Undergoing Preimplantation Genetic Testing Research Articles

P-239 Comparative study between two commercial vitrification kits on survival and clinical outcome for blastocyst frozen embryo transfer

Abstract Study question To compare the post thaw survival and clinical outcome of blastocyst (D5/D6) transfer between two different commercially available kits for freezing and thawing. Summary answer There is significant difference on survival of post thaw blastocyst and clinical outcomes between Kitazato (Japan) and Irvine (USA) vitrification media. What is known already Vitrification is widely used in embryo preservation for its reliable outcome and the ease of use. Different brands of commercial freezing and thawing kits are available. The main difference between the two commercial vitrification kits used in this study are the different extracellur cryoprotectant used in which Kitazato contain trehalose supplemented with hydroxypropyl cellulose while Irvine contain sucrose with dextran serum supplement. Recent study suggested that cell apoptosis in specimen vitrified in trehalose media is lower in comparison to sucrose. Study design, size, duration Retrospective study was carried out from 2022-2023. Patient with uterine abnormalities, history of recurrent implantation failure (RIF), patient that underwent Preimplantation Genetic Testing (PGT) and those transferring D2/3 embryo were excluded. Survivability of post thaw blastocyst and clinical outcome of frozen blastocyst transfer were compared between Kitazato (N = 374) and Irvine (N = 253) vitrification kit. The data was analyzed using t-test and Anova. Participants/materials, setting, methods Blastocysts that were vitrified and warmed with vitrification kit from the same manufacturer were grouped together and compared. Patient in each vitrification arms were divided into two main age groups which are <37 years old and ≥37 years old. The survivability and clinical outcome of the two respective vitrification kits was compared between the age group, blastocyst grading prior to freezing (Good, Fair, Poor) and the day of blastocyst freezing (D5 or D6). Main results and the role of chance The overall mean post thawed survival was higher in Kitazato (95.8%) compare to Irvine (91.8%) (p < 0.001). In patient age <37 years old, Kitazato group performed better when compare to Irvine group. Post thawed survival using Kitazato warming was significantly better (p < 0.001) on good (AA,AB,BA) and fair graded (BB) blastocyst with mean survival of 97.1% vs 93.4% and 94.9% vs 87.6% respectively. Similarly, post thawed survival of D5 and D6 blastocysts were significantly higher in Kitazato group when compared to Irvine group with mean survival of 96.7% vs 92.8% and 94.8% vs 90.3% respectively. However no significant difference was observed in post thawed survival outcome for patient aged ≥ 37 years old between both groups in the same study parameters. In terms of clinical outcomes, there is also no difference observed between Kitazato group and Irvine group on beta hCG positive rates. Interestingly, the Irvine group shown to have significantly higher clinical pregnancy compare to Kitazato group (53% vs 60% respectively) but no significant difference was observed in sub group for patient aged < 37 years old and ≥ 37 years old. Furthermore, the implantation rate (IR) in Irvine group group (55.7%) is also significantly higher compare to the Kitazato group (50.5%). Limitations, reasons for caution Potential confounder factors such as stimulation protocol and the embryologist experience were not factored in may have affected the outcome of the study. Sample size can be increased to improve confidence in this study. Wider implications of the findings This study demonstrates that the post thaw survivability is better in the Kitazato group while the clinical outcome is better in the Irvine group. Thus this can give a better understanding of the respective commercial vitrification kits and provide insight for laboratories to choose for their clinical practices. Trial registration number Not Applicable

P-512 Combined use of genetic diagnosis and algorithms for embryo quality assessment using artificial intelligence improves selection of embryos leading to clinical pregnancy

Abstract Study question Analyze the results of our genetic diagnosis program, evaluating the morphokinetics of the embryo using an automated algorithm, based on its morphokinetic development and quality. Summary answer Transfers with a higher KIDScore D5 v3.2 euploid embryo have a significantly higher chance of achieving pregnancy. What is known already Embryo selection using morphokinetic algorithms is a technique that allows us to select embryos with better quality and perform more transfers of a single embryo improving the clinical outcomes of treatments, this technique combined with the study of aneuploidies and genetic diseases help us to increase the chances of success in embryo implantation and in the results of live newborn at home. Currently, there are techniques available for their diagnoses that, despite the impact of the processes, allow us to select euploid embryos with greater potential for implantation. Study design, size, duration Retrospective study between 1/5/2022 and 31/12/2023 of patients undergoing preimplantation genetic testing for aneuploidy (PGTA) and preimplantation genetic testing for monogenic diseases (PGTM). A single euploid embryo was transferred. Transferred embryos are compared according to KIDScore D5 v3.2. Participants/materials, setting, methods A total of 171 cycles have been performed with PGT, 97 patients have had at least one euploid embryo or transferable mosaic. The single embryos transferred euploids are compared according to the KIDScore D5 v3.2 automated system that ranks each embryo from one to ten, based on their morphokinetic development and quality. Implantation rate, clinical gestation rate and miscarriage are analyzed. Main results and the role of chance A total of 84 single embryo transfers were performed after PGT with an implantation rate of 60.71% and a clinical gestation rate per transfer of 52.38%. According to the morphokinetic KIDScore D5 v3.2 of the transferred euploid embryo, patients with negative pregnancy had a mean KIDScore D5 v3.2 of 6.5±1.3 and for patients with positive gestation it was 7.5±1.4 (S). The mean maternal age of patients with negative and positive pregnancy was 32.8±3.5 and 34.8±3.9, respectively (NS). For the clinical gestation rate, the mean KIDScore D5 v3.2 was 7.48±1.6 (S). The miscarriage rate was 7.84%. Limitations, reasons for caution The main challenges encompass genetic and reproductive counseling and the most efficient approach to blastocyst biopsy. Wider implications of the findings Our data suggest that transfers with a higher KIDScore D5v3.2 euploid embryo have a significantly higher probability of achieving pregnancy. The evolutionary gestation rate is significantly higher when the morphokinetics of the embryo are better. Further studies with a larger number of cases are needed to better assess this trend. Trial registration number not applicable. non-clinical trials

P-529 Optical Genomic Mapping (OGM) enable cryptic chromosomal rearrangements in a couple with recurrent miscarriages undergoing preimplantation genetic diagnosis

Abstract Study question Could Optical Genomic Mapping (OGM) detect chromosomal structural variations (SVs) below the detection limit of conventional cytogenetic diagnostics in couples with recurrent miscarriages? Summary answer The Next-generation cytogenetics of Optical Genomic Mapping confirmed a cryptic chromosomal translocation in a couple with recurrent miscarriages undergoing preimplantation genetic testing for aneuploidies (PGT-A). What is known already Chromosomal structural variants (SVs) significantly contribute to infertility, recurrent pregnancy losses and congenital defects... Approximately 2-4% of couples facing recurrent miscarriages carry chromosomal abnormalities. Currently, their detection is based on convencional cytogenetic methods (karyotyping, FISH or array-CGH), however cryptic chromosomal rearrangements are difficult to identify. Next-generation cytogenetics such as OGM is able to detect SVs with a resolution of < 500 kb, allowing a better understanding of the cause of patient’s infertility. In our study, we utilized OGM to confirm a cryptic translocation in a couple experiencing recurrent miscarriages, aiming to enhance our understanding of the underlying causes of their infertility. Study design, size, duration Our study focuses on a non-consanguineous couple diagnosed with recurrent pregnancy loss that attended to our private fertility clinic to undergo assisted reproductive treatment (ART) with their autologous semen, oocytes and PGT-A. Conventional cytogenetic technique was performed on both partners by G-banding karyotyping. PGT-A was performed by NGS (Veriseq, Illumina). Optical Genomic Mapping (OGM) served as a diagnostic tool, complemented by lymphocyte FISH analysis to confirm chromosomal structural variations in these infertile patients. Participants/materials, setting, methods Ultra High Molecular Weight genomic DNA (UHMW gDNA) was extracted from frozen human peripheral blood of both partners. UHMW gDNA was quantified using Qubit dsDNA assay BR kit and Qubit 4 Fluorometer (Thermo Fisher Scientific). We used Saphyr system (Bionano Genomics) for cytogenetic diagnosis. The consensus genomic map was constructed by de novo assembly in the Bionano Solve data analysis software. SVs were identified by aligning Optical Genomic Map with the reference genome GRCh38/hg38. Main results and the role of chance The couple had suffered four prior miscarriages and undergoing three PGT-A cycles. Both partners provided a normal karyotype. Among 13 biopsied embryos, four showed CNVs on chromosomes 2(p24.3è25.3, 12Mb) and 6(p24.1è25.3, 11Mb). Following this result we suspected that the couple might has a cryptic chromosomal structural abnormalities. To test our hypothesis, we performed OGM. We analysed UHMW gDNA samples from both partners. OGM generated an average of 823.67Gb. The average N50 molecule length (≥150kb) was 318kb. The average mapping rate was 90.75% and the average labeling rate was 15.30 labels per 100kb. The average effective coverage depth was 242X. OGM successfully identified a cryptic translocation in the male partner ogm[GRch38]t(2;6)(p24.3;24.1), while the female partner showed no abnormalities. Translocation breakpoints identified by OGM were Chr2:12.560.508 and Chr6:11.622.593. The translocated segments were near to the detection limit of conventional cytogenetic analysis and with similar pattern that could be the reason for missed detection by karyotyping. Importantly, we identified one gene disrupted by OGM rearrangement breakpoint: MIR3681HG. A previous study considered that CNV loci of 2p24.3, including MIR3681HG gene, could be associated with spermatogenesis. The lymphocyte FISH analysis finding corroborated the OGM result. Limitations, reasons for caution OGM is a very powerful method for detecting chromosomal structural variants in human genomes and is able to detect cryptic structural variants. However, this is a very new diagnostic application, so there is limited experience with OGM for correlating structural variations results to infertility diagnosis in couples with recurrent miscarriages. Wider implications of the findings A balanced cryptic translocation has been identified in a couple with recurrent miscarriages. Our case report demonstrates the high diagnostic capability of Optical Genomic Mapping for cryptic SVs. OGM technology allows a more accurate cytogenetic diagnosis by detecting cryptic structural variants with high resolution in infertile patients with recurrent miscarriages. Trial registration number not applicable

P-515 The morphokinetic signature of high-level mosaic embryos in time-lapse ART analysis

Abstract Study question Do high-level mosaic embryos exhibit differences in developmental speed compared to euploid or low-level mosaic embryos? Summary answer High-level mosaic embryos exhibit slower development than euploid and low-level mosaic embryos during the blastulation and blastocyst formation stages. What is known already Time-lapse technology (TLT) has facilitated the real-time observation of human embryo development without disrupting culture conditions, generating a wealth of morphokinetic data linked to outcomes such as implantation potential, blastocyst development, and ploidy status. However, the developmental patterns of mosaic embryos based on their aneuploidy level using TLT remain not well understood. Consequently, we aimed to assess the clinical significance of morphokinetic parameters in human embryo development, with a specific focus on embryos categorized by their mosaic status. Study design, size, duration This retrospective study examined 385 blastocysts undergoing preimplantation genetic testing for aneuploidy (PGT-A) between January and December 2022. Out of the total, 191 expansion blastocysts underwent PGT-A biopsy on day 5. The PGT-A results, which categorized embryos into euploid, aneuploid, and mosaic based on their genetic composition, were determined through next-generation sequencing (NGS) by an analysis company. Mosaic embryos were further categorized into two groups: high-level mosaicism (>30% aneuploidy cells) and low-level mosaicism (≤30%). Participants/materials, setting, methods Differences in development rates were assessed through the examination of morphological and morphokinetic parameters. These parameters included the time to second polar body extrusion (tPB2), time to pronuclei appearance and fading (tPNa, tPNf), time to reach 2–8 cells (t2, t3, t4, t5, t6, t7, t8, t9), and time to the onset of blastulation and blastocyst formation (tSB, tB). Statistical analyses, incorporating ANOVA and post-hoc tests, were employed to compare developmental timings across different groups. Main results and the role of chance When categorizing mosaic embryos based on their level, high-level mosaic embryos showed a significantly slower developmental rate than low-level mosaic embryos, evident at both the blastulation stage (tsB, high-level vs. low-level: 101.80h vs. 97.3h, p=0.003) and blastocyst stage (tB, high-level vs. low-level: 109.98h vs. 105.92h, p=0.005). In comparison to euploid embryos, low-level mosaic embryos demonstrated comparable outcomes. Conversely, high-level embryos exhibited a significant difference in both blastulation (tsB, euploidy vs. high-level: 98.85h vs. 101.80h, p=0.026) and blastocyst (tB, euploidy vs. high-level: 106.73h vs. 109.98h, p=0.011) stages. These findings suggested that the mosaic level of embryos might influence developmental kinetics, proposing this approach as an additional tool for the selection of mosaic embryos in the context of embryo transfer. Limitations, reasons for caution This study focused on morphokinetic parameters captured using time-lapse imaging, which may not fully represent the complex cellular dynamics within embryos. Furthermore, the categorization of mosaic embryos into high-level and low-level groups based on the percentage of mosaicism (>30% and ≤30%, respectively) might not capture all embryo developmental potential. Wider implications of the findings The findings suggest valuable predictive potential in assessing the development of aneuploid and mosaic embryos. Further research into the development of euploid, aneuploid, and mosaic embryos, as well as high-level and low-level mosaic embryos, could provide valuable insights for ART practices. This could significantly impact ART decision-making and patient outcomes. Trial registration number not applicable

P-190 Impact of low lactate culture media in embryo development and IVF outcomes

Abstract Study question Can low lactate culture media improve embryo utilization rate in artificial reproductive technology (ART) cycles? Summary answer Embryos cultured in low lactate media showed a significant improved blastocyst utilization rate compared to embryos cultured in standard culture media. What is known already Culture media plays a key role in the embryonic development in the IVF lab. Currently, different culture medias are commercially available, yet no consensus on the most suitable media composition has been reached. Recently, minimizing metabolic stress for the embryo to enhance implantation and pregnancy rates has been suggested. Research on the metabolism of mammalian preimplantation embryos revealed that excess lactate in the culture medium, in addition to pyruvate and glucose, can burden metabolic efficiency, as embryos naturally produce lactate during energy production. Study design, size, duration The prospective, monocentric study was designed and conducted at the Kinderwunsch Institut Schenk GmbH (Dobl, Austria). A total of 60 women with an average age of 32.5 undergoing ICSI (intracytoplasmic sperm injection) treatment were included. All of them had at least 6 mature (MII) oocytes at time of insemination. Sibling oocytes were cultured in two different culture medias and outcomes were compared. Patients undergoing preimplantation genetic testing were excluded. Participants/materials, setting, methods Mature (MII) oocytes of each patient (total 621 MII oocytes) were inseminated by ICSI and then equally split into two culture medias: standard (313 oocytes) and low lactate (308 oocytes) covered with mineral-oil overlay and incubated in time-lapse under the same conditions. A total of 428 embryos were obtained. Embryos were evaluated using morphokinetic criteria. The fertilization rate, embryo utilization rate, pregnancy rate and time to Blastocyst were analyzed. Main results and the role of chance No differences were observed in the number of fertilized oocytes (n = 211 and 217 in the standard and low lactate media, respectively). The overall embryo utilization rate was significantly different (p < 0.008) resulting in 52% in standard media and 64.3% using low lactate media. Interestingly, age stratification (18-30; 31-35; 36-41) revealed overall higher embryo utilization rates when using low lactate media in all age groups. Blastocyst utilization rate was significantly different (p < 0.007) 43,1% and 58,05% in the standard and low lactate media, respectively. Regarding the time to blastocyst, it was observed that in the standard culture media 80% of the blastocysts reached blastocyst stage on D + 5 whereas in the low lactate media 92,07% reached blastocyst stage on D + 5, which was also significantly different (p < 0.001). No differences in pregnancy and live birth rates were observed. Limitations, reasons for caution The number of patients may be seen as a study limitation. Results should be confirmed with a bigger sample size. Wider implications of the findings Low lactate media could help improve IVF outcomes due to higher embryo utilization rate leading to better cumulative pregnancy rates. Excessive lactate concentration in culture media could negatively influence embryo metabolism. Trial registration number not applicable

Dependency of mitochondrial quantity on blastocyst timeline obscures its actual effect to pregnancy outcomes.

To explore the correlation between mitochondrial quantity and the blastocyst development timeline as well as their respective contributions to early pregnancy. A retrospective study was conducted using a dataset comprising 2,633 embryos that underwent preimplantation genetic testing for aneuploidy (PGT-A) between January 2016 and December 2023. The study was divided into three subsets to address distinct aspects: the representativeness of a single trophectoderm (TE) biopsy for mitochondrial quantity (n=43), the correlation between morphokinetic features and mitochondrial quantity (n=307), and the association analysis among mitochondrial quantity, blastocyst timeline factor, and reproductive outcomes (n=2,283). Distribution assessment of mitochondrial quantity across an individual blastocyst involved the identification within multiple biopsies and spent culture media. Timeline evaluation included correlating mitochondrial quantity with time-lapse datasets. Finally, multivariate logistic regression models, incorporating potential effectors alongside mitochondrial quantity, were employed to analyze their respective contributions to early pregnancy endpoints. Of distribution assessment, mitochondrial quantity exhibited an even distribution across the entire trophectoderm (Spearman's ρ=0.82), while no detectable mtDNAs in the corresponding spent culture media. Then the timeline correlation study revealed significant association between mitochondrial quantity and blastocyst features of both the day of expanded blastocyst formation (95% Confidence intervals, CIs: 0.27~4.89, p=0.03) and the timing of expanded blastocyst formation (tEB) (95% CIs: -0.24~-0.01, p=0.04) in the regression model, indicating a strong dependency between mitochondrial quantity and the blastocyst development timeline. For the contribution to early pregnancy, multivariate logistic regression models showed that the day of expanded blastocyst formation contributed to four endpoints persistently: positive for HCG (odd ratio, OR: 0.71, p=0.006), gestational sac (OR: 0.78, p=0.04), fetal heartbeat (OR: 0.71, p=0.004), and progression to 14 weeks (OR: 0.69, p=0.002). Contrastingly, no notable correlation was observed between the mitochondrial quantity and these endpoints. Strong interaction was observed between mitochondrial quantity and the blastocyst timeline, particularly the timing of expanded blastocyst formation. It suggests that the primary determinant influencing pregnancy outcomes lies in the time-dependent parameter of blastocyst rather than in the specific mitochondrial quantity.

Excessive proliferating cell nuclear antigen attenuates endometrial adhesive capacity and decidualization in patients with recurrent implantation failure.

Does the expression of proliferating cell nuclear antigen (PCNA) in the endometrium regulate endometrial receptivity in patients with recurrent implantation failure (RIF)? A high abundance of PCNA attenuates endometrial adhesive capacity and decidualization in patients with RIF. Aberrant expression of PCNA has been discovered in multiple infertility-related disorders. However, the expression pattern and role of PCNA in the establishment of endometrial receptivity and endometrial decidualization in patients with RIF remain unclear. We analysed the expression of PCNA in mid-secretory endometrial tissues from 24 patients with RIF and 24 healthy women. Additionally, PCNA expression levels were measured in proliferative and mid-secretory phase endometrial tissue samples from women with regular menstrual cycles and in decidual tissue samples taken from ten women during normal early pregnancy (n = 10 per phase for each group). The function and regulatory mechanisms of PCNA in endometrial adhesive capacity and endometrial decidualization were investigated using BeWo spheroids, Ishikawa cells, and human endometrial stromal cells (HESCs). The expression of PCNA in mid-secretory endometrial tissues of patients with RIF and women with normal endometrium and in endometrial tissue at different stages of the menstrual cycle and in decidualized tissues was analysed by RT-qPCR, western blot, and immunohistochemistry staining (IHC). Furthermore, the number of BeWo spheroids directly attached to the Ishikawa cell monolayers, and the potential molecular mechanisms involved, were compared between cells overexpressing PCNA and a control group. Additionally, the effect and regulatory mechanisms of PCNA on the decidualization of HESCs in vitro were investigated. Our findings indicated that the abundance of PCNA was dramatically greater in mid-secretory endometrial tissues from patients with RIF than in those from women with healthy endometrium. The expression of PCNA increased in the proliferative phase of the menstrual cycle but decreased gradually in the mid-secretory phase and in decidual tissues. Interestingly, PCNA was expressed in both human endometrial epithelial cells (HEECs) and HESCs. In Ishikawa cells, PCNA overexpression dramatically reduced the endometrial adhesive capacity by inhibiting the expression of adhesion molecules (E-cadherin and integrin β3) and activating the FAK/paxillin signalling pathway. Furthermore, in HESCs, PCNA overexpression attenuated endometrial decidualization by activating the AKT/β-catenin signalling pathway and increasing tight junctions between cells by upregulating ZO-1 and occludin expression. In addition, PCNA-ELAVL1 interactions were confirmed by coimmunoprecipitation in decidualized HESCs. N/A. The functional analysis of PCNA was limited by the number of human endometrial tissues. A larger sample size is required to further explore the potential roles of PCNA during embryo implantation. Moreover, the present results should be taken with caution, as only a few of the embryos that were transferred in RIF patients population underwent preimplantation genetic testing for embryonic chromosome aneuploidies (PGT-A), despite embryo ploidy testing being significant in the diagnosis of unexplained RIF. High PCNA expression attenuates endometrial adhesive capacity and decidualization in patients with RIF. These findings provide new insights into the potential mechanisms underlying the occurrence of implantation failure. This work was supported by the National Natural Science Foundation of China (82101698), Shandong Provincial Natural Science Foundation (ZR2021MH012), and the Science and Technology Plan of Yantai (2023YD021 and 2022YD031). The authors have no conflicts of interest to disclose. N/A.

A heatmap for expected cumulative live birth rate in preimplantation genetic testing for monogenic disorders and chromosomal structural rearrangements.

Our objective is to predict the cumulative live birth rate (CLBR) and identify the specific subset within the population undergoing preimplantation genetic testing for monogenic disorders (PGT-M) and chromosomal structural rearrangements (PGT-SR) which is likely to exhibit a diminished expected CLBR based on various patient demographics. We performed a single-centre retrospective cohort study including 1522 women undergoing 3130 PGT cycles at a referral centre for PGT. A logistic regression analysis was performed to predict the CLBR per ovarian stimulation in women undergoing PGT-M by polymerase chain reaction (PCR) or single-nucleotide polymorphism (SNP) array, and in women undergoing PGT-SR by SNP array, array comparative genomic hybridization (CGH) or next-generation sequencing (NGS). The mean age of women was 32.6years, with a mean AMH of 2.75µg/L. Female age and AMH significantly affected the expected CLBR irrespective of the inheritance mode or PGT technology. An expected CLBR < 10% was reached above the age of 42years and AMH ≤ 1.25µg/L. We found no significant difference in outcome per ovarian stimulation between the different PGT technologies, i.e. PCR, SNP array, array CGH and NGS. Whereas per embryo transfer, we noticed a significantly higher probability of live birth when SNP array, array CGH and NGS were used as compared to PCR. In a PGT-setting, couples with an unfavourable female age and AMH should be informed of the prognosis to allow other reproductive choices. The heatmap produced in this study can be used as a visual tool for PGT couples.

Time-lapse imaging of morula compaction for selecting high-quality blastocysts: a retrospective cohort study.

Before blastocyst development, embryos undergo morphological and metabolic changes crucial for their subsequent growth. This study aimed to investigate the relationship between morula compaction and blastocyst formation and the subsequent chromosomal status of the embryos. This retrospective cohort study evaluated embryo development (n = 371) using time-lapse imaging; 94 blastocysts underwent preimplantation genetic testing for aneuploidy (PGT-A). The embryos were classified as fully (Group 1, n = 194) or partially (Group 2, n = 177) compacted. Group 1 had significantly higher proportions of good- and average-quality blastocysts than Group 2 (21.6% vs. 3.4%, p = 0.001; 47.9% vs. 26.6%, p = 0.001, respectively). The time from the morula stage to the beginning and completion of compaction and blastocyst formation was significantly shorter in Group 1 than in Group 2 (78.6 vs. 82.4h, p = 0.001; 87.0 vs. 92.2h, p = 0.001; 100.2 vs. 103.7h, p = 0.017, respectively). Group 1 embryos had larger surface areas than Group 2 embryos at various time points following blastocyst formation. Group 1 blastocysts had significantly higher average expansion rates than Group 2 blastocysts (653.6 vs. 499.2μm2/h, p = 0.001). PGT-A revealed a higher proportion of euploid embryos in Group 1 than in Group 2 (47.2% vs. 36.6%, p = 0.303). Time-lapse microscopy uncovered a positive relationship between compaction and blastocyst quality and its association with embryo ploidy. Hence, compaction evaluation should be prioritized before blastocyst selection for transfer or cryopreservation.

Comparing blastocyst euploid rates between the progestin-primed and gonadotrophin-releasing hormone antagonist protocols in aneuploidy genetic testing: a randomised trial protocol

IntroductionProgestin can inhibit the pituitary luteinising hormone (LH) surge during ovarian stimulation for in vitro fertilisation (IVF) and studies show progestin-primed ovarian stimulation (PPOS) is effective in blocking the LH...

Unlocking Infertility: Enhancing Pregnancy Rates With Personalized Embryo Transfers Using Optimal Time for Endometrial Receptivity Analysis in Recurrent Implantation Failure Patients Undergoing In Vitro Fertilization.

Background Infertility remains a significant global challenge, and recurrent implantation failure (RIF) poses a considerable concern in assisted reproductive technology. Understanding the factors contributing to implantation failure is essential for developing accurate diagnostic tools and treatment strategies. Endometrial receptivity (ER) during the window of implantation is crucial for successful embryo implantation in in vitro fertilization (IVF) procedures. Molecular-based endometrial receptivity analysis and next-generation sequencing provide insights into ER, but there is a lack of research on these in the Indian population, particularly in patients with RIF. This retrospective cohort study evaluates the effectiveness of Optimal Timing for Endometrial Receptivity Analysis (OpERA)-guided personalized embryo transfer (pET) in Indian patients with a history of RIF. Methodology The study includes 158 female patients with a history of failed embryo transfers who underwent OpERA testing before frozen embryo transfer. Patients were categorized based on the number of previous failed transfers. OpERA outcomes were assessed, and clinical outcomes were compared between groups undergoing preimplantation genetic testing for aneuploidy (PGT-A) with and without OpERA. Endometrial preparation involved hormone replacement therapy, and OpERA testing was performed at the Neuberg Centre for Genomic Medicine using RNA extraction, cDNA conversion, and sequencing. Results OpERA outcomes showed no significant differences in receptive rates among patient groups. Group 3, with three or more failed transfers, exhibited significantly higher biochemical pregnancy rates (BPRs), clinical pregnancy rates (CPRs), and abortion rates (ARs) compared to Groups 1 and 2. OpERA with PGT-A showed significantly higher BPR, implantation rate, CPR, and lower AR compared to OpERA without PGT-A. Conclusions OpERA-guided pET, especially with PGT-A, demonstrated improved pregnancy outcomes, particularly in patients with a history of RIF. The study emphasizes the importance of OpERA in determining optimal transfer timing, moving beyond the traditional reliance on embryo quality alone.OpERA presents promise in predicting pregnancy outcomes for Indian patients with previous IVF failures. The integration of OpERA and PGT-A represents a significant advancement in personalized reproductive medicine, offering new hope for individuals grappling with infertility complexities.

A clinical predictive model for live birth in women of advanced age undergoing PGT cycles.

The trend of delaying childbirth has resulted in a growing number of advanced-aged women who are opting for preimplantation genetic testing (PGT) to screen for monogenic diseases or structural chromosomal rearrangements (PGT-M and PGT-SR). This increase in demand necessitates the development of a clinical predictive model for live birth outcomes in these women. Therefore, the objective of this study is to construct a comprehensive predictive model that assesses the likelihood of achieving a successful live birth in advanced-aged women undergoing PGT-M and PGT-SR treatments. A retrospective cohort study of 37-45-year-old women undergoing preimplantation genetic testing for monogenic disease or structural chromosomal rearrangement cycles from 2010 to 2021 was conducted at a university hospital reproductive centre. The purpose was to develop a clinical predictive model for live birth in these women. The main outcome studied was the cumulative live birth rate in the first or subsequent cycles. Developing a decision tree enabled a comprehensive study of clinical parameters and expected outcomes. The analysis included 158 women undergoing 753 preimplantation genetic testing cycles. The cumulative live birth rate was 37.342% (59/158). Decision tree analysis revealed that women aged ≤ 40.1 or women > 40.1 with one or more top-quality transferable embryos in their first cycle had the best chance for a live baby (56% and 41%, respectively). Those older than 40.1 without top-quality embryos and seven or fewer dominant follicles had no live births. A Kaplan-Meier curve showed that for autosomal dominant diseases, there was a negligible increase in live birth rate after three cycles, compared to six cycles in autosomal recessive inheritance. In older women, the chance of delivering after repeated cycles is higher in those with at least one top-quality unaffected embryo in their first preimplantation genetic testing cycle. Additional preimplantation genetic testing cycles after three in carriers of an autosomal dominant disorder and six in those with an autosomal recessive disorder should be considered prudently.

Clinical outcomes in carriers of insertional translocation: a retrospective analysis of comprehensive chromosome screening results

To evaluate the clinical outcomes in the carriers of insertional translocation (IT). Retrospective case series. University-affiliated reproductive medical center. Twenty-three couples with ITs. No direct interventions were involved; however, this study included patients who underwent preimplantation genetic testing for structural chromosomal rearrangements (PGT-SR). Outcome of preimplantation genetic testing for structural chromosomal rearrangements and percentage of blastocysts available for transfer. Among 23 IT carriers, 15 were simple interchromosome ITs (type A), 3 were intrachromosome IT carriers (type B), and 5 were interchromosome IT carriers combined with other translocations (type C). A total of 190 blastocysts from 30 cycles were biopsied, 187 embryos were tested successfully, and only 57 blastocysts (30.5%) from 21 patients were available for transfer (normal or balanced). The unbalanced rearrangement rate of type C was 79.2% (42/53), and the proportion of type A was 50.0% (57/114), which was significantly higher than that of type B (5%, 1/20). In type A, the probability of embryos harboring unbalanced rearrangement in female carriers was 56.0% (51/91), which was higher than that in male carriers (26.1%, 6/23). Furthermore, the haploid autosomal length value of the inserted fragment was correlated linearly with the incidence of abnormal embryos. In type A gametes, most gametes produced by 2:2 separation without crossover, and no 3:1 separation gamete was observed. The chance of identifying normal or balanced and mosaic blastocysts per mature oocytes in patients with ITs are 16.6% (67/404). Greater IT complexity results in fewer transferable embryos. For simple interchromosome ITs, female carriers and those with higher haploid autosomal length values have a higher risk of producing embryos with unbalanced rearrangement.

#116 : Prevalence of Aneuplodies in Chromosomes 13, 18, 21, X and Y in Embryos Generated by ICSI of Fertile Couples

Background and Aims: It has previously been reported that up to 70% of the embryos generated in an ART lab are chromosomally abnormal. Abnormalities in chromosomes 13, 18, 21, X and Y have been observed to be as common as 63% in embryos of sub-fertile couples. The aim of this study was to find if these abnormalities are just as common in embryos of fertile couples who have no history of an affected child. Method: A retrospective cohort study was conducted on 409 embryos obtained from 113 couples undergoing preimplantation genetic testing at an ART center in Pakistan, between 1 January 2021 and 31 December 2022. The inclusion criteria was that fertile couples who had come for gender selection for family balancing. Age range of female partners was 28 years to 38 years. Male partners had normal semen parameters as per the World Health Organization (WHO) guidelines. Results: A total of 70.7% (289/409) were unaffected by aneuploides in the five chromosomes. There was a significant difference in the ratio of normal to abnormal embryos. Out of the normal embryos, there was no significant difference in the male (34.5%) and female (36.1%) embryos. There was no significant difference in the number of abnormal female and male embryos (p=0.127). Among the aneuploidies in the five chromosomes, trisomy 21 was the most common (n=27; 6.6%). Conclusion: Approximately 20% of sub-fertile couples have an unknown etiology. Our results indicate that the five commonly known aneuploidies found in embryos of sub-fertile patients have a low prevalence in fertile patients. It is worth noting that this study is limited by the fact that abnormalities in sub-fertile couple were not analyzed and statistics from other already published studies was relied upon. Nonetheless, this data has provided perspectives for counselling couples of unexplained sub-fertility.

Transcriptomic profiles reveal the characteristics of oocytes and cumulus cells at GV, MI, and MII in follicles before ovulation

BackgroundThe oocyte and its surrounding cumulus cells (CCs) exist as an inseparable entity. The maturation of the oocyte relies on communication between the oocyte and the surrounding CCs. However, oocyte evaluation is primarily based on morphological parameters currently, which offer limited insight into the quality and competence of the oocyte. Here, we conducted transcriptomic profiling of oocytes and their CCs from 47 patients undergoing preimplantation genetic testing for aneuploidy (PGT-A). We aimed to investigate the molecular events occurring between oocytes and CCs at different stages of oocyte maturation (germinal vesicle [GV], metaphase I [MI], and metaphase II [MII]). Our goal is to provide new insights into in vitro oocyte maturation (IVM).ResultsOur findings indicate that oocyte maturation is a complex and dynamic process and that MI oocytes can be further classified into two distinct subtypes: GV-like-MI oocytes and MII-like-MI oocytes. Human oocytes and cumulus cells at three different stages of maturation were analyzed using RNA-seq, which revealed unique transcriptional machinery, stage-specific genes and pathways, and transcription factor networks that displayed developmental stage-specific expression patterns. We have also identified that both lipid and cholesterol metabolism in cumulus cells is active during the late stage of oocyte maturation. Lipids may serve as a more efficient energy source for oocytes and even embryogenesis.ConclusionsOverall, our study provides a relatively comprehensive overview of the transcriptional characteristics and potential interactions between human oocytes and cumulus cells at various stages of maturation before ovulation. This study may offer novel perspectives on IVM and provide a reliable reference data set for understanding the transcriptional regulation of follicular maturation.

Obstetric and Neonatal Outcomes After Transferring More Than One Embryo in Patients With Preimplantation Genetic Testing.

To compare obstetric and neonatal outcomes after single embryo transfer (SET) compared with multiple embryo transfer (MET) from frozen-thawed transfer cycles of embryos that underwent preimplantation genetic testing for aneuploidies (PGT-A). We conducted a retrospective cohort study from the SART CORS (Society for Assisted Reproductive Technology Clinic Outcome Reporting System) national database. Clinical and demographic data were obtained from the SART CORS database for all autologous and donor egg frozen-thawed transfer cycles of embryos that underwent PGT-A between 2014 and 2016, after excluding cycles that used frozen oocytes, fresh embryo transfer, and transfers of embryos from more than one stimulation cycle. Multivariable linear and log-binomial regression models were used to estimate the relative and absolute difference in live-birth rate, multiple pregnancy rate, gestational age at delivery, and birth weight between SET compared with MET. In total, 15,638 autologous egg transfer cycles and 944 donor egg transfer cycles were analyzed. Although the live-birth rate was higher with MET compared with SET in the autologous oocyte cycles (64.7% vs 53.2%, relative risk [RR] 1.24, 95% CI, 1.20-1.28), the multiple pregnancy rate was markedly greater (46.2% vs 1.4%, RR 32.56, 95% CI, 26.55-39.92). Donor oocyte cycles showed similar trends with an increased live-birth rate (62.0% vs 49.7%, RR 1.26, 95% CI, 1.11-1.46) and multiple pregnancy rate (54.0% vs 0.8%) seen with MET compared with SET. Preterm delivery rates and rates of low birth weight were significantly higher in MET compared with SET in both autologous and donor oocyte cycles and were also higher in the subanalysis of singleton deliveries that resulted from MET compared with SET. Despite some improvement in live-birth rate, nearly half of the pregnancies that resulted from MET of embryos that underwent PGT-A were multiples. Compared with SET, MET is associated with significantly higher rates of neonatal morbidity, including preterm delivery and low birth weight. The transfer of more than one embryo that underwent PGT-A should continue to be strongly discouraged, and patients should be counseled on the significant potential for adverse outcomes.

Growth hormone supplementation during ovarian stimulation in women with advanced maternal age undergoing preimplantation genetic testing for Aneuploidy

BackgroundStudies have shown that supplementation with recombinant human GH (rh-GH) during ovarian stimulation (OS) may improve the ovarian response and clinical outcomes of IVF. However, it remains unclear whether GH is associated with the ploidy status of embryos, and therefore, is unable to explain the underlying reason for the effect of GH on IVF outcomes. This study aimed to investigate whether GH supplementation in women with advanced maternal age (AMA) during OS is related to an increased probability of obtaining euploid blastocysts.MethodsThis was a single center retrospective cohort study. The data of all women aged 38–46 years who underwent their first preimplantation genetic testing for aneuploidy (PGT-A) cycle between January 2021 and June 2022 were reviewed. Patients in the GH group received 4 IU/day subcutaneous GH supplementation from the beginning of OS to the trigger day, and patients in the control group did not. A total of 140 patients in the GH group and 272 patients in the control group were included after 1:2 propensity score matching.ResultsThe baseline and cycle characteristics between the two groups were similar. The proportion of cycles which obtained euploid blastocysts was significantly higher in the GH group than that in the control group (41.43% vs. 27.21%, P = 0.00). The GH group had a significantly higher euploid blastocyst rate per cohort (32.47% vs. 21.34%, P = 0.00) and mean euploid blastocyst rate per cycle (per biopsy cycle 0.35 ± 0.40 vs. 0.21 ± 0.33, P = 0.00; per OS cycle 0.27 ± 0.38 vs. 0.16 ± 0.30, P = 0.02). However, the benefit of GH was more significant in patients aged 38–40 years, but not significant in patients aged 41–46 years. Pregnancy outcomes were similar between the two groups after embryo transfer.ConclusionsGH supplementation during OS is associated with a significantly increased probability of obtaining euploid blastocysts in women aged 38–40 years, but this benefit is not significant in women aged 41–46 years. Our results explained the underlying reason for the effect of GH on IVF outcomes in existing studies, and might be helpful for AMA patients undergoing PGT-A cycles to obtain a better outcome meanwhile to avoid over-treatment.Trial registrationNCT05574894, www.clinicaltrials.gov.

Effectiveness of atosiban in women with previous single implantation failure undergoing frozen-thawed blastocyst transfer: study protocol for a randomised controlled trial

BackgroundUterine contractions may interfere with embryo implantation in assisted reproductive technology. To reduce these contractions and improve success rates, the oxytocin antagonist atosiban has been suggested for administration during embryo...

Usable blastocysts developed from in-vitro-matured metaphase I oocytes in preimplantation genetic testing cycles

Research questionAre blastocysts derived from in-vitro-matured metaphase I (MI) oocytes less likely to produce usable embryos for transfer compared with those derived from in-vivo-matured oocytes in cycles undergoing preimplantation genetic testing (PGT)? DesignThe primary outcome was usable blastocyst rate, which was compared between blastocysts derived from in-vitro-matured MI oocytes after ovarian stimulation and from in-vivo-matured oocytes. Logistic regression analysis using generalized estimating equations was used to control for confounders in the analysis of factors that may influence the chance of a blastocyst being usable and in the comparison of embryological outcomes. Student's t-test, Mann–Whitney U test, chi-squared tests or Fisher's exact tests were used to compare clinical and pregnancy outcomes. ResultsA total of 1810 injected metaphase II (MII) oocytes from 154 PGT cycles involving 154 couples were included in this study. A total of 1577 MII oocytes were in-vivo-matured and 233 were in-vitro-matured MI oocytes. The usable blastocyst rate was similar between the in-vitro-matured MI oocyte group and the in-vivo-matured oocyte group (adjusted RR 0.97, 95% CI 0.40 to 2.34). Three live births were achieved using usable blastocysts derived from in-vitro-matured MI oocytes. ConclusionsIf in-vitro-matured MI oocytes can be fertilized and develop into blastocysts, their ability to provide usable embryos for transfer is similar compared with those developed from in-vivo-matured oocytes. These blastocysts could be considered valuable for women with few viable embryos in assisted reproductive technology cycles.

Conventional IVF is feasible in preimplantation genetic testing for aneuploidy.

To investigate the feasibility of the application of conventional in vitro fertilization (cIVF) for couples undergoing preimplantation genetic testing for aneuploidies (PGT-A) with non-male factor infertility. To evaluate the efficiency of sperm whole-genome amplification (WGA), spermatozoa were subjected to three WGA protocols: Picoplex, ChromInst, and multiple displacement amplification (MDA). In the clinical studies, 641 couples who underwent PGT-A treatment for frozen embryos between January 2016 and December 2021 were included to retrospectively compare the chromosomal and clinical outcomes of cIVF and intracytoplasmic sperm injection (ICSI). Twenty-six couples were prospectively recruited for cIVF and PGT-A treatment between April 2021 and April 2022; parental contamination was analyzed in biopsied samples; and 12 aneuploid embryos were donated to validate the PGT-A results. Sperm DNA failed to amplify under Picoplex and ChromInst conditions but could be amplified using MDA. In frozen PGT-A cycles, no significant differences in the average rates of euploid, mosaic, and aneuploid embryos per cycle between the cIVF-PGT-A and ICSI-PGT-A groups were observed. The results of the prospective study that recruited couples for cIVF-PGT-A treatment showed no paternal contamination and one case of maternal contamination in 150 biopsied trophectoderm samples. Among the 12 donated embryos with whole-chromosome aneuploidy, 11 (91.7%) presented uniform chromosomal aberrations, which were in agreement with the original biopsy results. Under the Picoplex and ChromInst WGA protocols, the risk of parental contamination in the cIVF-PGT-A cycles was low. Therefore, applying cIVF to couples with non-male factor infertility who are undergoing PGT-A is feasible.