; two polar bodies 16\u0026ndash;18 hours post-ICSI. The fertilized zygotes were cultured till Day5/6 using standard incubation protocol. The embryo development assessment was done on Day5/6 and grading of the blastocyst was done as per the standard grading system (The Istanbul consensus workshop on embryo assessment).\u003c/p\u003e\n\u003ch3\u003eBiopsy, Tubing \u0026 Vitrification\u003c/h3\u003e\n\u003cp\u003eOnly grade 1\u0026amp; 2 fully expanded blastocysts were selected for biopsies. Number of embryos to be biopsied and consent forms were obtained from the patients. LASER assisted hatching was performed to facilitate the herniation of enough number of trophectoderm (TE) cells and once the appropriate number of cells were herniated, the biopsy was performed by aspirating six to eight TE cells by using a biopsy needle (Blastomere aspiration needle, Cook, USA). Aspirated TE cells were then transferred into a PCR tube that contained Phosphate buffer saline (PBS) provided by the genetic lab (Igenomix, India) carefully in the presence of an eyewitness The biopsied cells were then stored in a deep freezer at -21 degrees Celsius until they were shipped to the genetic lab.\u003c/p\u003e \u003cp\u003eThe collapsed blastocysts were further cultured for 1\u0026ndash;2 hours post biopsies for their expansion and then vitrified using a kitazato vitrification kit (Kitazato BioPharma, Tokyo, Japan) and stored on a cryotop in liquid nitrogen (LN2)\u003c/p\u003e\n\u003ch3\u003eFrozen Embryo Transfer (Fet)\u003c/h3\u003e\n\u003cp\u003eFor subsequent FET, the endometrium was prepared by giving estrogen support. Once the endometrium attains adequate thickness (\u0026gt;\u0026thinsp;8mm), luteal phase support was started and, FET was planned. On the day of transfer, the thawing of one/two PGT-A euploid embryos was done by using the Kitazato Thawing kit (Kitazato BioPharma, Tokyo, Japan). The thawed blastocyst was cultured for 2 hours for survival confirmation in one step culture medium at 37 degrees Celsius. One or two thawed blastocyst was transferred into the uterus by using an ET catheter (Emtrac set, Gynetic, Belgium). Fourteen days after ET, the patient underwent a urine pregnancy test and pregnancy was confirmed by Beta Hcg levels in the blood (\u0026gt;\u0026thinsp;50mIU/ml).\u003c/p\u003e\n\u003ch3\u003eStatistics\u003c/h3\u003e\n\u003cp\u003eThe variables were calculated and presented in absolute numbers and percentages. The chi-Square test was performed to analyze the difference between the variables. Differences were considered significant at P\u0026thinsp;\u0026lt;\u0026thinsp;0.05.\u003c/p\u003e"},{"header":"Result","content":"\u003cp\u003eIn this study, a total of 267 patients were included as per the inclusion criteria. Out of the 267 patients, 53 patients who had undergone biopsy followed by euploid blastocyst transfer were grouped as PGT-A (n\u0026thinsp;=\u0026thinsp;53, Study group) and the rest of the patients where the blastocyst transfer was done entirely based on the morphological analysis of the embryo alone were grouped as Non-PGT-A (n\u0026thinsp;=\u0026thinsp;214, Control group). The study group includes patients with AMA\u0026thinsp;+\u0026thinsp;PGT-A and the control group refers to patients with AMA\u0026thinsp;+\u0026thinsp;Non-PGT-A (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eThe clinical pregnancy rate was found to be considerably higher in the PGT-A group (71.6% vs 51%, p\u0026thinsp;=\u0026thinsp;0.007) (Fig.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e2\u003c/span\u003e) which indicates that transferring a PGT-A selected euploid embryo can increase the overall pregnancy outcome in the IVF cycle. Further, the miscarriage rate was found significantly lower in the PGT-A group as compared to the Non-PGT-A group (11% vs 25%, p\u0026thinsp;=\u0026thinsp;0.02) (Fig.\u0026nbsp;\u003cspan refid=\"Fig7\" class=\"InternalRef\"\u003e3\u003c/span\u003e), which demonstrates that PGT-A may detect most of the chromosomal anomalies responsible for early miscarriages in the embryonic stage. There is no significant difference observed in terms of live birth rate in both groups (62.2% vs 51%, p\u0026thinsp;=\u0026thinsp;0.14) (Fig.\u0026nbsp;\u003cspan refid=\"Fig8\" class=\"InternalRef\"\u003e4\u003c/span\u003e). The PGT-A group also had a significantly higher overall implantation rate (53% vs 33%, p\u0026thinsp;=\u0026thinsp;0.007) (Fig.\u0026nbsp;\u003cspan refid=\"Fig9\" class=\"InternalRef\"\u003e5\u003c/span\u003e), which demonstrates that transferring a PGT-A selected euploid embryo increases its chances of implantation due to its normal genetic makeover.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"No\" id=\"Taba\" border=\"1\"\u003e \u003ccolgroup cols=\"5\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003eS.No.\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eVariables\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eNon-PGT-A\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003ePGT-A\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003eP-Value\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e1.\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eNo. of FET\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e214\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e53\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e3.\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eClinical Pregnancy Rate (CPR)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e51%\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e71.6%\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e0.007\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e4.\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eMiscarriage Rate (MR)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e25.4%\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e11.3%\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e0.02\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e5.\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eImplantation Rate (IR)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e33%\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e53%\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e0.007\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e6.\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eLive Birth Rate (LBR)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e51%\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e62.2%\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e0.14\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eTable No.1: Comparison of different clinical outcomes between the Non- PGT-A and PGT-A groups.\u003c/p\u003e"},{"header":"Discussion","content":"\u003cp\u003eThe primary objective of pre-implantation genetic testing for aneuploidy (PGT-A) in ART is to select the most competent embryo for transfer, after analyzing its genetic makeover or genotype, thus improving the reproductive outcome, as studies have shown that aneuploidy rates in IVF embryos are high, especially in AMA patients (\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eOur study showed a significant increase in overall pregnancy rate (PR), implantation rate (IR), and lower miscarriage rate (MR) in AMA women who had undergone PGT-A as when compared to AMA women without PGT-A (Table\u0026nbsp;1). There was not much difference observed in the live birth rate in both groups, even though the PGT-A group had slightly increased live birth rates as compared to Non PGT-A group (62.2% vs 51%, p\u0026thinsp;\u0026lt;\u0026thinsp;0.14), but this was statistically not significant.\u003c/p\u003e \u003cp\u003eA lot of studies have emphasized the use of PGT-A in the AMA group of patients and elaborated on the benefits of selecting a euploid embryo for transfer. One such study by Hsiao-Ling Lee and his team, has demonstrated that blastocyst biopsy with array comparative genomic hybridization (aCGH) is a reliable method for detecting euploid embryos for transfer and supported the use of PGT-A in AMA patients (\u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e18\u003c/span\u003e). In Another study, it was concluded that testing embryos for common chromosomal aberrations using PGT-A resulted in fewer embryo transfers, lower miscarriages, and increased pregnancy rates in AMA patients (\u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e). In his recent study, Ubaldi et al. have emphasized the significance of TE biopsy, vitrification, and CCS for single embryo transfer in AMA patients (\u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e20\u003c/span\u003e, \u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e21\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eRecurrent miscarriage (RM) is a multifactorial disorder defined by two or more losses. Hodes-Wertz et al. found that idiopathic RM is mostly caused by aneuploid embryos and that PGT-A with aCGH could decrease MR and improve PR (\u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e22\u003c/span\u003e). IVF/PGT-A appears to lower the miscarriage risk when compared with natural conception.\u003c/p\u003e \u003cp\u003eHowever, there are few studies comparing the live birth rate in AMA patients followed by TE biopsy \u0026amp; euploid embryo transfer. In this regard, Lee et al. have supported that the application of TE biopsy and PGTA could improve the live birth rate in women aged 40\u0026ndash;43 years (18). The ultimate goal of ART treatment is to achieve a healthy live birth. To accomplish this goal, we need a tool to select the euploid embryos from the cohort of same to minimize the risk of transferring aneuploid embryos. PGT-A can be a helpful tool for selecting chromosomally competent embryos for transfer in the ART cycle especially in the AMA group of patients where the rate of chromosomal anomalies is higher. Also, transferring one or two PGT-A-selected euploid embryos can lower the multiple gestational pregnancies and can avoid the health risk associated with it to the mother (23). The use of PGT-A in AMA patients can also reduce the number of IVF cycles to achieve a healthy live birth and the time taken for conceiving as well.\u003c/p\u003e \u003cp\u003eHowever, PGT-A has its own limitations such as, it is an invasive technique as it requires TE biopsies to be performed on day5 blastocysts, also the cost of testing each blastocyst is expensive.\u003c/p\u003e \u003cp\u003eMoreover, many AMA women have a poor ovarian reserve, which limits the number of blastocysts available for testing, thereby further reducing the chances of having at least one euploid embryo for transfer from the cohort. In such cases, the patient may end up with no blastocyst for transfer in the IVF cycle or needs to go for another cycle. Due to these factors, the use of PGT-A in the ART cycle on a routine basis is debatable. The assessment of necessity and counseling of patients is of utmost priority.\u003c/p\u003e"},{"header":"Conclusion","content":"\u003cp\u003eWe found PGT-A to be useful in women of advanced maternal age, where transferring a single euploid embryo revealed better reproductive outcomes when compared to, transferring a blastocyst based on its morphological characteristic alone. As mentioned, PGT-A has its own limitations but at present, it is the only efficient strategy available to minimize the age-related reproductive risk in AMA women. Therefore, a greater number of studies with larger sample size is required to support the live birth data.\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eAcknowledgment\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors would like to thank the staff and laboratory personnel at Oasis Fertility for their generous support and assistance throughout this study. Authors would also want to thank all the health workers who were involved in the care for the patients.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eFunding\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThis research did not receive any specific grant from funding agencies in the public, commercial or not-for-profit sectors.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eDisclosure\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors have no relevant conflicts of interest to declare. Interventions performed as a part of this retrospective study have relevant informed patient consents duly signed.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAuthor\u0026rsquo;s contribution\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eKrishna Mantravadi and Durga G Rao conceived the study. Pooja Chauhan collected and analyzed the data. Beena Rawat helped preparing the manuscript and statistics. All authors were involved in writing the manuscript. \u0026nbsp;\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\u003cli\u003e\u003cspan\u003eDevesa M, Tur R, Rodr\u0026iacute;guez I, Coroleu B, Mart\u0026iacute;nez F, Polyzos NP. Cumulative live birth rates and number of oocytes retrieved in women of advanced age. A single centre analysis including 4500 women \u0026ge; 38 years old. Hum Reprod. 2018;33(11):2010\u0026ndash;7.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eCetinkaya MB, Siano LJ, Benadiva C, Sakkas D, Patrizio P. Reproductive outcome of women 43 years and beyond undergoing ART treatment with their own oocytes in two Connecticut university programs. J Assist Reprod Genet. 2013;30(5):673\u0026ndash;8.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eTsafrir A, Simon A, Revel A, Reubinoff B, Lewin A, Laufer N. Retrospective analysis of 1217 IVF cycles in women aged 40 years and older. Reprod Biomed Online. 2007;14(3):348\u0026ndash;55.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eLeridon H. Can assisted reproduction technology compensate for the natural decline in fertility with age? A model assessment. Hum Reprod. 2004;19(7):1548\u0026ndash;53.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eMills M, Rindfuss RR, McDonald P, te Velde E. Why do people postpone parenthood? Reasons and social policy incentives. Hum Reprod Update. 2011;17(6):848\u0026ndash;60.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eCimadomo D, Fabozzi G, Vaiarelli A, Ubaldi N, Ubaldi FM, Rienzi L. Impact of Maternal Age on Oocyte and Embryo Competence. Front Endocrinol (Lausanne). 2018;9:327.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eJanny L, Menezo YJ. Maternal age effect on early human embryonic development and blastocyst formation. Mol Reprod Dev. 1996;45(1):31\u0026ndash;7.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eCapalbo A, Hoffmann ER, Cimadomo D, Ubaldi FM, Rienzi L. Human female meiosis revised: new insights into the mechanisms of chromosome segregation and aneuploidies from advanced genomics and time-lapse imaging. Hum Reprod Update. 2017;23(6):706\u0026ndash;22.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eFranasiak JM, Forman EJ, Hong KH, Werner MD, Upham KM, Treff NR, et al. The nature of aneuploidy with increasing age of the female partner: a review of 15,169 consecutive trophectoderm biopsies evaluated with comprehensive chromosomal screening. Fertil Steril. 2014;101(3):656 \u0026ndash; 63.e1.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eMagnus MC, Wilcox AJ, Morken NH, Weinberg CR, H\u0026aring;berg SE. Role of maternal age and pregnancy history in risk of miscarriage: prospective register based study. Bmj. 2019;364:l869.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eNybo Andersen AM, Wohlfahrt J, Christens P, Olsen J, Melbye M. Maternal age and fetal loss: population based register linkage study. Bmj. 2000;320(7251):1708\u0026ndash;12.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eGrande M, Borrell A, Garcia-Posada R, Borobio V, Mu\u0026ntilde;oz M, Creus M, et al. The effect of maternal age on chromosomal anomaly rate and spectrum in recurrent miscarriage. Hum Reprod. 2012;27(10):3109\u0026ndash;17.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eFaddy MJ, Gosden RG, Gougeon A, Richardson SJ, Nelson JF. Accelerated disappearance of ovarian follicles in mid-life: implications for forecasting menopause. Hum Reprod. 1992;7(10):1342\u0026ndash;6.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eCheng JM, Liu YX. Age-Related Loss of Cohesion: Causes and Effects. Int J Mol Sci. 2017;18(7).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eSteuerwald N, Cohen J, Herrera RJ, Sandalinas M, Brenner CA. Association between spindle assembly checkpoint expression and maternal age in human oocytes. Mol Hum Reprod. 2001;7(1):49\u0026ndash;55.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eVan Blerkom J. Mitochondrial function in the human oocyte and embryo and their role in developmental competence. Mitochondrion. 2011;11(5):797\u0026ndash;813.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003ede Lange T. How telomeres solve the end-protection problem. Science. 2009;326(5955):948\u0026ndash;52.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eLee HL, McCulloh DH, Hodes-Wertz B, Adler A, McCaffrey C, Grifo JA. In vitro fertilization with preimplantation genetic screening improves implantation and live birth in women age 40 through 43. J Assist Reprod Genet. 2015;32(3):435\u0026ndash;44.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eSacchi L, Albani E, Cesana A, Smeraldi A, Parini V, Fabiani M, et al. Preimplantation Genetic Testing for Aneuploidy Improves Clinical, Gestational, and Neonatal Outcomes in Advanced Maternal Age Patients Without Compromising Cumulative Live-Birth Rate. J Assist Reprod Genet. 2019;36(12):2493\u0026ndash;504.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eRubio C, Bellver J, Rodrigo L, Castill\u0026oacute;n G, Guill\u0026eacute;n A, Vidal C, et al. In vitro fertilization with preimplantation genetic diagnosis for aneuploidies in advanced maternal age: a randomized, controlled study. Fertil Steril. 2017;107(5):1122\u0026ndash;9.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eUbaldi FM, Cimadomo D, Vaiarelli A, Fabozzi G, Venturella R, Maggiulli R, et al. Advanced Maternal Age in IVF: Still a Challenge? The Present and the Future of Its Treatment. Front Endocrinol (Lausanne). 2019;10:94.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eHodes-Wertz B, Grifo J, Ghadir S, Kaplan B, Laskin CA, Glassner M, et al. Idiopathic recurrent miscarriage is caused mostly by aneuploid embryos. Fertil Steril. 2012;98(3):675\u0026ndash;80.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eMeldrum DR. Introduction: Preimplantation genetic screening is alive and very well. Fertil Steril. 2013;100(3):593\u0026ndash;4.\u003c/span\u003e\u003c/li\u003e\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":true,"hideJournal":false,"highlight":"","institution":"","isAcceptedByJournal":true,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true},"keywords":"Advanced Maternal Age, PGT-A, Blastocyst, Live Birth, Embryo Aneuploidy","lastPublishedDoi":"10.21203/rs.3.rs-2124932/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-2124932/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003ch2\u003ePURPOSE\u003c/h2\u003e \u003cp\u003eDoes preimplantation genetic testing for aneuploidy (PGT-A) in embryos help women of advanced maternal age (AMA) achieve better reproductive outcomes?\u003c/p\u003e\u003ch2\u003eMETHODS\u003c/h2\u003e \u003cp\u003eMulticenter, controlled retrospective study, a total of 267 patients (n\u0026thinsp;=\u0026thinsp;267) were recruited, of which 53 patients (PGT-A group) consented to PGT-A, followed by euploid embryo transfer, whereas the remaining 214 patients (non-PGT-A group) underwent embryo transfer of un-screened morphologically graded blastocysts.\u003c/p\u003e\u003ch2\u003eRESULTS\u003c/h2\u003e \u003cp\u003eA significant increase in the clinical pregnancy rate was observed in the PGT-A group when compared to the non-PGT-A group (71.6% vs. 51%, p\u0026thinsp;=\u0026thinsp;0.007), while the miscarriage rate was found to be lower in the PGT-A group compared to the non-PGT-A group (11% vs. 25%, p\u0026thinsp;=\u0026thinsp;0.02). The live birth rates observed in either group were statistically nonsignificant (62.2% vs 51%, p\u0026thinsp;=\u0026thinsp;0.14). In the PGT-A group, similarly, the implantation rate was found to be significantly higher than in the non-PGT-A group (53% vs. 33%, p\u0026thinsp;=\u0026thinsp;0.007).\u003c/p\u003e\u003ch2\u003eCONCLUSION:\u003c/h2\u003e \u003cp\u003eThe data suggest that PGT-A testing in women of advanced maternal age can improve their reproductive outcomes.\u003c/p\u003e","manuscriptTitle":"Role of Preimplantation genetic testing in Indian women with advanced maternal age to optimize Reproductive Outcomes","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2022-10-07 20:20:11","doi":"10.21203/rs.3.rs-2124932/v1","editorialEvents":[{"type":"communityComments","content":0}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"320d0f35-1063-4099-8c3e-b519fba5be7c","owner":[],"postedDate":"October 7th, 2022","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"published-in-journal","subjectAreas":[],"tags":[],"updatedAt":"2024-03-25T09:51:51+00:00","versionOfRecord":{"articleIdentity":"rs-2124932","link":"https://doi.org/10.1142/S2661318224500014","journal":{"identity":"fertility-and-reproduction","isVorOnly":true,"title":"Fertility \u0026 Reproduction"},"publishedOn":"2024-02-07 09:51:51","publishedOnDateReadable":"February 7th, 2024"},"versionCreatedAt":"2022-10-07 20:20:11","video":"","vorDoi":"10.1142/S2661318224500014","vorDoiUrl":"https://doi.org/10.1142/S2661318224500014","workflowStages":[]},"version":"v1","identity":"rs-2124932","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-2124932","identity":"rs-2124932","version":["v1"]},"buildId":"GqpaHPwrfC8PjnIFayRh5","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}