Undergoing IVF Cycles Research Articles

Use of tamoxifene-controlled ovarian hyperstimulation for fertility preservation before breast cancer treatment: A prospective cohort study with a 5-year follow-up

PurposeFertility issues are of great concern for young women undergoing treatment for breast cancer (BC). Fertility preservation (FP) protocols using controlled ovarian stimulation (COS) with letrozole have been widely used with overall good results. However, letrozole cannot be used in every country in this context. This study aimed to assess the efficacy of tamoxifen for COS in women with early BC undergoing FP. MethodsThis multicentric prospective study included patients aged 18–40, diagnosed with stage I, II and III invasive BC, undergoing tamoxifen-COS before adjuvant or neoadjuvant chemotherapy (NAC). The primary endpoint was the efficacy of tamoxifen-COS protocol evaluated by the number of oocytes collected and vitrified. Secondary endpoints included the time interval before chemotherapy, breast cancer (BC) recurrence rates, and reproductive outcomes. ResultsNinety-five patients were included between 2014 and 2017, aged 31.5 ± 4 years on average. 37.9 % received NAC and 62.1 % received adjuvant chemotherapy. FP procedure was successful in 89.5 % of the cycles. The mean number of collected and vitrified oocytes was 12.8 ± 7.9 and 9.8 ± 6.2, respectively. The mean duration of COS was 10.4 ± 1.9 days. Median time before chemotherapy initiation was 3.6 weeks (IQR 3.1; 4.1) for women receiving NAC. Five-year relapse-free and overall survival rates were in-line with those expected in this population. Twenty-one women had spontaneous full-term pregnancies, while 5 underwent IVF cycles with frozen-thawed oocytes, without pregnancy. ConclusionTamoxifen-COS protocols appear to be feasible before adjuvant or NAC treatment in young BC patients and efficient in terms of oocyte yield.

P-525 Is there an impact of chromosomal structural rearrangements on IVF laboratory outcomes in PGT-SR cycles? a propensity score matching-based study

Abstract Study question Are IVF laboratory outcomes altered in PGT-SR cycles, and how are these outcomes affected by the type of rearrangement and the carrier’s gender? Summary answer PGT-SR cycles showed a decrease in overall blastulation and top-quality blastocyst rates compared to PGT-A cycles. Transferable blastocyst rate correlates with chromosomal structural rearrangement type. What is known already Chromosomal structural rearrangements (SR) are associated with altered gametogenesis, resulting in the formation of gametes with chromosomal imbalances. For affected couples, undergoing IVF cycles with PGT-SR is recommended to select embryos with a normal or balanced karyotype. Scarce data exist on IVF laboratory outcomes in couples undergoing PGT-SR cycles. Findings regarding blastocyst rate, mainly focused on high-quality blastocysts, remain unclear. In addition, structural rearrangement carriers yield fewer transferable blastocysts than patients undergoing PGT-A. Conclusive evidence regarding the influence of carrier gender and type of chromosomal SR on the development of transferable blastocysts is lacking, with existing studies yielding contradictory results. Study design, size, duration This retrospective cohort study was conducted at a single university-affiliated obstetrics and fertility center between January 2019 and November 2023. The analysis involved 558 patients undergoing IVF cycles, including 136 in PGT-SR and 422 in PGT-A cycles. Participants/materials, setting, methods Trophectoderm biopsies from day 5/6/7 blastocysts underwent comprehensive chromosome screening utilizing validated NGS. Propensity Score Matching with a 1:1 match ratio was performed to balance baseline characteristics between the PGT-SR and PGT-A groups. The impact of chromosomal structural rearrangements on laboratory outcomes was assessed using a logistic regression model. Subsequently, within the PGT-SR population, the association between the carrier’s gender and the transferable blastocyst rate was evaluated through logistic regression, adjusting for potential confounders. Main results and the role of chance There was no significant difference in the fertilization rate between the PGT-A and PGT-SR groups (OR = 1.26, 95% CI 1.01-1.60, p-value=0.14). However, the PGT-SR group exhibited a significantly lower overall blastulation rate (36.7%) compared to the PGT-A group (42.8%) (OR = 0.76, 95% CI 0.63-0.93, p-value=0.03). Notably, a significant difference was observed in the top-quality blastocyst rate, with PGT-A patients showing a rate of 21.7% versus 9.57% in PGT-SR patients (OR = 0.45, 95% CI 0.29-0.69, p-value<0.01). In the PGT-SR cohort, female carriers constituted 44.1%, while male carriers were 56.6%. Reciprocal translocations represented 56.5% of the structural rearrangements, with the remaining 43.5% being other types. When comparing female to male carriers, no significant association was observed between the transferable blastocyst rate and the carrier’s gender (OR = 0.90, 95% CI = 0.35-2.31, p-value=0.83). However, a notable association was identified between the transferable blastocyst rate and the type of structural chromosomal rearrangements (OR = 0.16, 95% CI = 0.09-0.29, p-value<0.01). Specifically, in cases where the patient carried a chromosomal SR other than reciprocal translocation, the transferable blastocyst rate was 83.2%, in contrast to 44.9% in patients with reciprocal translocations. Limitations, reasons for caution The retrospective design of this study represents a limitation, and the limited sample size of patients with structural chromosomal rearrangements restricts the statistical power. Wider implications of the findings These findings reveal that patients with SR yield fewer embryos for biopsy than those undergoing PGT-A, showing a reduced rate of transferable blastocysts in cases of reciprocal translocations. Consequently, providing comprehensive counseling and managing expectations for couples undergoing PGT-SR is essential. Trial registration number not applicable

O-212 A need to standardize: the use and posterior recall of toxic consumables could be avoided with the appropriate testing protocols

Abstract Study question Could the recall of toxic products have been avoided if the samples had been tested with a more sensitive Mouse Embryo Assay (MEA)? Summary answer Standardizing MEA protocols with sensitive methodologies could prevent the potential harm imparted on human embryos and avoid the need to recall initially unidentified toxic products. What is known already Dozens of consumables are used during an IVF cycle, with a potential effect on gametes/embryos in culture. Bioassays, such as the MEA, are commonly used to pinpoint embryo-toxic products/batches before their release. No strict regulations exist on how to perform these tests, although several variables have been described to play a major role on the assay’s sensitivity and its capacity to detect embryo-toxicity. Toxic products/batches that are not identified may need to be recalled. However, recalls are typically issued after the products/batches are suspected of hindering embryo development in clinical laboratories, with the loss of irreplaceable biological samples. Study design, size, duration Only in 2023, four important batch recalls have been issued by regulatory bodies or manufacturers, concerning such diverse products as oil, media, capillaries, or follicle aspiration needles (FAN). Reportedly, thousands of couples undergoing IVF cycles may have been affected by this undetected toxicity in samples that were already MEA-tested by the manufacturers or external testing laboratories, following an undisclosed methodology. Eight of these lots (2xmedia, 3xcapillaries, 2xFAN) were retested with a standardized sensitive MEA protocol. Participants/materials, setting, methods The samples were retested following a high-sensitivity and previously published protocol, using 1-cell stage embryos from a sensitive hybrid mouse strain (B6CBAF1), supplementing with 5mg/ml HSA, and maintaining a low embryo density per droplet. A total of 462 embryos were cultured, allocated to the different test groups (21 + 42 embryos for Medium 1 and 2; 63 + 84+147 embryos for Capillaries 1, 2 and 3; 42 + 42 embryos for FANs 1 and 2, respectively). Main results and the role of chance Embryos were cultured at 37.3oC, 6%CO2 and 7%O2 for 5 days (96h). The expanded blastocyst formation rate (BFR) in each group was assessed as the end-point of the assays, and an 80% was considered as the toxicity-marking threshold. When cultured in Medium 1 and Medium 2, supplemented with 5mg/ml HSA, only 0/21 (0%) and 7/63 (11.1%) of embryos reached the expanded blastocyst stage, respectively. Capillaries and FANs were tested with an indirect method, in which the samples were first incubated with a small volume of control culture medium, and the extract was used to culture the embryos. With this approach, Capillaries 1, 2 and 3 yielded a 23.8% (15/63), 22.6% (19/84) and 0.7% (1/147) BFR, respectively. Similarly, when testing FANs 1 and 2, only 66.7% (28/42) and 19.1% (8/42) of the embryos were able to form a blastocyst. Unfortunately, the recalled oil batches were not available for retest. In parallel to each test, a control group was cultured with standard culture medium without contact with any of the samples; the BFR in all control groups was above 80%. These surprisingly low results suggest that toxicity was present in these samples in high concentrations, evidencing the need for their recall. Limitations, reasons for caution The methodology used in this project successfully identified all the recalled samples as embryo-toxic. However, the same protocol might not prove equally sensitive when testing other products, as the sensitivity of the MEA has been shown to be highly dependent on numerous external factors and to vary between different toxins. Wider implications of the findings Recent events could lead to the belief that the MEA is an insensitive bioassay to assess embryo-toxicity. Far from true, attention should be paid to the specific methodology used in each case. The lack of regulations and standardization translates into huge differences in sensitivity, even leading to completely contrary results. Trial registration number not applicable

P-196 Early blastocyst formation on day 4 is associated with improved clinical outcome and advocates for embryo quailty assessment on day 4

Abstract Study question Is early cavitation on day 4 associated with improved clinical outcome? Summary answer Early blastulation on day 4 is a positive predictor of implantation after single blastocyst transfer on day 5. What is known already In order to select the blastocyst with the highest implantation potential for transfer or freezing, several morphological and/or morphokinetic parameters have been proposed. For instance, early compaction on day 3 (D3) or blastocyst expansion on day 5 (D5) have been reported to be relevant when selecting embryos for transfer. However, day 4 has often been overlooked and very few studies focused on early blastulation on day 4 (D4). Indeed, data on early blastulation on day 4 is scarce. It was however recently demonstrated that embryos transferred on day 4 could provide acceptable pregnancy rates. Study design, size, duration This monocentric retrospective cohort study was conducted in all consecutive couples undergoing IVF cycles in 2017-2022 with single blastocyst transfer and observation on day 4. Participants/materials, setting, methods Morphological and morphokinetic assessment was performed by 2 operators at 93±2h, 116±2h (and 140±2h) post insemination, on day 4, 5 (and 6) respectively. A total of 26 507 D4 embryos were included in this study and grouped according to the observation of early cavitation or not : 23 244 were in the D4 control group (absence of early cavitation on D4) and 3263 were in the D4 blastocyst group (observation of early cavitation on D4). Main results and the role of chance The proportion of embryos used clinically for either transfer or freezing on day 5 or 6 was significantly higher in the D4 blastocyst group than in D4 control group (86.7% vs 50.1% respectively, p < 0.05). Concerning pregnancy rate, cinical pregnancy rate was significantly higher in the D4 blastocyst group than in D4 control group (p < 0.05). In the D4 control group, 5051 embryos were transferred and clinical pregnancy rate is 28.7%. In the D4 blastocyst group, 1286 embryos were transferred (fresh or frozen) and clinical pregnancy rate is 40.2%. Limitations, reasons for caution The retrospective and monocentric design calls for caution when considering the generalizability of our findings. Wider implications of the findings We postulate that D4 embryo quality assessment should be widely implemented in any blastocyst program in order to improve embryo quality assessment and subsequently clinical outcome. Trial registration number not applicable

O-255 Ovarian Response Prediction Index (ORPI) as a predictor tool for ovarian response and clinical pregnancy in IVF/ICSI cycle : A Prospective Cohort Study

Abstract Study question Can incorporation of the Ovarian Response Prediction Index (ORPI), integrating AMH, AFC and age, offer a reliable prognostication of ovarian stimulation response in ART cycles? Summary answer The study affirms that ORPI, a three-variable index, emerges as a robust predictor for both ovarian response and clinical pregnancy rates within IVF/ICSI cycles. What is known already In the realm of individualized Controlled Ovarian Stimulation, precision in forecasting ovarian response proves pivotal. Ovarian reserve, influenced by age, AMH, and AFC, exhibits considerable heterogeneity among women of analogous age. While AMH and AFC function as pivotal markers, a holistic approach, exemplified by ORPI, stands out as a more comprehensive tool for predicting ovarian response. Study design, size, duration This prospective cohort study enrolled 302 subjects undergoing IVF cycles at a tertiary infertility center spanning from 1st January 2022 to 31st December 2023. Rigorous inclusion criteria mandated age ≤38 years, a BMI between 20–25 kg/m2, regular menstrual cycles, intact bilateral ovaries, no history of ovarian surgeries, absence of severe endometriosis, and no evidence of endocrine disorders. Participants/materials, setting, methods Subjects adhering to the inclusion criteria underwent venous blood sampling for AMH measurement and transvaginal ultrasound for AFC assessment. The ORPI was meticulously computed as (AMH × AFC) / patient age. Key endpoints encompassed the total oocytes retrieved, mature MII oocytes, and occurrences of clinical pregnancy. Rigorous statistical methodologies were employed for robust analysis. Main results and the role of chance The mean values for age (years), AMH (ng/mL), ORPI, AFC, and fertilized oocytes were 31.75 ± 4, 3.75 ± 3.39, 1.06 ± 1.22, 7.1 ± 2.94, and 6.11 ± 3.26, respectively. A highly significant positive correlation (p-value < 0.0001) was observed between ORPI and the numbers of oocytes and MII oocytes, with correlation coefficients of 0.714 and 0.746, respectively. Interpretation of the area under the ROC curve revealed that ORPI was a significant predictor of obtaining ≥4 oocytes at a cutoff point of > 0.2, with an area under the curve of 0.907. Similarly, for MII oocytes ≥4, ORPI was a significant predictor at a cutoff point of > 0.2, with an area under the curve of 0.937. In the case of positive clinical pregnancy, ORPI emerged as a significant predictor at a cutoff point of > 0.75, with an area under the curve of 0.822. The study underscored a substantial positive correlation between ORPI and both oocyte and MII oocyte counts, affirming ORPI’s pivotal role as a predictor for clinical pregnancy. Limitations, reasons for caution Limitations due to the constraint of sample size temper the generalizability of findings. The study did not account for live birth rates, and a comparative analysis with alternative markers was omitted. A more expansive, meticulously designed study is imperative. Wider implications of the findings ORPI, an innovative ovarian response biomarker, showcases considerable potential for predicting both ovarian response and clinical pregnancy rates. Its integration into individualized Controlled Ovarian Stimulation regimens holds promise for augmenting the counselling and prognostication process for patients embarking on IVF/ICSI cycles. Trial registration number MCDH/2023/28

Expression of miRNA from spent pre-implantation embryos culture media

This study examines the expression of three microRNAs (hsa-miR-661, hsa-miR-21–5p, hsa-miR-372–5p) in spent pre-implantation embryos culture media to identify possible new non-invasive biomarkers of embryo competence, predictive of development to the blastocyst stage. A preliminary analysis on 16 patients undergoing IVF cycles was performed by collecting and stored spent culture media on the fifth/sixth day of embryo culture. Expression of miRNAs was evaluated according to the embryos’ fate: 1) NE/DG: non-evolved or degenerate embryos; 2) BLOK: embryos developed to the blastocyst stage. Preliminary results revealed a higher miRNAs expression in NE/DG spent media. To elucidate the roles of these miRNAs, we employed a robust bioinformatics pipeline involving: 1) in-silico miRNA Target Prediction using RNAHybrid, which identified the most-likely gene targets; 2) Construction of a Protein-Protein Interaction network via GeneMania, linking genes with significant biological correlations; 3) application of modularity-based clustering with the gLay app in Cytoscape, resulting in three size-adapted subnets for focused analysis; 4) Enrichment Analysis to discern the biological pathways influenced by the miRNAs. Our bioinformatics analysis revealed that hsa-miR-661 was closely associated with pathways regulating cell shape and morphogenesis of the epithelial sheet. These data suggest the potential use of certain miRNAs to identify embryos with a higher likelihood of developing to the blastocyst stage. Further analysis will be necessary to explore the reproducibility of these findings and to understand if miRNAs here investigated can be used as biomarkers for embryo selection before implantation into the uterus or if they may be reliable predictors of IVF outcome.

Association between blood lead levels and unfavorable IVF outcomes: potential involvement of endoplasmic reticulum stress response in granulosa cells.

To investigate the relationship between blood lead levels (BLLs) and IVF clinical outcomes in infertile females and to further explore the possible involvement of granulosa cell (GC) endoplasmic reticulum (ER) stress in the process. One hundred twenty-three infertile women undergoing IVF cycles were included in the current study. All participants were divided into three (low, medium, and high) groups determined by BLL tertiles. Gonadotropin releasing hormone (GnRH) agonist regimen for ovarian stimulation was used for all patients, with follicular fluids being collected on the day of oocyte retrieval. Lactate dehydrogenase (LDH) levels in follicular fluid and the endoplasmic reticulum stress-signaling pathway of granulosa cells (GCs) were examined. The oocyte maturation rate and high-quality embryo rate on cleaved stage decreased significantly as BLL increased. For lead levels from low to high, live birth rate (68.29%, 56.10%, 39.02%; P=0.028) showed negative correlations with BLLs. Also, follicular fluid Pb level and LDH level was significantly higher in the high lead group versus the low group. Binomial regression analysis revealed significant negative correlation between BLLs and live birth rate (adjusted OR, 0.38; 95% CI, 0.15-0.95, P=0.038). Further analysis of the endoplasmic reticulum stress (ER stress) signaling pathway of GCs found that expressions of GRP78, total JNK, phosphorylated JNK, and CHOP increased and BCL-2 decreased with increasing BLLs. BLLs are negatively associated with final clinical outcomes in IVF patients that may be related to increased ER stress response and GC apoptosis. Thus, reducing Pb exposure before IVF procedures may improve final success rates.

P-236 IL-6 levels in follicular-fluid and day5/6 spent culture drops indicate its role as a biomarker of oocyte maturation and an embryokine influencing embryo developmental competence

Abstract Study question To elucidate the role of IL-6 in folliculogenesis and embryo development during human in-vitro culture Summary answer High IL-6 levels in follicular-fluid positively correlate with oocyte maturity status whereas higher IL-6 secretome in day5/day6 spent culture media inversely correlates with blastocyst/ICM grades What is known already Although some studies have attempted to explore the correlation of FF IL-6 with fertilization and embryo development rates; the results have been inconclusive. Similarly, studies comparing FF IL-6 levels with endometrial receptivity and pregnancy outcomes have been ambiguous and contradictory. Recent research has reported improvement in bovine blastocyst ICM grades in culture media supplemented with IL-6. No study has yet evaluated the role of IL-6 as a biomarker of oocyte maturity nor investigated IL-6 as an embryokine that could regulate embryo developmental competence in humans. We have employed the FF metabolomic and Secretomic approach for IL-6 in human IVF cycles. Study design, size, duration Prospective study in women (25-35 years) undergoing IVF cycles (n = 256) for unexplained/tubal-factor infertility from 2018-2020. Only those women whose FF and day5/6 spent culture media sample could be obtained, were included. Women with endometriosis, PID, Genital Kochs were excluded. IL-6 levels were measured by diagnostic kit in spent micro-drops and FF (pooled per patient per cycle). Cycles were divided into Low and High IL-6 groups (FF: ≤ 47.5 and >47.5pg/ml; Spent Medium: <0.5 and ≥0.5pg/ml). Participants/materials, setting, methods Antagonist cycles involving COH with r-FSH and ICSI for all. Noted oocyte Nuclear maturity (GV/MI/MII). Oocyte Cytoplasmic maturity graded as Top and non-Top, based on presence of granulation, refractile bodies, sER, visco-elasticity. Post ICSI, oocytes cultured individually in 50µl single-step media micro-drops till day3 and shifted to fresh 50µl micro-drops for extended culture till day5/6. Blastocyst gradation was done using Gardner’s classification for expansion, ICM and trophectodermal cells. Statistical analysis done using graph-pad prism VI Main results and the role of chance High FF/IL-6 group (n = 103) had significantly higher proportion of MII (92.08 vs. 76.33%; p < 0.001) and top-quality (88.10 vs. 70.0%; p < 0.001) oocytes and fertilization rates (89.45 vs. 77.30%; p = 0.009) compared to Low FF/IL-6 group (n = 153) although total number of days of stimulation/gonadotropin dose, mean oocytes retrieved did not differ significantly between the two groups. Cleavage (84.64 vs. 75.92%; p = 0.061) and blastocyst formation (53.86 vs. 42.37%; p = 0.053) rate, although higher, did not differ significantly between the two FF/IL-6 groups. FF/IL-6 level correlated strongly with overall oocyte maturity status (Pearson r = 0.61, 95% CI 0.52-0.8). Threshold FF/IL-6 level for obtaining top-quality MII oocytes was >39.2pg/ml (ROCAUC = 86%, Sensitivity = 78%, Specificity = 83%). In D5/6 spent culture drops, likelihood of top-quality blastocyst formation (expansion grade ≥3 and ICM/TE grades AA/AB/BA) increased by 41% (odds ratio: 0.44, 95%CI: 0.4-0.5; χ2 p < 0.0001) if IL-6 concentration was <0.5 pg/ml. Lower culture-drop IL-6 levels correlated strongly with top ICM grades (Pearson r = −0.66, 95% CI 0.6-0.7). Highest IL-6 levels were found in spent culture-drops containing grade-C blastocysts whether on day5 or day6. IL-6 levels were comparatively higher in day6 than in day5 micro-drops, correlating with higher percentage of top-quality blastocysts on day5 than on day6. Threshold micro-drop IL-6 level for top-grade blastocyst <0.42 pg/ml. Limitations, reasons for caution We estimated IL-6 levels in pooled FF (per patient, per cycle). Although cumbersome & time-consuming, it would be worthwhile to evaluate individual FF (per follicle, per patient, per cycle). Also, measuring IL-6 in spent media involving group embryo culture, may facilitate study of the probable paracrine effects of this ‘embryokine’. Wider implications of the findings IL-6 is a pleiotropic cytokine and its role/s in folliculogenesis, embryo development and endometrial receptivity is not yet clearly etched out. This study attempts to elucidate the functions of IL-6 in a systematic manner by deriving threshold values for obtaining top quality oocytes and blastocysts in human in-vitro culture conditions. Trial registration number NOT APPLICABLE

P-095 Increased sperm high DNA stainability is associated with a diminished chance of fertilization with IVF cycles: a retrospective study using a propensity score-matching analysis

Abstract Study question To investigate the association of high DNA stainability (HDS) with fertilization, embryonic development and clinical outcome after IVF treatment. Summary answer Increased HDS levels might be associated with decreased fertilization of IVF cycle. What is known already The HDS parameter of sperm suggests loosening and weakening of chromatin condensation structure resulting from the lack of full histone-to-protamine exchange. Elevated HDS may negatively correlate with sperm quality, including sperm morphology and concentration. Study design, size, duration Couples that underwent IVF cycles from January 2016 to December 2020 were retrospectively studied, including a total of 2604 target cycles after IVF treatment consisting of 676 cycles in the HDS > 15% group and 1928 cycles in the HDS ≤ 15% group. In addition, 548 cycles undergoing fresh IVF-embryo transplantation treatment consisting of 151 cycles in the HDS > 15% group and 397 in the HDS ≤ 15% group were included. Participants/materials, setting, methods HDS was assessed by sperm chromatin structure assay method. Couples were classified into HDS > 15% group and HDS ≤ 15% group based on the HDS thresholds recommended by the manufacturer’s instructions. After controlling the bias between groups by the propensity score-matching method, the Mann-Whitney U test or Chi-squared test was used to evaluate the differences between groups. Regression analysis was used to assess the effect of HDS levels on fertilization, embryo quality, and clinical outcome of IVF cycles Main results and the role of chance No significant differences were observed between the HDS > 15% group and HDS ≤ 15% group regarding the number of fertilized oocytes (10.00 [6.00, 15.00] vs. 10.00 [6.00, 15.00], P = 0.230), the number of 2 pronuclei zygotes (8.00 [4.00, 12.00] vs. 8.00 [5.00, 12.00], P = 0.311), the number of cleaved zygotes (10.00 [5.00, 15.00] vs. 10.00 [6.00, 15.00], P = 0.301), and the number of high-quality embryos on day 3 (5.00 [2.00, 9.00] vs. 5.00 [3.00, 9.00], P = 0.729). However, linear regression analysis showed that HDS levels negatively impacted fertilization (adjusted B value = −0.015, 95% CI: −0.028 ∼ −0.002, P = 0.022) and 2 pronuclei formation (adjusted B value = −0.015, 95% CI: −0.029 ∼ 0.000, P = 0.046). The implantation rate (47.1% vs. 50.2%, P = 0.418), clinical pregnancy rate (60.9% vs. 64.5%, P = 0.440), ongoing pregnancy rate (83.7% vs. 86.7%, P = 0.475), early miscarriage rate (16.3% vs. 13.3%, P = 0.475), late miscarriage rate (1.1% vs. 3.5%, P = 0.232), and live birth rate (82.6% vs. 83.2%, P = 0.896) were not significantly different between groups. Linear regression analysis showed that HDS did not affect the implantation rate. Binary logistic regression analysis also showed that HDS did not affect clinical pregnancy, ongoing pregnancy, early miscarriage, late miscarriage, and live birth. Limitations, reasons for caution The main weakness of this study was the nature of the retrospective design, and additionally, patients were not randomized. Wider implications of the findings Although our results are not entirely new, it can be considered as reliable because 2604 IVF cycles and 548 fresh IVF-ET cycles were included and subsequently controlled the bias between groups This study suggested that HDS might be an indicator in predicting the fertilization ability of sperm in IVF treatment. Trial registration number not applicable

P-365 The composition of the endometrial microbiota, as determined by MicroBioMap®, significantly affects clinical outcomes in assisted reproduction treatments

Abstract Study question How does the presence of specific pathogenic or health-promoting microbial species in the endometrium relate to female fertility in assisted reproduction treatments (ART)? Summary answer Detection of endometrial pathogens using MicroBioMap® is associated with poor ART outcomes, whereas sterile endometria show the highest pregnancy and live birth rates. What is known already Vaginal eubiosis is characterised by low microbial diversity and dominance of Lactobacillus, which displays bioregulatory functions and prevents the growth of pathogenic microorganisms. Vaginal dysbiosis is directly linked to infertility and poor pregnancy outcomes. However, the endometrial microbiota composition and its role in fertility are more controversial. Robust reports show that chronic endometritis (CE) is highly prevalent among infertile patients, whose outcomes improve after antibiotic treatment. However, the role of Lactobacillus in this compartment, whether it also displays bioregulatory functions, or if it is essential and beneficial for women undergoing ART, is still unclear. Study design, size, duration This is a case-control retrospective study analysing the endometrial microbiota in the window of implantation (WOI) and the ART outcomes of 456 women undergoing IVF cycles in egg donation regimes with/without PGT-A (177 and 163 respectively), or using their own eggs in cycles with/without PGT-A (91 and 25 respectively), between March 2016 and April 2021 in IVF-Life Alicante (Spain). ART results after single embryo transfer (sET) of patients with different endometrial microbiota profiles were compared. Participants/materials, setting, methods Presence of 18 microbial CE pathogens and 4 Lactobacillus species was determined by high-throughput qPCR (MicroBioMap®) in 456 endometrial samples obtained during the WOI (determined by ER Map® receptivity test) of a hormone replacement therapy cycle. Clinical outcomes (clinical pregnancy, CP; live birth, LB) of sETs performed during the WOI (guided by ER Map®) were compared in relation to the endometrial microbiota profile. In PGT-A cycles, embryo ploidy was determined by NGS (VeriSeq-MiSeq) before sET. Main results and the role of chance According to the microbial profile detected by MicroBioMap®, patients were categorised as i) Lactobacillus only (L, n = 199), ii) Pathogens only (P, n = 56), iii) Lactobacillus + pathogens (L+P, n = 59) or iv) Undetected (U, n = 142). Absence of endometrial microorganisms was significantly associated to ART success. U-patients showed significantly higher CP (64.2% vs 50.8%, Fisheŕs exact p=0.046) and LB rates (52.9% vs 37.2%, p=0.03) after euploid sET than patients with microorganisms. These differences were more pronounced in patients receiving donor eggs (CP 70.2% vs 50.8%, p=0.026; LB 59.5% vs 40.2%, p=0.046). Presence of Lactobacillus in isolation in the endometrium showed a detrimental effect compared to U-environments. A trend towards statistical significance was detected, with L-patients showing lower LB rates compared to U-patients after euploid sET (38.8% vs 52.9%; p = 0.084). Detection of endometrial pathogens significantly reduced ART success rates. P-patients had significantly lower CP rates (64.2% vs 41.2%, p=0.038) after euploid sET than U-patients, and significantly lower CP (70.2% vs 36%, p=0.006) and LB rates (59.5% vs 21.7%, p=0.004) if receiving donor eggs. Interestingly, Lactobacillus dominance in pathogen-positive patients seemed to improve outcomes after euploid sET compared to pathogen-dominated patients (CP 56.7% vs 42.2%; LB 41.7% vs 31%). Limitations, reasons for caution This is a retrospective study. Randomised-control trials, non-selection studies and/or other investigations including antimicrobial treatments are needed to confirm these results and the extent of any clinical benefits. Endometrial biopsies were obtained transcervically. Although utmost care was taken, risk of contamination from the lower genital tract cannot be completely excluded. Wider implications of the findings Our results suggest that sterile endometria provide the best conditions for embryo implantation and pregnancy. Lactobacillus presence in endometrium might not be essential for pregnancy, and could even be detrimental if found in isolation. Presence of Lactobacillus as a bioregulator might only be beneficial if other pathogens co-colonise the tissue. Trial registration number Not applicable

General anesthesia with propofol during oocyte retrieval and in vitro fertilization outcomes: retrospective cohort study

General anesthesia is frequently administered during oocyte retrieval. Its effects on the outcomes of IVF cycles are uncertain. This study investigated whether administration of general anesthesia (specifically propofol) during oocyte retrieval affects IVF outcomes. A total of 245 women undergoing IVF cycles were included in this retrospective cohort study. IVF outcomes of 129 women who underwent oocyte retrieval under propofol anesthesia and 116 without anesthesia were compared. Data were adjusted for age, BMI, estradiol on triggering day and total gonadotropin dose. The primary outcomes were fertilization, pregnancy and live birth rates. A secondary outcome was the efficiency of follicle retrieval associated with the use of anesthesia. Fertilization rate was lower in retrievals under anesthesia compared to without (53.4% ± 34.8 vs. 63.7% ± 33.6, respectively; p = 0.02). There was no significant difference in the ratio of expected to retrieved oocytes between retrievals with and without anesthesia (0.8 ± 0.4 vs. 0.8 ± 0.8, respectively, p = 0.96). The differences in pregnancy and live birth rates between the groups were not statistically significant. General anesthesia administered during oocyte retrieval may have adverse effects on the fertilization potential of oocytes. This impact on the developmental potential of oocytes may lead to negative IVF outcomes and should be investigated further.

8999 Reproductive Outcomes and Overall Prognosis of Patients with Asherman's Syndrome Undergoing IVF Cycles

<h3>Study Objective</h3> To estimate the impact of Asherman's Syndrome (AS) following hysteroscopic adhesiolysis on short term reproductive outcomes and the time to achieve pregnancy in infertile patients during IVF cycles. <h3>Design</h3> Retrospective cohort study. <h3>Setting</h3> Tertiary university affiliated medical center. <h3>Patients or Participants</h3> Fifty-one infertile patients who were treated for Asherman's syndrome (AS) and underwent IVF cycles (study group). Patients were matched with controls in a 1:1 ratio, according to their age and etiology of infertility (control group). <h3>Interventions</h3> The collected date was obtained from medical records and telephonic survey. <h3>Measurements and Main Results</h3> There was no difference in the mean number of embryo transfer per patient between the two groups (4.9 ±4.6 vs. 6.22 ±4.3, p=0.78). The mean endometrial thickness before embryo transfer was significantly higher among women in the control group (8.7 ±1.8 vs. 6.95±1.7, respectively, p=0.001). Women with AS had significantly lower cumulative live birth rate and significantly higher miscarriage rate as compared to women in the control group (23.5% vs 56.8, p=0.001 and 41% vs. 15.6%, p=0.008 accordingly). The mean endometrial thickness was significantly higher among women with AS who had live birth (8.2 (±1.4) 6.9 (±1.2), p=0.001). The Overall time to conception was significantly longer among with AS (407.962 ±74.7 Vs 785.6 <h3>Conclusion</h3> AS has an impact on the reproductive potential in infertile patients following hysteroscopic adhesiolysis. Endometrial thickness was shown to be an important predictor for live birth among women with AS that utilizing IVF.

P-695 Anti-muellerian Hormone (AMH) and Plasminogen Activator Inhibitor (PAI-1) are robust follicular-fluid (FF) biomarkers predictive of follicle maturity and oocyte quality in women undergoing IVF cycles

Abstract Study question Is AMH the in-vivo trigger that correlates with PAI-1 in the follicular-fluid micro-milieu to influence maturity and quality of oocytes derived in vitro during IVF? Summary answer Lower FF-AMH levels correlate with top quality oocytes whereas Lower FF-PAI-1 levels correlate with follicle-maturity as reflected in mature MII oocytes retrieved in IVF cycles What is known already Ovulation is a time-coordinated tightly regulated proteolytic event. PAI-1, primarily produced by granulosa cells, is a major form of PAI in FF of human preovulatory follicles, allowing localized degradation and regulating ovulation.While ovulation process is rendered redundant in IVF cycles; nevertheless FF PAI-1 concentration may influence maturity of oocyte within the follicle. AMH is exclusively produced by granulosa cells of pre-antral &amp; early-antral ovarian follicles in the adult female and secreted into FF. Although AMH is widely accepted as an ovarian-reserve marker, very few studies have explored direct putative correlation of FF-AMH with oocyte quality in stimulated IVF cycles. Study design, size, duration Prospective study of n = 1150 (Power of study &amp;gt;85%) women (mean age 30.22±4.25 years, BMI 23.97±4.53, W/H ratio 0.88±0.06) undergoing antagonist stimulation protocol with r-FSH (September 2015-December 2018). On day of ovum pick-up, original aspirate per follicle was pooled per patient. AMH and PAI-1 were evaluated in pooled FF by ELISA method using diagnostic kits. These protein-biomolecules were expressed as ratio of total protein content. SerumE2, LH, P on d/hCG trigger were also estimated by RadioImmunoAssay. Participants/materials, setting, methods Elderly women (age&amp;gt;35 years), women with polycystic-ovaries, endometriosis were excluded. Embryo-transfer (ET) was done either on Day3 or day5. Luteal-phase was supported with micronized progesterone. All cycles were divided into Low FF-AMH (≤1.72 ng/mg protein), High FF-AMH (&amp;gt;1.72ng/mg protein) and Low FFPAI-1 (≤592.1 ng/mg protein), High FFAI-1 (&amp;gt;592.1 ng/mg protein) groups. Oocyte quality was graded as poor, average and top based on morphological assessment of COC-complex, zona-pellucida (thickness, regularity) and cytoplasm (granularity, vacuolation, refractile bodies). Main results and the role of chance Mean Age, Baseline D3-Serum AMH, E2 d/hCG and number of eggs retrieved did not differ significantly between low vs. High FF AMH groups as well as Low vs. High FF-PAI-1 groups. Low FF AMH group displayed significantly higher percent of top-quality oocytes (70.38±11.5 vs. 52.16±8.3;p&amp;lt;0.0001) and higher fertilization rates (88.68±10.7 vs. 72.5±9.2%; p = 0.004) compared to High FF AMH group. Incidences of total fertilization failure were significantly higher in high FF AMH (3.25% vs. 1.1%, p = 0.0002) compared to Low-FFAMH group. FF-AMH correlated with oocyte-quality (Pearsonr=0.48); lower-levels increased the odds of top-quality oocytes. Thresholds of FF-AMH for top quality oocytes was &amp;lt;1.8 ng/mg protein;ROCAUC:85%) Low FF PAI-1 group had significantly higher Percentage of MII oocytes (93.55 vs. 74.8%; p &amp;lt; 0.0001), Rates of Fertilization (86.35 vs. 71.04, p = 0.008) and cleavage (79.19 vs. 69.85;p=0.03) compared to High FF-PAI-1 group. FF-PAI-1 correlated with oocyte-maturity (Pearsonr=0.51), lower-levels increased the odds of obtaining MII-oocytes. Cutoff of FF PAI-1 for mature MII oocytes was &amp;lt;504.4 ng/mg protein;ROCAUC:76%). FF-AMH showed strong positive correlation with FF-PAI-1 (Pearsonr=0.66) Limitations, reasons for caution Several other molecules in the follicular-fluid milieu may be responsible for maturity status and quality of oocytes. Although these are apparently the most plausible of markers; role of other factors cannot be ruled out. Also, it may be better to study mono-follicular aspirates rather than pooled FF for each patient. Wider implications of the findings FF-levels of AMH and PAI-1 as non-invasive endogenous biomarkers of oocyte maturity and quality in-vitro may facilitate better understanding of the physiological pathway governing in-vivo ovulation process. This study reiterates ‘Follicle-maturation’ is the rate-determining step that sets off a cascade mechanism triggering oocyte-maturation and quality further influencing fertilization, embryo development. Trial registration number In patients with polycystic ovarian syndrome (PCOS), are serum androgen levels (total and free testosterone) associated with the risk of endometrial hyperplasia or cancer?

P-411 Blood concentrations of specific (PCBs) congeners and abortion rates in healthy women undergoing IVF cycles living in areas with high and low pollution. (EcoFoodFertility Project)

Abstract Study question Human exposure to polychlorinated biphenyls (PCBs) exists despite its ceased production since 1977. PCBs congeners harm reproductive health, but data remain inconclusive. Summary answer Evaluate relationship between blood concentrations of specific (PCBs) congeners and abortion rates in women undergoing IVF cycles living in areas with high and low pollution. What is known already PCBs are a class of synthetic chemicals that were widely used in industry until the 1970s and are found in many consumer products as refrigerants in electrical equipment, as flame retardants in plastics production and electronics. Continuous exposure to PCBs is of concern because they fall into the group of endocrine disruptors (EDCs), PCB congeners can spread throughout the female reproductive tract. Indeed, they have been found in human follicle fluids, placenta, amniotic fluid in embryos and fetuses Despite multiple reproductive health effects in women was reported, the results are sometimes inconsistent between experimental and epidemiological studies. Study design, size, duration In the frame of EcoFoodFertility Project, a cross sectional study conducted in Campania Region (Italy) to assess environmental impact on fertility and human health, blood samples were collected during the IVF cycles of the 65 women (aged 27.8 + 3.6) no smokers, BMI 23.5 – 27.9, normal menstruation, in two areas with high and low pollution between 2017 and 2019. The PCBs congeners we investigated were: estrogenic congeners (31,44,52,101,153), anti-estrogenic congeners (77,105,110,114,126,156,169) and other (28,118,138,180). Participants/materials, setting, methods Group A (n = 30) living in low polluted area (LPA), alto Medio Sele in province of Salerno, and group B (n = 35) living in highly polluted area (HPA), known as the “Land of Fires” due to the many sources of pollution. The blood sample was taken on the day of the oocyte pick-up and PCBs were measured by Gas Chromatography Coupled to (tandem) Mass Spectrometry. Statistical analyses were performed by GraphPad Prism 9. Main results and the role of chance Blood concentrations of PCBs are expressed as wet weight (nanograms per gram of serum) and as standardised lipid concentrations (nanograms per gram lipid). The PCB 169 congener was found in the highest concentration, 229.62 ng/g and, on average, accounted for 46.2 % of ΣPCBs tested. The PCB 77 (p &amp;lt; 0.05) and 118 ((p &amp;lt; 0.001) congeners were significantly higher in participants permanently living in HPA (Group B). For the other congeners, no significant variation was found. No significant variations between the two groups for oocyte pick-up rate, metaphase I and II oocyte ratio was found, fertilisation rate and pregnancy rate. Furthermore, the probability of embryo implantation was similar between the two groups, but the abortion rate was 3.0 times higher in group B. The probability of live births is 85.7% in group A and 53.85% in group B.These findings, even if the results between PCBs exposure and reproductive outcomes are due to the different congeners of these chemical compounds and unknown confounding factors and exposure times may be implicated, however suggest an association between the detection of specific PCB congeners and increased abortion rates in women undergoing IVF cycles that cannot be understimated. Limitations, reasons for caution Although we have set up a strict control and survey protocol, our study also has limitations due to the relatively small sample size of the groups and as with almost all epidemiological studies on environmental impact, there may be confounders or co-exposures that could interfere with the observed associations. Wider implications of the findings Although PCBs are no longer produced, the exposure remains widespread due to their long biological half-life, accumulation in the food chain and human health impacts are still reported. Our findings confirm these concerns, therefore, the study of these old chemicals is still relevant and important and should be expanded. Trial registration number not applicable

P-656 Effect of COVID-19 vaccination on clinical outcome in fully vaccinated infertile women undergoing IVF/ICSI cycles at tertiary care centre: prospective observational study

Study questionDoes immune response to COVID-19 vaccination affect the clinical outcome in fully vaccinated infertile women undergoing IVF/ICSI cycles?Summary answerCOVID-19 IgG antibodies are present in follicular fluid post vaccination and higher immune response increases duration of gonadotrophins required and negatively impacts the IVF outcome.What is known alreadyRecent studies assessed the influence of COVID-19 infection and mRNA COVID-19 vaccine on the stimulation cycle characteristics and embryological variables of patients undergoing IVF cycle and found no effect on the IVF outcome in their immediate IVF cycle after recovery, except for a decreased number of top quality embryos. One study reported infection or mRNA vaccine results in rapid formation of anti-COVID IgG which can be detected in follicular fluid. This immune response did not lead to any significant negative effect on ovarian follicular function. There is a possibility that COVID-19 infection might affect numerous fertility-linked proteins.Study design, size, durationProspective observational study, conducted at Division of Reproductive Medicine of tertiary care institute. After taking informed consent, 32 patients who satisfy the inclusion and exclusion criteria with history of receiving two doses of Covishield or Covaxin vaccine with at-least 2 weeks from last dose, were recruited for IVF/ICSI cycles from December 2021 to January 2022, for assessing COVID-19 IgG antibodies in their follicular fluid.Participants/materials, setting, methodsWomen of 21-40 years with normal ovarian reserve and normal uterine cavity were included, those with history of COVID infection were excluded. All patients underwent GnRH antagonist protocol. Follicular fluid was collected at time of oocyte retrieval. After collecting oocytes, 400 microlitre of follicular fluid was stored at -80 and later thawed and analysed for SARS-CoV-2 IgG antibodies (ADVIA Centaur COV2G assay, Germany) which are expressed in index value and reported as reactive (≥1 index).Main results and the role of chanceOut of 32, 21 (65.6%) of the participants had received COVISHIELD (V1)and 11 (34.3%) received COVAXIN (V2). The mean gap between vaccine and the IVF cycle was 84.94 ± 52.65 days. The mean COVID IgG antibody titres (Index) were significantly higher in V1, 28.77±33.50 (0.34 -100), than V2 2.28±3.74(0.05-13.23), p<0.001. Patients with higher antibody titres, required longer duration of ovarian stimulation, rho=0.42, p = 0.017. Patients with higher COVID IgG antibodies were negatively correlated with clinical pregnancy rate (20.9 0± 29.68 vs 4.60 ± 6.28, p = 0.153). The time gap from the last dose of vaccine to IVF cycle had moderate negative correlation with percentage of grade-I embryos out of the total embryos fertilised (%), rho= -0.33, p = 0.068. Furthermore, higher gonadotropins doses were required in patients with high antibody titres, rho=0.25, p = 0.160, and amongst V1 vs V2, total dose of gonadotropins required was 3802.38±742.92 vs 3422.73±564.52, respectively, p=0.115. COVID IgG antibody titres had weak negative correlation with number of grade-I embryos, rho= -0.16, p = 0.396. The time gap from the last dose of vaccine to IVF cycle had a weak negative correlation with number of grade-I embryos, rho=-0.28, p = 0.124.Limitations, reasons for cautionThe main limitation of this study is small sample size. However, the study is currently ongoing, and these are the interim results of the same. As prospective studies with larger sample size would be required to assess the effect of different COVID-19 vaccines in different populations on the IVF outcomes.Wider implications of the findingsThe present study confirms the presence of COVID IgG antibodies in follicular fluid in vaccinated women, and proves that COVISHIELD vaccinated patients had higher antibody titres. Higher antibody titres require longer duration of stimulation and result in poorer outcomes so a longer interval from vaccine to IVF should be recommended.Trial registration numberNA

P-799 To evaluate the effect of Intra-ovarian platelet rich plasma instillation on the clinical outcome of women with diminished ovarian reserve: A prospective interventional study

Abstract Study question Does intra-ovarian instillation of platelet rich plasma (PRP) improves the clinical outcome of IVF cycles in women with Diminished Ovarian reserve (DOR)? Summary answer PRP instillation leads to consistent improvement in Antral follicle count (AFC), thus achieving clinical pregnancy rate of 33.3% per cycle in women with DOR. What is known already There is rising incidence of females with diminished ovarian reserve (DOR) especially among Asian ethnicity. With the emergence of regenerative medicine, multiple studies have evaluated the role of intra-ovarian PRP, demonstrating a beneficial role in improving ovarian reserve parameters (serum Follicular stimulating hormone (FSH), serum anti-Mullerian hormone (AMH), AFC). Despite its’ favorable effects on biochemical markers and AFC, data regarding improvement in clinical outcome remains elusive and led to inception of this study. Study design, size, duration A prospective interventional study was conducted at Division of Reproductive Medicine of a tertiary care institute. 41 infertile females aged 20-39 years with DOR (AMH &amp;lt;1.2 ng /ml; AFC&amp;lt;5) were enrolled in the study during a 6-month period beginning from August 2021. Participants/materials, setting, methods After informed consent, patients received fresh autologous PRP, prepared from 30 ml venous blood. 1.5ml of PRP instilled in each ovarian stroma between day 7-10 of menstrual cycle under sedation. Patients were followed up for three-consecutive months to assess ovarian reserve parameters including serum FSH, AMH and AFC. Patients showing significant improvement in parameters were recruited for fresh IVF cycles using Antagonist protocol with 1% transdermal testosterone. Outcomes were analysed using linear mix effect model. Main results and the role of chance The average platelet concentration in PRP was ∼10,00,000 platelets/µL. The mean age of enrolled patients was 31.22±4.16 years. Linear improvement in AFC (3.63 vs 6.98 vs 7.97 vs 6.90, p&amp;lt;0.001) was observed from baseline to three consecutive follow-up months with maximal response witnessed in second month in 57.1% of those undergoing IVF cycle. However, there was no significant difference in Serum FSH (p=0.11) and AMH (p=0.16) from the baseline post intervention. Of the 41 patients, 35 (85.3%) responded to the treatment and underwent IVF antagonist cycle. 5 out of 35 IVF cycles were cancelled mid-cycle due to poor ovarian response. The mean dose of gonadotropin requirement was 2667.5±281.1 IU (Follicular stimulating hormone) and 1400±337.3 IU (Human menopausal gonadotropin). The average number of oocytes retrieved was 5.7±2.2 whereas mean number of MII oocytes was 4.63±1.85. The fertilization rate and the cleavage rate were 92.4% and 74.1% respectively. Of the thirty patients, eight patients underwent day 2 transfer due to poor grade of embryos. Mean number of grade 1- day 3 embryos was 1.25±0.55 with surplus embryos available for cryopreservation in 14 patients. The overall clinical pregnancy rate per transfer was 33.33%. No adverse events were reported. Limitations, reasons for caution This was a prospective single arm study. A randomized controlled trial comprising a “no-treatment” arm would establish a Level-I evidence. However, “no-treatment” arm in a developing country like ours, imposes financial burden on the couple with no guaranteed clinical success and thus raising ethical concern and need for ovarian rejuvenation. Wider implications of the findings With the impetus to provide a biological child to these DOR women, intra-ovarian PRP instillation as a method of ovarian rejuvenation holds promising results. Evidently, PRP is not only effective in improving ovarian reserve but this translates into an improved reproductive outcome in a population, previously limited to oocyte donation. Trial registration number REF/2022/01/051033

P-460 SARS-CoV-2 infection of human ovarian cells: an in vitro model for the detection of the virus entry into the host cells

Study questionSevere acute respiratory syndrome coronavirus 2 (SARS-CoV-2) can enter and actively infect human follicular and granulosa cells?Summary answerFollicular Granulosa (GCs) and Cumulus cells (CCs) are susceptible to SARS-CoV-2 infection that is able to reproduce.What is known alreadyTo enter host cells, SARS-CoV-2 uses Spike S1 subunit to bind the receptor angiotensin-converting enzyme2 (ACE2), S2 subunit is cleaved by the host transmembrane serine protease 2 (TMPRSS2) or by cathepsin L (CSTL) to produce unlocked, fusion-catalyzing viral forms. CD147 (BSG) has been proposed as an additional host receptor for SARS-CoV-2. Female fertility is strictly dependent on oocyte quality and competence. ACE2 is highly expressed in the human ovaries and in the stromal endometrial cells, as well as in GCs and oocytes. The expression of ACE2 strongly suggests that it is potentially at a high risk of SARS-CoV-2 infection.Study design, size, durationIn order to analyze the presence of host receptors ACE2 and co-receptors TMPRSS2-CSTL and BSG and consequently the susceptibility of GCs and CCs to SARS-CoV2 infection. GCs and CCs were collected from about 25 patients undergoing IVF/ICSI cycles at the UOSA of Assisted Reproductive techniques, from March 2020 to October 2021 at the Center of Couple Sterility, Siena University Hospital.Participants/materials, setting, methodsGCs and CCs were collected from women undergoing IVF cycle. GCs were recovered from the follicular fluid according to the procedure previously published. After oocyte denuding, CCs were isolated and both GCs and CCs co-cultured with SARS-CoV-2 then the supernatant was used to infect VERO6 Cells. Host factors and SARS-CoV2 expression/localization were confirmed by RT-PCR, Western blot and Immunofluorescence. SARS-CoV2 infection and its effect on GCs and CCs were evaluated by Transmission and Immuno-electron microscopy.Main results and the role of chanceqRT-PCR analysis and WB showed that ACE2, TMPRSS2, BSG and CTSL transcripts were expressed in both GCs and CCs cells, even if at different levels. ACE2 transcript was significantly increased in the CCs (0.43 vs 0.15; p < 0.05) with respect to GCs; this datum is very interesting since CCs are the follicular cells able to establish intimate connections with the developing oocyte (e.g. gap junctions). Therefore, contrary to what can be assumed, human CCs cannot act as a barrier against virus entry into the oocyte. Coreceptor BSG and CTSL were the most expressed in GCs with respect to TMPRSS2, (0.7 vs 0.3 and 0.8 vs 0.4 respectively; p < 0.05). Ultrastructural evaluation of human GCs and CCs infected with SARS-CoV-2 was carried out at different time points post-infection (24h, 48h and 72h). Infected human GCs showed cell-associated virus-like particles, virions were approximately spherical, with a diameter outside the lipid bilayer ranging from 50-150 nm and ultrastructural characteristics consistent with those described for other coronaviruses. Complete virions were also observed inside the cytoplasm as single or small groups of particles, either dispersed or within large vesicles. Immunoelectron microscopy confirmed these particles as SARS-CoV2.Limitations, reasons for cautionThis is a human in vitro study, and we cannot predict all the implications in female fertility and related to the oocyteWider implications of the findingsWe provide evidence in favor of SARS-CoV-2 infection in GCs and CCs, the ovarian somatic cells that support oocyte development and competence acquisition. The close relationship between oocytes and follicular cells raises the hypothesis that these cells may represent a vehicle for the oocyte SARS-CoV-2 infectionTrial registration numberNot applicable

P-425 Premature follicular-phase progesterone increase in ART cycles and endometrial receptivity: evaluation of new, more reliable predictors

Abstract Study question Is plasma Progesterone (P) at HCG administration more sensitive in predicting endometrial receptivity in fresh IVF cycles than PFI (progesterone-follicles index) and POI (progesterone-oocytes index)? Summary answer Our data suggest that POI predicts endometrial receptivity defect better than the P assay alone, thus allowing better decision making. What is known already The premature increase in plasma P in the follicular phase of stimulated IVF cycles has been associated with a lower embryo implantation rate because of reduced endometrial receptivity. There is still no consensus on how this impediment to implantation should occur and what threshold should be used. Moreover,since the efficacy of P increase in conditioning reproductive outcomes is more intense in patients with normo- and poor-response and generally inversely proportional to the ovarian response to stimulation, PFI and POI, which better reflect the average amount of P secreted from the follicles, have been suggested as potential additional markers of receptivity. Study design, size, duration Retrospective analysis of fresh IVF-ET cycles. PFI was obtained dividing the plasma P level on the day of hCG administration by the number of follicles ≥14 mm; POI by the number of oocytes retrieved. Pearson correlations, ROC curves, multivariate logistic regression analysis and Cox regression were performed to evaluate the predictive value of PFI and POI for the main outcomes we considered: ongoing pregnancy rate (OPR) and live birth rate (LBR). Participants/materials, setting, methods Women undergoing IVF cycles at San Paolo University Hospital in Milan between 2016 and 2021 were enrolled. As exclusion criteria, we considered previous ovarian surgery, endometriosis, severe male factor and cycle interruption before the embryo transfer. We collected the following data: age of the woman; stimulation protocol; total amount of FSH administered; P and Estradiol (E2) levels and number of follicles &amp;gt;14 mm on induction day; number of oocytes collected; OPR and LBR. Main results and the role of chance A total of 662 fresh embryo transfers (ETs) were performed and 214 pregnancies were achieved. We observed a statistically significant difference in age, number of total embryos and POI between women who became pregnant and those who did not. Specifically, Pearson’s correlation revealed a significant positive correlation between pregnancy and number of total embryos (r = 0.165, p &amp;lt; 0.001), and a significant negative correlation between pregnancy and age (r = 0.201, p &amp;lt; 0.001) and POI (r=-0.096, p = 0.013). Multivariate logistic regression revealed POI to be the most significant predictive parameter of pregnancy, with a odds ratio of 4.374 (p = 0.007, CI: 1.496-12.797). The ROC curve confirmed the highest predictive value of POI for OPR. Finally, Cox regression showed that POI &amp;lt;0.18 was correlated with better OPR (odds ratio=1.419, p = 0.012), suggesting the potential clinical impact of identifying an optimal cut-off of POI to reconsider patient management. Limitations, reasons for caution The results of this study are limited by the small population size. Wider implications of the findings Since the effect on endometrial receptivity of increased plasma P is inversely proportional to ovarian response,we suggest using a marker that accounts for response characteristics. POI has been found to be a reliable index for predicting IVF-ET cycle outcomes,supporting a limit beyond which the fresh embryo transfer should be reconsidered. Trial registration number N/A

Comparative Study of Effectiveness of Yognidra and Antioxidants on Semen Quality in Sub Fertile Male Patients Undergoing IVF Treatment at Wardha Region, India

Background: There are various reasons of infertility in male which can be managed with pharmacotherapy and psychological methods .Physiological and Psychological factors such as low self esteem, stigma, depression anxiety etc which leads to stress contribute to infertility related causes. Yoga (Asanas, Pranayama and Meditation) shows effects on improving psychological factors but no studies are conducted on yognidra related with male infertility. A comparison between yognidra and antioxidant will be efficient in improving quality of semen and can increase ART success rate and Pregnancy incidence by treating psychological and physiological factor.&#x0D; Aim and Objectives: To study the effectiveness of yognidra and antioxidant on semen quality in subfertile male patients undergoing IVF cycles.&#x0D; To sensitize all subfertile males about the study.&#x0D; To assess the effect of yoganidra in subfertile males.&#x0D; To assess the effect of antioxidant in subfertile males.&#x0D; To compare and analyze the semen quality after giving both types of interventions&#x0D; Methodology: 60 infertile patients visiting Infertility clinic Wardha test tube baby centre ,AVBRH, Sawangi Meghe, Wardha will participate in the study. Consent form will be signed by patient. The patients will be divided in to 2 groups of Group number 1 with (30 subjects) antioxidants and Group number 2 will be given (30 subjects) Yognidra. Comprehensive history of patient will be taken in the prescribed format and external genitalia will be clinically examined at Surgery OPD. Study the semen quality and divide according to WHO criteria .Assessment and comparison of semen quality of both the groups before and after intervention .Data of two groups will be analysed and compared.&#x0D; Expected Results: The study shows a comparison between yognidra and antioxidant will be efficient in improving quality of semen and can increase ART success rate and Pregnancy incidence by treating psychological and physiological factor.&#x0D; Conclusion: The current research study will help to understand and explore the utility of yognidra at Infertility clinics and assist practitioner to engage subfertile couples undergoing IVF treatment in the practice of these technique. This study will enhance the quality of support programmes and psychological research. Incorporating psychological intervention provided by yognidra will act as a complementary therapy during infertility treatment.

Myoinositol versus metformin pretreatment in GnRH-antagonist cycle for women with PCOS undergoing IVF: a double-blinded randomized controlled study

Objective To study the effects of myoinositol (Myo) in comparison to metformin (Met), in reducing the risk of OHSS and improving ART outcome in PCOS women undergoing IVF. Design Double-blinded randomized controlled trial (CTRI/2018/05/014196). Setting ART Clinic, AIIMS, New Delhi patients: 102 infertile PCOS women undergoing IVF cycles were enrolled after evaluating for eligibility and allotted as 50 in group 1 (Myo) and 52 in group 2 (Met) after randomization. Interventions Recruited patients received Myo 2 g twice daily (group 1) and Met 850 mg twice daily (group 2). Pre- and post-treatment clinical (menstrual pattern, BMI), hormonal profile (LH, FSH, testosterone, prolactin [PRL], and AMH), biochemical parameters (HOMA IR, fasting glucose, and insulin), ovarian with antral follicle count (AFC) and side effect profile were assessed. After 3 months of therapy, patients were recruited for IVF cycle by antagonist protocol was involving controlled ovarian stimulation, cycle monitoring, oocyte recovery, insemination of oocytes and follow up with fertilization, cleavage, transfer of good grade cleavage embryos, or blastocysts pregnancy outcomes and OHSS incidence and medications was continued until the day of OPU. Main outcome measures Primary outcome was OHSS and clinical pregnancy rate including spontaneous, IVF, and cumulative pregnancy rate including FET. Secondary outcome was ART outcomes and the change in biochemistry and hormonal profile between groups and inter group after medications at 12 weeks. Results Incidence of OHSS (Myo 5 (10.0) (n = 50), Met 10 (20.0) (n = 50) p .07) was not statistically different between groups. Clinical pregnancy rate (Myo 18 (36.0) (n = 50), Met 9 (18.0) (n = 50) p .04) cumulative pregnancy rate including FET (Myo 16 (43.2) (n = 37) vs. Met 10) 22.7) (n = 44) p .05) and spontaneous conception (prior to IVF) Myo 13 (26.0) (n = 50), Met 6 (12.0) (n = 50) p .07) was significantly high in Myo group. No between group difference in ovarian stimulation outcomes including duration and dosage of gonadotropins, E2, P4 levels, number of follicles >14 mm on day of trigger. Number of oocytes retrieved and grade of maturity were similar between groups. Fertilization, cleavage and number of good grade embryos were significantly higher in Myo group. However, implantation rate and number of embryos for freezing were similar between groups. Myo had improvement in fasting insulin, HOMA, Sr.AMH, and SHBG suggesting decreased insulin resistance Conclusions Myo is equally beneficial as Met in reducing the risk of OHSS and has better ART outcome in PCOS women undergoing antagonist cycles.