History Of Recurrent Implantation Failure Research Articles

P-518 Differentially gene expression identified by RNA seq in aneuploid human blastocysts is more influenced by the phenomenon of aneuploidy than the involved chromosome

Abstract Study question What insights do RNA-Seq profiles provide about gene expression in aneuploid blastocysts with small and large chromosomes compared to euploid blastocysts? Summary answer The difference in gene expression is affected by aneuploidy and is more related to the presence of aneuploidy than the size of the aneuploid chromosomes. What is known already Embryonic aneuploidy is one of the main common causes of implantation failure and miscarriage. RNA-Seq investigations of the embryo in the early stages of preimplantation indicate that with the gradual replacement of the embryonic profile and the reduction of the oocyte residues, the gene expression profile in the blastocyst is more dependent on the embryo’s genome. A recent study by comparing the gene expression profile of euploid blastocysts and specific aneuploidies such as trisomy 21, trisomy 18, XXY and XO syndromes using RNA-Seq, showed how the type of aneuploidy might affect gene expression. Study design, size, duration This study was conducted on surplus embryos of the patients with history of recurrent implantation failure or recurrent miscarriage following PGT by Day-5 trophectoderm biopsy. Inclusion criteria involved maternal age ≤37, normal parental karyotypes, and ovulation by long stimulation protocol. As advanced maternal age and ovarian stimulation method can affect the aneuploidy rate and expression profile, embryos belonging to mothers > 37 years and included only patients with the long stimulation protocol were excluded. Participants/materials, setting, methods The normal embryos and those with single aneuploidy were selected for comparison of gene expression profiles using RNA sequencing. The single aneuploid embryos were classified in two groups: with defined aneuploidy(DA) of small chromosomes (including chromosomes 16, 20, 21, 22), and with undefined aneuploidy(UA) as single aneuploids for other chromosomes. To neutralize the effect of genomic content on gene expression profile in DA group, embryos were pooled reciprocally (trisomy and monosomy for the same chromosome). Main results and the role of chance Principal Component Analysis (PCA) validated embryos classification considering their differentially expressed genes (DEG) clustering into three distinct groups. To minimize the role of chance, only genes with significant adjusted P-values in both STAR and Hisat data analyses were included in consequent interpretations. In addition to comparison between and among the DA, UA and normal groups, the entire set of aneuploid groups was compared with the normal group. The DEG in both aneuploid groups compared to the normal group represented eminent upregulation rather than downregulation. Gene Ontology analyses indicated an upregulation of transcription, ubiquitination, autophagy, mitochondrial, chromosome segregation and DNA repair pathways in both DA and UA compared to normal group. Overexpression of the DNA repair pathway and other validated genes of interest in aneuploid embryos was in agreement with qPCR findings. Translation and protein synthesis pathways also showed overexpression as part of the functional response to aneuploidy. Upregulation of apoptotic process pathway was evident in the UA group. While comparison of gene expression between DA and UA showed overexpression of 143 genes, but cellular lipid synthesis was the only pathway with a significant difference. The distribution of DEG on chromosomes in the DA group was not associated with selected aneuploidies. Limitations, reasons for caution The number of biological replicates in each category was limited in order to provide more confident data by minimizing the time interval between providing the samples and RNA sequencing. Larger studies might change some affected pathways due to differences in identified DEG. Wider implications of the findings This study shed light on the black box of human preimplantation aneuploidy. Considering that the transcriptome is affected by the genomic status and some debates on genomic based current PGT approaches, this study is a step for selection of healthy and viable embryos by revealing the molecular mechanisms. Trial registration number not applicable

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-507 Prediction of clinical efficacy of PGT-A based on maternal age and embryo quality in patients with recurrent implantation failure (RIF) or recurrent pregnancy loss (RPL)

Abstract Study question Can the utility of PGT-A be predicted by selecting embryos based on both maternal age and embryo quality in patients with RIF or RPL? Summary answer The predictive value of PGT-A can potentially be determined by assessing maternal age and embryo quality of experiencing patients with RIF or RPL. What is known already Embryo quality and euploid status are crucial determinants in selecting embryos for euploid embryo transfer to achieve a successful pregnancy. This is because embryos with high morphological scores may still exhibit aneuploidy, making morphologic evaluation alone limited. This limitation is particularly important in patients with a history of RIF and RPL. Aneuploidies occur during meiosis in the embryo and increase significantly with age, becoming a major contributor to implantation failure, leading to unsuccessful pregnancies, miscarriages, and congenital birth defects. Study design, size, duration This retrospective study, conducted at a single center, assessed embryo quality in 927 blastocysts derived from 221 cases of patients with RIF and 863 blastocysts derived from 204 cases of patients with RPL. The study spanned from January 2022 to November 2023. Blastocysts were scored using the KIDScore provided by the Embryoscope® time-lapse system (Vitrolife, Göteborg, Sweden) following the manufacturer’s protocols and subsequently underwent next-generation sequencing (NGS)-based PGT-A. Participants/materials, setting, methods A total of 927 RIF and 863 RPL patients’ blastocysts were analyzed, resulting in 256 and 242 euploid embryos, respectively, using NGS. Euploid embryos were classified by morphokinetics (good quality and medium quality) and maternal age groups (≤35, 36-37, 38-39, 40-41, 42-43, 44-45, ≥46). In RIF, 62.9% (161/256) of euploid embryos were classified as good quality, and 37.1% as medium quality. In RPL, 59.1% (143/242) were classified as good quality, and 40.9% as medium quality. Main results and the role of chance The euploid embryo rate decreases in both good- and medium-quality embryos as the patient’s age increases. For RIF patients: ≤35 (47.1% and 22.7%), 36-37 (46.7% and 19.8%), 38-39 (37.5% and 16.4%), 40-41 (37.5% and 11.5%), 42-43 (31.8% and 10.0%), 44-45 (0% and 11.1%), ≥46 (0% and 0%). For RPL patients: ≤35 (46.0% and 25.4%), 36-37 (45.9% and 25.0%), 38-39 (43.8% and 15.1%), 40-41 (27.9% and 18.0%), 42-43 (26.3% and 11.7%), 44-45 (0% and 12.5), ≥46 (0% and 0%). Moreover, ongoing pregnancy or live birth rates also decrease as the patient’s age increases, while the miscarriage rate increases. For RIF patients: ≤35 (57.8% and 16.1%), 36-37 (50.0% and 18.8%), 38-39 (46.2% and 25.0%), 40-41 (43.5% and 28.6%), 42-43 (36.4% and 33.3%), 44-45 (0% and 100%). For RPL patients: ≤35 (59.5% and 12.0%), 36-37 (52.0% and 18.8%), 38-39 (47.8% and 31.3%), 40-41 (43.5% and 37.5%), 42-43 (40.0% and 42.9%), 44-45 (0% and 100%). However, embryo transfer was not possible for those over 46 due to all embryos being aneuploidy. Therefore, PGT-A is valuable for patients under 42, proves beneficial for reducing the abortion rate, and achieving a high pregnancy rate for those aged 42-43, and should be considered for patients over 44. Limitations, reasons for caution This data is from a retrospective study, and well-designed prospective studies with data from multiple centers, larger samples, and extended study periods are needed to validate the current results. Additionally, obtaining information on demographic characteristics, detailed embryo information, and overall clinical outcomes can further enhance the current results. Wider implications of the findings These findings suggest that individualizing each case based on both maternal age and embryo quality is helpful in predicting the utility of PGT-A. Such personalized considerations can result in significant benefits for patients by optimizing outcomes, reducing cycle times, and consequently lowering related costs. Trial registration number not applicable

Evaluating the Impact of Long-Term GnRH Agonist Therapy on Pregnancy Outcomes in Endometriosis-Associated Implantation Failure and Pregnancy Loss

Purpose This study aimed to investigate the efficacy of long-term gonadotropin-releasing hormone (GnRH) agonist therapy in preventing endometriosis progression and relieving symptoms, particularly on pregnancy outcomes during thawed embryo transfer in patients experiencing endometriosis and recurrent implantation failure or recurrent pregnancy loss. Methods In individuals with clinical endometriosis and a history of recurrent implantation failure or recurrent pregnancy loss, we conducted a comparative analysis of clinical outcomes between those undergoing long-term GnRH agonist treatment for symptom relief, such as menstrual pain, followed by embryo transfer using Hormone Replacement Therapy (HRT) cycle, and those undergoing embryo transfer using an HRT cycle without GnRH agonist treatment. The study examined various clinical outcomes between the two groups. Results The primary outcomes included live birth rate (LBR), miscarriage rate, biochemical pregnancy rate, and perinatal complications. The GnRH agonist group showed significantly higher LBR than the control group (37.50% vs. 13.04%; p=0.02). Multivariable logistic regression analysis, adjusted for age and gravidity, showed significantly higher LBR in the GnRH agonist group compared to the control group (odds ratio: 15.3; 95% confidence interval: 2.30, 102.00; p=0.005). Conclusions The findings of this study suggested that employing a GnRH agonist in the embryo transfer protocol is effective for patients with endometriosis experiencing recurrent implantation failure or recurrent pregnancy loss.

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 CASE OF HYPERTRIGLYCERIDEMIC PANCREATITIS SECONDARY TO TACROLIMUS AND ESTRADIOL USE FOR IN VITRO FERTILIZATION

Hypertriglyceridemia is a metabolic condition with multifactorial etiology that may cause life-threatening complications, such as pancreatitis. In pregnant women, hypertriglyceridemia may be secondary to physiological changes during pregnancy or may be exacerbated by medications. The combination of tacrolimus and estradiol is sometimes used in in vitro fertilization (IVF) for women with a history of recurrent implantation failure. This case report discusses a case of acute pancreatitis caused by hypertriglyceridemia secondary to the use of tacrolimus and estradiol.

Impact of endometrial compaction on reproductive outcomes after cryotransfer of euploid embryos in a modified natural cycle: protocol for a prospective cohort study.

Embryo implantation is a complex and poorly understood process. Most studies to date have focused on the analysis of the endometrium at the end of the estrogenic phase, while the available data on its importance after secretory transformation are limited and inconsistent. Current evidence does not allow for a conclusive interpretation of the changes observed in the pre-implantation endometrium, whether in the natural or replacement cycle, and their relevance in the development of a pregnancy or the implications for clinical practice. Multicenter prospective observational cohort study. Based on our sample size calculation, the study group will consist of 206 women (exposed or "compaction" group: 103 women with a decrease of ≥ 5% in endometrial thickness between the estrogenic phase and the day of embryo transfer; non-exposed "non-compaction" group: 103 women with similar or greater endometrial thickness between these time points). The main objective of this study is to compare the ongoing pregnancy rates in natural cycles for euploid embryo transfer in patients who present endometrial compaction at the time of transfer versus those who with a stable or greater endometrial thickness with respect to the estrogenic phase. The estimated duration of the study is 30 months. Inclusion criteria are: 18 to 50 years of age, with primary or secondary infertility, subjected to endometrial preparation in a modified natural cycle for transfer of a genetically euploid blastocyst, from their own oocyte or oocyte donation, with a normal uterine cavity. Exclusion criteria are: uterine or endometrial disease (e.g., multiple myomatosis, severe adenomyosis, Asherman syndrome, refractory endometrium), conditions that prevent correct ultrasound assessment (tilted uterus), or a history of recurrent implantation failure or repeated miscarriages. The findings from this study will provide valuable insights into the potential influence of the "endometrial compaction" phenomenon on reproductive outcomes during natural cycle endometrial preparation. By examining this aspect, we aim to contribute to a better understanding of the factors that may impact successful outcomes in fertility treatments.

Endometrial proteomic profile of patients with repeated implantation failure.

Successful embryo implantation, is the initiating step of pregnancy, relies on not only the high quality of the embryo but also the synergistic development of a healthy endometrium. Characterization and identification of biomarkers for the receptive endometrium is an effective method for increasing the probability of successful embryo implantation. Endometrial tissues from 22 women with a history of recurrent implantation failure (RIF) and 19 fertile controls were collected using biopsy catheters on 7-9 days after the peak of luteinizing hormone. Differentially expressed proteins (DEPs) were identified in six patients with RIF and six fertile controls using isobaric tag for relative and absolute quantitation (iTRAQ)-based proteomics analysis. Two hundred and sixty-three DEPs, including proteins with multiple bioactivities, such as protein translation, mitochondrial function, oxidoreductase activity, fatty acid and amino acid metabolism, were identified from iTRAQ. Four potential biomarkers for receptive endometrium named tubulin polymerization-promoting protein family member 3 TPPP3, S100 Calcium Binding Protein A13 (S100A13), 17b-hydroxysteroid dehydrogenase 2 (HSD17B2), and alpha-2-glycoprotein 1, zinc binding (AZGP1) were further verified using ProteinSimple Wes and immunohistochemical staining in all included samples (n=22 for RIF and n=19 for controls). Of the four proteins, the protein levels of TPPP3 and HSD17B2 were significantly downregulated in the endometrium of patients with RIF. Poor endometrial receptivity is considered the main reason for the decrease in pregnancy success rates in patients suffering from RIF. iTRAQ techniques based on isotope markers can identify and quantify low abundance proteomics, and may be suitable for identifying differentially expressed proteins in RIF. This study provides novel evidence that TPPP3 and HSD17B2 may be effective targets for the diagnosis and treatment of non-receptive endometrium and RIF.

O-291 Clinical Application of Noninvasive PGT for Patients with Recurrent Pregnancy Loss or Repeated Implantation Failure

Abstract Study question Has noninvasive PGT been allied with the patients experiencing recurrent pregnancy loss (RIF) and recurrent implantation failure (RPL)? What are the clinical beneficiaries? Summary answer Embryo selection through noninvasive PGT can reduce the miscarriage rate in patients experiencing RPL and improve the clinical pregnancy rate in patients experiencing RIF. What is known already Chromosomal abnormalities exist in early human embryos, especially in patients with RPL and RIF. Therefore, embryos are usually evaluated through pre-implantation genetic testing for aneuploidy (PGT-A). However, a biopsy is an important concern for undetermined health risks. Meanwhile, the researchers observed genomic DNA contents in the embryo culture medium. Since then, multiple studies have been published using culture medium for analyzing chromosomal ploidy. In 2016, Xu et al. first reported a noninvasive chromosome screening (NICS) assay using a spent blastocyst culture medium. Nevertheless, the clinical application of NICS has not been evaluated in patients experiencing RPL or RIF. Study design, size, duration We designed a retrospective cohort study including 303 subjects from July 2018 to May 2021, according to the records of the Reproductive Centre at the Second Affiliated Hospital of Wenzhou Medical University. Patients experiencing RPL or RIF who received the NICS for aneuploidy were included in the NICS group, while those who underwent conventional morphology embryo transfer during the same period were included in the non-NICS group. Participants/materials, setting, methods We included women with a history of RPL (≥2 pregnancies) or RIF (≥3 implantations), exclusion criteria were antiphospholipid syndrome (APS), diabetes, hypothyroidism, or other severe complications. Routine IVF/ICSI was performed based on sperm quality. The embryos were placed in droplets. Approximately 30 mL of blastocyst medium from each embryo was transferred into cell lysis buffer. Whole-genome amplification was performed using culture media, followed by library preparation using ChromInst (Yikon Genomics; EK100100724 NICS Inst Library Preparation Kit). Main results and the role of chance For the patients experiencing RPL, the miscarriage rate per FET was significantly lower in the NICS group than in the non-NICS group (17.9% vs. 42.6%), whereas the ongoing pregnancy rate (40.7% vs. 25.0%) and live birth rate (38.9% vs. 20.6%) were significantly higher in the NICS group compared to the non-NICS group. Nevertheless, no differences were identified in pregnancy rates per patient between the NICS and non-NICS groups (49.6% vs. 44.9%). For the patients experiencing RIF, the pregnancy rates per FET were significantly higher in the NICS group than in the non- NICS group (46.9% vs. 28.7%), whereas the live birth rate and ongoing pregnancy rate per FET and per patient were no significant difference in the NICS group than in the non-NICS group. Nevertheless, no differences were identified in the miscarriage rate per clinical pregnancy between the NICS and non-NICS groups (23.3% vs. 25.9%). Limitations, reasons for caution As a retrospective study, patients in the NICS groups had different clinical prognoses than those in the non-NICS groups, introducing bias in the study results. Wider implications of the findings Non-invasive PGT can be used not only for aneuploidy detection but also for a comprehensive evaluation of morphology and DNA concentration, mosaic ratio, resolution, etc. The accumulation of clinical outcomes can also be combined with the clinical data of patients as an index to predict the clinical outcomes of embryos. Trial registration number NA

P-729 Factors affecting live birth rates in frozen-thawed single euploid blastocyst transfer cycles

Abstract Study question What are the factors affecting the rates of live birth in frozen-thawed euploid embryo transfer cycles? Summary answer Some demographic characteristics, frozen embryo transfer (FET) cycle parameters, and embryo characteristics of patients affect live birth rates in euploid embryo transfer cycles. What is known already The aim of assisted reproductive techniques (ART) is to achieve live birth as soon as possible. Preimplantation genetic testing (PGT) is an embryo selection technique recommended to shorten the live birth time. In addition, PGT reduces the number of failed ART cycles by eliminating embryos with chromosomal abnormalities that will not implant or will cause miscarriage. Despite these, the live birth rate after euploid embryo transfer cycles is still not at the expected level. Study design, size, duration This retrospective cohort study was conducted between January 2011 and May 2021 at the IVF and Reproductive Genetics Centre, Memorial Sisli Hospital, Istanbul, Turkey. The study involved a total of 2783 frozen-thawed euploid embryo transfer cycles. Participants/materials, setting, methods 2873 euploid FET cycles were analysed in the study to determine factors affecting live birth rates. 30 parameters thought to affect live birth were identified and analysed by logistic regression analysis on the effects of live birth. These parameters include patients' demographics, fresh cycle and FET cycle characteristics, and embryo-related parameters on live birth effects in euploid embryo transfer cycles. Main results and the role of chance 53% of 2783 euploid embryo transfer cycles have resulted in a live birth. The birth rate was 0,5 times less in the group with a body mass index (BMI) of higher than 29 compared to the group with a BMI of less than 25 (p < 0.05). In cycles in which embryos of good-to-moderate-to-poor quality were transferred, the chance of live birth was lower than in cycles in which embryos of top quality were transferred (OR:0.837 OR:0.515 OR:0.528, respectively). The live birth rate was higher in cases where endometrial preparation was performed in natural cycles than in artificial cycles (OR:1.67, p < 0.05). Embryo transfers without separate blastomeres had a higher live birth rate than embryo transfers with separate blastomeres (OR:1.416, p < 0.006). The live birth rate was 0.7 times lower in the group that did not undergo embryo re-expansion 4 hours after thawing (p < 0.04). Embryo transfer on the sixth day was associated with a lower live birth rate than embryo transfer on the fifth day. (OR:0.6, p < 0.02). The group with a history of recurrent implantation failure (RIF) had a lower live birth rate than the group without a history of RIF (OR:0.5) Limitations, reasons for caution This study is a retrospective study. Wider implications of the findings In euploid embryo transfer cycles, clinicians and embryologists must know the parameters affecting live birth to guide treatment and select the embryo to be transferred first. In addition, the patient should be informed and guided about these factors. Trial registration number Not applicable

Human Leukocyte Antigen Alleles Compatibility and Immunophenotypic Profile Associations in Infertile Couples.

The maternal immune system has a major role in the successful embryo implantation and maintenance of the pregnancy. This study aimed to investigate the maternal immunophenotyping profile (percentage of Natural Killer [NK] cellsand the CD4/CD8 [cluster designation] ratio in peripheral blood lymphocytes)and the HLA (Human Leukocyte Antigen)-DQA1 alleles sharing in infertile couples. This cross-sectional study included 78 women who had experienced at least two spontaneous miscarriages and 110 women with a history of recurrent implantation failures after in vitro fertilization (IVF) or intracytoplasmic sperm injection (ICSI) and embryo transfer (ET) (IVF-ET failures). The NK cell percentage and the CD4/CD8 ratio were determined by flow cytometry. Genotyping of the HLA-DQA1alleles was carried out for all women and their partners, and couple HLA-DQA1 compatibility was expressed as the percentage of commonHLA-DQA1 alleles (totaling 35 alleles)shared between spouses to the sum of the unique alleles observed. In women with recurrent miscarriages, high values (%) of the NK population with a median (interquartile range [IQR]) of 10.3% (7.7% to 12.5%) and CD4/CD8 ratio (1.7) (1.5 to 2.1) were found. In women with IVF-ET failures, the (%) NK population (10.5%) (8.6% to 12.5%) and CD4/CD8 ratio (1.8) (1.5 to 2.1) were similarly increased (p=0.390, and p=0.490, respectively). The proportion of women with >10% NK cells was 53.8% and 58.2% in women with miscarriages and IVF-ET failures, respectively (p=0.554). The prevalence of HLA-DQA1*5 allele carriage was elevated in women with miscarriages as well as those with IVF-ET failures (52.6% and 61.8%, respectively; p=0.206). The proportion of couples with high (>50%) HLA-DQA1 sharing was 65.4% in the group with miscarriages and 73.6% in the group with IVF-ET failures, respectively (p=0.222). The CD4/CD8 ratio was statistically significantly positively correlated with the (%) NK population in women with IVF-ET failures (rho = 0.297, p=0.002) and with the (%) HLA-DQA1 sharing in the group with miscarriages (rho = 0.266, p=0.019). The couples in which both spouses were carriers of the HLA-DQA1*5 allele had an increased probability of high (>50%) HLA-DQA1 compatibilitycompared with the couples in which neither of the spouses carried the allelein the miscarriage group (OR = 24.3, 95% CI: 3.0 to 198.9, p<0.001), and the IVF-ET failure group (OR = 10.5, 95% CI: 2.2 to 49.8, p<0.001). The peripheral NK (%) population and CD4/CD8 ratio, as well as the prevalence of the HLA-DQA1*5 allele, were elevated in women with recurrent miscarriages and IVF-ET failures. Furthermore, these couples with negative reproductive outcomes had a high percentage of HLA-DQA1 allele similarity. The presence of the HLA-DQA1*5 allele in spouses was strongly associated with overall couple HLA-DQA1 compatibility, implying that it could be used as a surrogate marker for assessing overall immunological compatibility in infertile couples.

Is it necessary for young patients with recurrent implantation failure to undergo preimplantation genetic testing for aneuploidy?

To determine whether preimplantation genetic testing for aneuploidy (PGT-A) can improve the pregnancy outcomes of patients aged under 38 years who have a history of recurrent implantation failure(RIF). Retrospective cohort study. We retrospectively studied the pregnancy outcomes of RIF patients aged under 38 years from January 2017 to December 2021.178 patients were divided into two groups according to whether they underwent PGT-A: the PGT-A group(n=59)and the control group(n=119).In the PGT-A group, we compared the euploidy rate of the different quality and developmental rate blastocysts. In both groups,the patients were the first frozen-thaw single blastocysts transfer after the diagnosis of RIF. Among the pregnancy outcomes, the clinical pregnancy rate was assessed as the primary outcome. The spontaneous abortion rate and ongoing pregnancy rate were the secondry outcomes. The generalized estimation equation was used to adjust for the blastocysts derived from the same patients. Multivariate logistic analysis models were used to compare the pregnancy outcomes between the two groups. In the PGT-A group, 293 blastocysts obtained from59 patients underwent PGT-A. The proportions of euploidy, aneuploidy and mosaic blastocysts were 56.31%, 25.60% and 18.09%, respectively. A comparison of the euploidy rates of different quality blastocysts showed that the rate of good-quality blastocysts was significantly higher than that of poor-quality blastocysts (67.66% vs 46.88%; odds ratio [OR], 2.203; 95%confidence interval[CI], 0.943-3.612; P=0.002). However, no significant difference was observed in the different developmental rates blastocysts. Compared with Day 5 blastocysts, the euploidy rates of Day 6 and Day 7 blastocysts were not significantly different(61.54%vs51.91%; OR,0.945; 95%CI, 0.445-2.010; P=0.884; and 61.54%vs47.37%; OR, 1.106; 95%CI, 0.774-1.578; P=0.581, respectively).As for the pregnancy outcomes, the clinical pregnancy rate was significantly increase after the use of PGT-A compared with the control group(71.19%vs56.30%; OR, 0.538; 95%CI, 0.262-1.104; P=0.039). However, the spontaneous abortion rates and ongoing pregnancy rates were not significantly different between the control and PGT-A groups (21.43% vs 19.40%; aOR,0.727; 95%CI,0.271-1.945; P=0.525; and55.93% vs 45.38%; aOR, 0.649; 95%CI, 0.329-1.283; P = 0.214,respectively). PGT-A improved the clinical pregnancy rate after blastocyst transfer in RIF patients aged under 38 years.

P-413 Non-autologous human platelet lysate stimulates in vitro proliferation of primary human endometrial cells from patients with recurrent implantation failure

Abstract Study question Does patient diagnosis impact induction of primary human endometrial cell proliferation by commercial non-autologous human platelet lysate (HPL)? Summary answer HPL stimulates in vitro proliferation of both primary endometrial epithelial cells (EECs) and stromal cells (ESCs) from patients with and without recurrent implantation failure (RIF). What is known already Inadequate endometrial receptivity and thickness are major causes for RIF. Our preliminary results suggested that non-autologous HPL stimulates in vitro proliferation of primary human EECs and ESCs isolated from patients with a history of RIF, with and without a thin endometrium (TE). In addition, using an in vitro model of embryo attachment, we reported that 48-hour treatment with HPL significantly augments the attachment of trophoblast spheroids (generated with HTR-8/SVneo cells) to EECs, suggesting an increase in endometrial receptivity following treatment. This suggests that HPL may standardize future clinical treatments for a TE and endometrial origins of RIF. Study design, size, duration Endometrial tissue was collected from five patients without RIF (control) and eighteen RIF patients at the CReATe Fertility Centre, Toronto, Canada. The eighteen RIF patients were further classified into three groups (N = 6 each) based on endometrial phenotype: 1) proliferative phase RIF only (without a TE), 2) secretory phase RIF only (without a TE), and 3) secretory phase RIF+TE (with a TE). Primary EECs and ESCs were enzymatically isolated and cultured separately. Participants/materials, setting, methods Primary EECs and ESCs were serum-starved with serum-free culture media (SFM) for 24 hours and then treated for 48 hours with the following treatment media: SFM (negative control), or SFM supplemented with 1% HPL for EECs, or 10% HPL for ESCs. Cell viability and proliferation were assessed using the metabolic assay PrestoBlue reagent and immunocytochemistry to quantify cells actively expressing the nuclear proliferation marker Ki67. Main results and the role of chance The metabolic assay demonstrated that 48-hour treatment with non-autologous HPL stimulates a similar significant increase in EEC viability and proliferation for all patient groups. EECs from patients without RIF (control) had the highest fold increase (1.49-fold, P &amp;lt;0.001), followed by proliferative phase RIF only (1.41-fold, P &amp;lt;0.001), secretory phase RIF+TE (1.39-fold, P &amp;lt;0.001), and secretory phase RIF only (1.24-fold, P &amp;lt;0.05). For ESCs, HPL stimulated a significant 2.69-fold increase in cell viability and proliferation for the secretory phase RIF+TE patient group (P &amp;lt;0.01). Pairwise comparison of the ratio of actively proliferating (Ki67+) cells between SFM and HPL treatment revealed that EECs isolated from control patients had the highest and most significant increase by 27.3% (P &amp;lt;0.05) after HPL treatment. Although not statistically significant, EECs isolated from all RIF patients displayed a similar trend of increased Ki67+ cells after HPL treatment. Whereas HPL treatment significantly increased the ratio of Ki67+ cells for ESCs isolated from both the proliferative phase (by 23.9%, P &amp;lt;0.05) and secretory phase (by 29.9%, P &amp;lt;0.05) endometrium of RIF only patients. Limitations, reasons for caution Although our data suggests that HPL treatment significantly stimulates primary endometrial cell proliferation in vitro, our sample size per group is small (N = 5-6). A larger sample size and future randomized controlled trials are needed to determine the efficacy of HPL as a treatment for a TE and/or RIF. Wider implications of the findings Our study provides the groundwork to improve clinical treatment of a TE and endometrial origins of RIF. We anticipate that in addition to stimulating cell proliferation, HPL will also induce a broad transcriptional response towards improved endometrial receptivity. We will next focus on characterizing the transcriptomic profile following HPL treatment. Trial registration number not applicable

Retraction notice to ‘The impact of an endometrial receptivity array on personalizing embryo transfer for patients with infertility: a meta-analysis’ [F&S Reviews 3 (2022) 157-173]

<h3>Objective</h3> To investigate the effectiveness of the endometrial receptivity array (ERA) test on in vitro fertilization (IVF) outcomes. <h3>Evidence Review</h3> The PubMed/MEDLINE, ScienceDirect, and Scopus databases were searched. We included studies investigating IVF outcomes between patients who underwent standard embryo transfer and personalized embryo transfer (PET) guided by the ERA results. The main outcome measures were the implantation rate, clinical pregnancy rate, ongoing pregnancy rate, miscarriage rate, and live birth rate (LBR). The random-effects model was applied to pool data. We evaluated cohort and randomized controlled trial studies using the Strengthening the Reporting of Observational Studies in Epidemiology guidelines and the Cochrane Collaboration tool. <h3>Results</h3> Seventeen studies (7,052 patients, 4 randomized controlled trials, and 13 cohort studies) were included in this meta-analysis. Personalized embryo transfer on the basis of ERA was not found to optimize the gestational outcomes, including the implantation rate (risk ratio [RR], 1; 95% confidence interval [CI], 0.83–1.2), clinical pregnancy rate (RR, 0.99; 95% CI, 0.85–1.15), ongoing pregnancy rate (RR, 0.99; 95% CI, 0.89–1.11), and miscarriage rate (RR, 1.12; 95% CI, 0.81–1.54). Further subgroup analysis indicated that patients undergoing PET in the first IVF cycle had a markedly increased LBR (RR, 1.24; 95% CI, 1.03–1.49). Conversely, PET among patients with a history of recurrent implantation failure did not enhance the LBR (RR, 0.86; 95% CI, 0.64–1.36). <h3>Conclusion</h3> Endometrial receptivity array shows no significant improvement in IVF outcomes except in the LBR for patients undergoing the first IVF cycle. Additional research is required to ascertain the efficacy of ERA before achieving wider usage. <h3>PROSPERO Number</h3> CRD42021255124.

Endometrial HLA-F expression is influenced by genotypes and correlates differently with immune cell infiltration in IVF and recurrent implantation failure patients.

Is human leukocyte antigen (HLA)-F protein expressed in mid-secretory endometrium, and are its expression levels influenced by HLA-F gene polymorphisms and correlated with the abundance of uterine natural killer (uNK) cells and anti-inflammatory M2 macrophages? HLA-F protein is expressed in mid-secretory endometrium, and levels are correlated with immune cell infiltration, plasma progesterone concentrations and HLA-F single-nucleotide polymorphisms (SNPs), however, women experiencing recurrent implantation failure (RIF) show differences when compared to women attending their first IVF treatment. The immunomodulatory HLA class Ib molecules HLA-G and HLA-F are expressed on the extravillous trophoblast cells and interact with receptors on maternal immune cells. Little is known regarding HLA-F expression in endometrial stroma and HLA-F function; furthermore, HLA-F and HLA-G SNP genotypes and haplotypes have been correlated with differences in time-to-pregnancy. Primary endometrial stromal cell (ESC) cultures (n = 5) were established from endometrial biopsies from women attending IVF treatment at a fertility clinic. Basic HLA-F and HLA-G protein expression by the ESCs were investigated. A prospective controlled cohort study was performed including 85 women with a history of RIF and 36 control women beginning their first fertility treatment and with no history of RIF. In some analyses, the RIF group was divided into unknown cause, male infertility, female infertility, and both female and male infertility. Endometrial biopsies and blood samples were obtained the day equivalent to embryo transfer in a hormone-substituted cycle. HLA protein expression by ESCs was characterized using flow cytometry and western blot. In the cohort study, the specific immune markers HLA-F and HLA-G, CD56 and CD16 (NK cells), CD163 (M2 macrophages), FOXP3 (regulatory T cells) and CD138 (plasma cells) were analysed by immunohistochemistry and a digital image analysis system in endometrial biopsies. Endometrial receptivity was assessed by an endometrial receptivity array test (the ERA® test). Endometrial biopsies were examined according to modified Noyes' criteria. SNPs at the HLA-F gene and HLA-G haplotypes were determined. HLA-F protein is expressed in the endometrium at the time of implantation. Furthermore, the HLA-F protein levels were different according to the womeńs HLA-F SNP genotypes and diplotypes, which have previously been correlated with differences in time-to-pregnancy. Endometrial HLA-F was positively correlated with anti-inflammatory CD163+ M2 macrophage infiltration and CD56+ uNK cell abundance for the entire cohort. However, this was not the case for CD56+ in the female infertility RIF subgroup. HLA-F levels in the endometrial stroma were negatively correlated with plasma progesterone concentrations in the RIF subgroup with known female infertility. Conversely, HLA-F and progesterone were positively correlated in the RIF subgroup with infertility of the male partner and no infertility diagnosis of the woman indicating interconnections between progesterone, HLA-F and immune cell infiltration. Glandular sHLA-G expression was also positively correlated with uNK cell abundance in the RIF subgroup with no female infertility but negatively correlated in the RIF subgroup with a female infertility diagnosis. Immunohistochemistry analyses of endometrial biopsies and DNA sequencing of HLA genes. Data will be shared upon reasonable request to the corresponding author. The control group of women attending their first IVF treatment had an anticipated good prognosis but was not proven fertile. A significant age difference between the RIF group and the IVF group reflects the longer treatment period for women with a history of RIF. The standardization of hormonal endometrial preparation, which allowed consistent timing of endometrial and blood sampling, might be a strength because a more uniform hormonal background may more clearly show an influence on the immune marker profile and HLA class Ib levels in the endometrium by other factors, for example genetic polymorphisms. However, the immune marker profile might be different during a normal cycle. The findings further highlight the importance of HLA-F and HLA-G at the implantation site and in early pregnancy for pregnancy success. Diagnostic measures and modulation of the complex interactions between HLA class Ib molecules, maternal immune cells and hormonal factors may have potential to improve fertility treatment. This work was supported by the Region Zealand Health Sciences Research Foundation and the Zealand University Hospital through the ReproHealth Research Consortium ZUH. The authors declared there are no conflicts of interest.

Assessment of in vitro fertilization outcomes after hysteroscopic isthmoplasty in recurrent implantation failure.

The aim of this study was to assess the in vitro fertilization (IVF) treatments outcomes of women with recurrent implantation failure (RIF) after hysteroscopic isthmoplasty for isthmocele. This retrospective, observational study included 61 patients with RIF and isthmocele who presented to IVF Unit of Hüma Obstetrics and Gynecology Hospital between January, 2018 and December, 2020. Thirty-one patient who underwent hysteroscopic isthmoplasty-before IVF procedure-were included. Remaining 30 patients underwent IVF without hysteroscopic isthmoplasty were included as controls. The fertility outcome were compared between groups. After 3months, spontaneous pregnancy was achieved in 18.4% (7/38) of patients with isthmocele who underwent hysteroscopic isthmoplasty. In the isthmoplasty group, implantation rate, clinical pregnancy rate, biochemical pregnancy rate, and the miscarriage rate were 41.9% (13/31), 35.5% (11/31), 6.5% (2/31), and 9.7% (3/31), respectively. The live birth rate after in vitro fertilization was 25.8% (8/31) in the isthmoplasty group and 3.3% (1/30) in the control group (p < 0.05). There were no significant differences in the IVF outcomes such as the biochemical pregnancy rate and miscarriage rate between the isthmoplasty and control groups. This study demonstrated that hysteroscopic isthmoplasty improves pregnancy and live birth rates for women with a history of RIF and isthmocele. Reproductive results of the IVF cycles after hysteroscopic correction of isthmocele were comparable to those of the patients without any uterine abnormalities and those with diagnosis of unexplained infertility.

Evaluation of the effect of intrauterine injection of platelet-rich plasma on the pregnancy rate of patients with a history of implantation failure in the in vitro fertilization cycle.

ABSTRACTObjectives:Implantation failure is a major problem in reproductive medicine, and despite the various methods described for treatment, there is little consensus on the most effective method. This study was conducted to investigate the effect of intrauterine injection of platelet-rich plasma (PRP) on the pregnancy rate of patients with a history of implantation failure in the in vitro fertilization (IVF) cycle.Study Design:In this study, women attending the infertility clinic of Ali ibn Abi Taleb Hospital, who had a history of implantation failure and were candidates for frozen embryo transfer (FET), were examined in Zahedan (Iran) in 2019. The patients were divided into two groups of PRP recipients and the control group. IVF was performed routinely, and in the PRP-receiving group, an intrauterine injection was given 48 h before embryo transfer (ET). The number of gestational sacs, the rate of implantation, the frequency of chemical and clinical pregnancies as well as the frequency of abortion were compared in the two groups. The collected data were analyzed by the SPSS software version 21Results:Ninety patients with a history of implantation failure participated in the study, and finally, the information of 85 patients was studied. The frequency of chemical pregnancy was 40% in the experimental group, 27% in the control group, and regarding clinical pregnancy, 33% in the experimental group and 24% in the control group, but there was no significant difference between the two groups. The rate of implantation, the mean thickness of the endometrium, and the frequency of abortion did not differ significantly between the two groupsConclusion:In patients with endometrial thickness greater than 8 mm with a history of recurrent implantation failure, intrauterine injection of PRP had no effect on the fertility outcome.

IVF outcomes after T-shaped uterine metroplasty in patients with recurrent implantation failure

BackgroundThe aim of this study was to assess the in vitro fertilization (IVF) treatments outcomes of women with recurrent implantation failure (RIF) after hysteroscopic metroplasty for T-shaped uterus. MethodsThis retrospective, observational study included 85 patients with RIF and T-shaped uterus who presented to In Vitro Fertilization Unit of Hüma Obstetrics and Gynecology Hospital between January, 2018 and August, 2021. The hysteroscopic metroplasty was performed before IVF procedure in 43 of patients included. Remaining 42 patents underwent IVF without hysteroscopic metroplasty were included as controls. The fertility outcome were compared between groups. ResultsAfter 3 months, spontaneous pregnancy was achieved in 10.4% (5/48) of patients with T-shaped uterus underwent hysteroscopic metroplasty. In the hysteroscopy group, implantation rate, clinical pregnancy rate, biochemical pregnancy rate and the miscarriage rate were 55.8% (24/43), 41.9% (18/43), 14% (6/43) and 11.6% (5/43, respectively. The live birth rate after in vitro fertilization was 30.2% (13/43) in the hysteroscopy group and 9.5% (4/42) in the control group (p<0.05). There were no significant differences in the IVF outcomes such as the biochemical pregnancy rate and miscarriage rate between the hysteroscopy and control groups. ConclusionThis study demonstrates that hysteroscopic metroplasty improves pregnancy and live birth rates for women with a history of RIF and dysmorphic uterus. Correction of T-shaped uterus with hysteroscopic metroplasty ensured favorable reproductive outcomes in RIF cases. However, conclusions must be taken carefully as this is an observational study. A prospective, randomized and controlled study is necessary to support these results.

Prevalence of peripheral blood natural killer cells ≥12% in women with recurrent implantation failure: study carried out in a private clinic of Mexico City

Objective: To report the incidence of Natural killer cells peripheral blood (pNK) ≥12% and his relationship with Recurrent Implantation Failure (RIF) in a population of Mexican women. Methods: Retrospective, cross sectional and observational study that included 74 women with history of RIF, which were subjected to a study to evaluate pNK cell levels in order to avoid complications in future pregnancies. 2 groups were formed: 1) Control and 2) RIF: women with a history of implantation failure and primary infertility (with no history of pregnancies or abortions). Results: Women in RIF were younger and had higher BMI, compared to the Control. Prevalence of patients with pNK cells ≥12% was statistically higher in RIF than in Control (66.6% vs 20%). RIF showed a significative increase of pNK≥1 cell concentration, compared to Control (12.9±4.6 vs 9.5±0.6, p=0.00). When only women with results of pNK ≥12% were evaluated, RIF showed a level numerically higher than that of the Control (16±1.8 vs 13.5±0.8). Conclusion: RIF may be the result of increased pNK concentrations and as observed in this study, slightly more than 60% of the Mexican population could be susceptible to abortions.

History of Recurrent Implantation Failure is Associated With the Incidence of Adverse Perinatal Outcomes in Singleton Live Births Following Frozen-Thawed Embryo Transfer Cycles.

ObjectiveTo investigate whether patients with a history of recurrent implantation failure (RIF) are associated with adverse perinatal outcomes in singleton live births following frozen-thawed embryo transfer (FET) cycles.DesignRetrospective cohort study.MethodsThis study analyzed the obstetric and neonatal outcomes of patients with and without a history of RIF who underwent FET cycles in a single reproductive center between January 2017 and October 2020. A total of 1,100 women with singleton live births beyond 28 weeks of gestation were included. The primary outcome measures were perinatal outcomes, especially gestational age, birthweight, preterm birth (PTB), large for gestational age (LGA), small for gestational age (SGA), congenital malformation rates, and premature rupture of the membranes (PROM). Multiple logistic regression was used to establish relationships between RIF and adverse perinatal outcomes after adjusting for relevant baseline demographics and cycle characteristics.Result(s)The RIF group showed a preferred transfer of two embryos and cleavage embryos compared with the control group (P <0.05). Regarding perinatal outcomes in singleton deliveries, women with RIF had increased rates of LBW (adjusted odds ratio [aOR] 2.027; 95% confidence interval [CI], 1.025–4.009), PTB (aOR 1.785; 95% CI, 1.050–3.036), and PROM (aOR 2.259; 95% CI, 1.142–4.467). The incidence of congenital malformations was similar between the two groups (4.1% vs. 2.4%; P = 0.759). Furthermore, multiple intrauterine procedures were associated with a statistically significant increased risk of PROM in RIF patients (aOR 1.537; 95% CI, 1.105–2.137).ConclusionsWomen with a history of RIF were associated with an increased risk of LBW, PTB, and PROM in singleton live births after FET cycles. In addition, multiple intrauterine procedures were independent risk factors for PROM.