Embryo Transfer Treatment Research Articles

OP-25 - Do abnormalities in an endometrial microbiota affect the pregnancy outcome of assisted reproductive technology in reproductive immunity?

Recently, the relationship between endometrial microbiota and repeated implantation failure (RIF) has been reported. However, it is still unclear the effect of the microbiota on pregnancy outcome of the patients with the assisted reproductive technology, especially a frozen embryo transfer (FET) treatment. We included 802 cycles with clinical results of pregnancy outcome after FET treatment at our clinic from December 2018 to January 2021. Endometrium fluid was collected before FET and microbiota was examined. We examined the relationship between endometrial microbiota and pregnancy outcome in 463 cycles in which the endometrial microbiome test results were available, pregnancy outcome was known, CD138 negative, and less than 38years (229 cycles were positive pregnancy and 234 cycles were negative pregnancy). Sampling was performed carefully avoiding contamination before FET treatment. Extracted genomic DNA was sequenced for the region of 16 S ribosomal RNA using next-generation sequencer (Endometrial microbiome test, Varinos, Japan). The sequencing data was assigned to bacterial taxonomy and the background-contaminated bacteria were excluded from the microbiome profile. Lactobacillus abundance was calculated. After concordance of patient background, Lactobacillus abundance between successful and unsuccessful pregnancy group was compared. We investigated the effect of microbiota in EF, especially Lactobacillus, for the infertile female with FET as an assisted reproductive treatment with large amount of cycles (802 cycles). First, all of cycles were divided into a successful pregnancy group (305/802) and an unsuccessful pregnant group (497/802), and then the abundance of Lactobacillus was analyzed (the pregnancy ratio was 38.0%). The mean abundance of Lactobacillus in EF was significantly higher in the successful pregnant group than in the unsuccessful pregnant group (70.2% ± 40.5 versus 63.5% ± 43.1, p=0.007). To further refine the conditions of each cycle, patient backgrounds of each cycle were compared. There was a significant difference in age and CD138 results between the successful pregnant group and unsuccessful pregnant group, so the analysis was performed with the limited cycles with CD138-negative and under 38 years (229/463 versus 234/463, respectively). However, even under this strict condition, the mean abundance of Lactobacillus was significantly higher in the pregnant group than in the unsuccessful pregnant group (72.1% ± 39.7 versus 61.1% ± 44.4, p=0.003). These results suggest that the endometrial environment with high Lactobacillus abundance is more conducive to pregnancy in FET treatment. The results of this study suggest that the endometrial microbiota at the time of embryo implantation, especially Lactobacillus genus, is highly relevant to pregnancy outcome, implying that disruption of the intrauterine microbiota may cause infertility. We need further studies to improve the endometrial microbiota of infertile women.

Association between duration of controlled ovarian stimulation and live birth rate in women undergoing In Vitro Fertilization: a SART CORS analysis

Background: In-Vitro Fertilization (IVF) treatment involves synchronization of multiple time-sensitive events, most of which are rate-limiting too. Controlled ovarian stimulation (COS) is one such event. The reproductive outcomes based on the duration of COS (d-COS) in a fresh, IVF embryo transfer (ET) are not well established and therefore, remains largely uncertain. Objective: To evaluate the association between d-COS and live birth rate (LBR) in women undergoing a fresh IVF-ET using autologous oocytes. Methods: A retrospective cohort study was conducted using a US nationwide IVF register – SARTCORS (Society for Assisted Reproductive Technology Clinic Outcomes Reporting System). From a total of 93,889 cycles, we included 56,666 fresh, autologous, IVF - ET treatment cycles from January 2014 through December 2015, with follow-up until October 2016. Adjusted odds and risk ratio with 95% confidence intervals were estimated while controlling for multiple demographic factors and other potential confounders. Variables and outcomes: The primary exposure variable was d-COS defined as the difference in days between gonadotrophin administration and oocyte retrieval. The primary outcome measure was live birth following a fresh IVF-ET. Secondary outcome measures included biochemical pregnancy rate, miscarriage rate, implantation rate and clinical pregnancy rate. Results: A total of 56,666 treatment cycles (mean [SD] age of 33.9 [4.47], BMI of 26.1 [6.02], AMH value of 2.19 [3.37]), and a baseline FSH value of 7.62 [3.49]) underwent a fresh IVF-ET. The LBR after a combined analysis for all ages and all protocols was 44.2 % (n = 25043). In the combined analysis, there was a statistically significant decrease in the live birth rate with LBR with d-COS beyond 10 days. The adjusted OR (95% CI) of LBR for a woman who had 11, 12, 13 and ≥14 days of COS, compared to optimal duration of 10 days was 0.97 (0.87-0.99), 0.94 (0.8-1), 0.83 (0.77-0.89) and 0.73 (0.68-0.79) respectively. The AOR (95% CI) of miscarriage rates for a woman who had 11, 12, 13 and ≥14 days of COS, compared to referent was 1.12 (1-1.26), 0.99 (0.87-1.12), 1.03 (0.90 -1.17) and 1.04 (0.90 - 1.2) respectively. With increasing d-COS, the implantation rate (IR) and clinical pregnancy rate (CPR) also showed a decreasing trend, as with other reproductive outcomes. The RR (95% CI) for implantation rate in a woman who had 11, 12, 13 and ≥14 days of COS, compared to referent was 0.97 (0.93-1), 0.97 (0.93-1.01), 0.91 (0.87-0.95) and 0.86 (0.82-0.9). The adjusted OR (95% CI) of CPR for a woman who had 11, 12, 13 and ≥14 days of COS, compared to referent was 0.95 (0.89-1.01), 0.93 (0.87-0.99), 0.8 (0.75-0.86) and 0.7 (0.65-0.75) respectively. Conclusions and Relevance: In this nationwide cohort study of women undergoing fresh IVF-ET using autologous oocytes, controlled ovarian stimulation lasting approximately 10-days was associated with an optimal live birth rate.

O-058 Obstetric outcomes of twin gestation after In-Vitro Fertilization and Embryo Transfer (IVT-ET) treatment in patients who underwent hysteroscopic septoplasty for subseptate uterus

Abstract Study question Is there an associated increased risk of preterm birth (PTB) in patients who underwent hysteroscopic septoplasty for subseptate uterus (SSU) with twin gestation through IVF-ET? Summary answer This study suggests an increased risk of PTB in patients, who underwent hysteroscopic septoplasty for SSU, and subsequently conceived twins after IVF-ET treatment. What is known already In general, the incidence of PTB is higher after IVF-ET treatment. In addition, Müllerian uterine anomalies, such as unicornuate uterus, are considered risk factors for PTB, even with singleton pregnancy. Furthermore, historical data suggest that some patients with uterine anomaly may have cervical incompetence. Twin gestation is associated with PTB in approximately 50% of cases. It is assumed that the obstetric outcome of twin gestation after successful septoplasty should be no different from those with normal uterine cavity. The impact of previous septoplasty for SSU on the obstetric outcomes of twin gestation after IVF-ET treatment has not been adequately studied. Study design, size, duration This retrospective cohort study included 377 fertility patients who received treatment between 2006-2019. The study group included 162 patients who underwent hysteroscopic septoplasty for SSU and subsequently conceived dichorionic-diamniotic (DI-DI) twins after IVF-ET treatment (Group 1). The control group included 215 patients who had normal endometrial cavity on hysteroscopy (#109) or Transvaginal 3D ultrasound scan with saline sonohysterogram (#106) and subsequently conceived DI-DI twins after IVF-ET treatment (Group 2). Participants/materials, setting, methods This study was conducted at an infertility clinic affiliated with an academic hospital. Demographic, clinical, and obstetric outcome data were collected for both groups and compared using appropriate statistical methods. PTB, severe PTB, extreme PTB, and peri-viable birth were defined as < 37 weeks, <32 weeks, <28 weeks and between 20 weeks-25 weeks 6 days gestation respectively. Main results and the role of chance There was no significant difference in mean age, BMI and infertility duration or in the incidence of smoking, primary infertility, tubal factors and unexplained infertility between the two groups. There was significantly lower mean baseline FSH levels (p < 0.01), lower incidence of male infertility (p < 0.001) and higher incidence of ovulatory disorders (p < 0.001) and endometriosis (p < 0.05) in Group 1 compared to Group 2. As expected, there was past history of lower parity rate (p < 0.01) and higher miscarriage rate (p < 0.001) in Group 1 compared to Group 2 prior to septum division. There was significantly higher incidence of PTB (72.8% vs 56.3%, p < 0.01), severe PTB (20.3% vs 8.2%, p < 0.01), extreme PTB (8.3% vs 3.9%, p < 0.05), and peri-viable birth (6.3% vs 1.9%, p < 0.05) in Group 1 compared to Group 2. There was no significant difference in cesarean section rate (82.8% vs 87.5%, p = 0.349) between Group1 and Group 2 respectively. There was significantly lower gestational age at birth in weeks (34.0 + 4.0 vs 35.6 + 3.0, p < 0.001), birth weight of the first newborn (2180.5 + 717.9 vs 2436.4 + 606.6, p < 0.001), and the second newborn (2235.2 + 674.2 vs 2398.0 + 537.9, p < 0.05) in grams in Group 1 compared to Group 2. Limitations, reasons for caution Our study has limitations being retrospective in nature. Data on cervical length measurements via ultrasound, if performed, was not available. However, recall bias was prevented by retrieving data from that submitted to the Society of Assisted Reproductive Technology. Additionally, procedural variation was reduced as this was a single-center study. Wider implications of the findings Our study suggests that twin gestation after IVF-ET in patients who underwent successful hysteroscopic septoplasty for SSU may increase the likelihood of PTB. Such patients may require increased surveillance with ultrasound cervical length and signs for preterm labor. Future research should focus on measures to reduce PTB in such patients. Trial registration number Not Applicable

P-787 Factors associated with live birth rate (LBR) and multiple live birth rate (mLBR): UK vs France registries

Abstract Study question Do the associations between age, previous attempts, frozen and day-5 transfers relative to LBR and mLBR differ between national registries (UK vs France)? Summary answer Both registries agree age negatively correlates with LBR and mLBR; disagree on effects of frozen transfers, previous attempts and day-5 transfers on LBR and mLBR. What is known already Due to the high risks of multiple pregnancy, the focus of ART success was shifted from having a high number of live birth rate (LBR) to one full-term, healthy baby and a low multiple live birth rate (mLBR). Elective single embryo transfer (eSET) has been an effective strategy for reducing the risk of mLBR but in most guidelines, it is recommended for young patients (<35 years) only. There is extensive evidence from large-scale studies characterising how factors, seen in eSET policies, affect LBR. However, only a few studies focus on mLBR. Study design, size, duration A retrospective analysis of two national registry datasets, HFEA (UK) (N = 442042; 2014-2018) and ABM (France) (N = 305142; 2014-2018), evaluating how age (<35 yrs, 35-37 yrs, 38-40 yrs, >41 yrs), fresh/frozen embryo transfer (ET), previous attempts (0, 1, 2, 3, 4, 5 attempts), and embryo stage (day-5 or day-3 transfers) impact LBR and mLBR following eSET (control group) and multiple embryo transfer (MET) (treatment group). Participants/materials, setting, methods Statistical analyses of the datasets were conducted using adjusted two-sided odds ratios from chi-squared tests with 95% confidence intervals (α = 0.05) with Bonferroni correction. Main results and the role of chance Two registries agreed on the negative correlation between age and LBR (HFEA: eSET OR:1.00-0.79-0.73-0.57, p < 0.001; MET OR:1.00-0.85-0.74-0.55, p < 0.001; ABM: eSET OR:1.00-0.75-0.71-0.52, p < 0.001; MET OR:1.00-0.80-0.69-0.50, p < 0.001) and mLBR following MET (HFEA OR:1.00-0.79-0.70-0.71, p < 0.001; ABM OR:1.00-0.74-0.74-0.59, p < 0.001). mLBR following eSET remained constant with increasing age in both datasets (HFEA OR:1.00-0.91-0.92-1.34, NS; ABM OR:1.00-0.87-1.02-1.13, NS). LBR was significantly higher for fresh vs frozen eSET in both datasets (HFEA OR:1.36, p < 0.001; ABM OR:1.83, p < 0.001). However, HFEA showed no significant difference in LBR between fresh and frozen MET (OR:0.98, NS), whereas LBR in fresh MET was significantly higher in ABM (OR:1.28, p < 0.001). There was disagreement on fresh vs frozen ET mLBR (HFEA: eSET OR:1.10, NS; MET OR:0.97, NS; ABM: eSET OR:0.81, p < 0.05; MET OR:1.36, p < 0.001). Registry results differed regarding the impact of previous attempts on LBR and mLBR.Day-5 ETs had significantly higher LBR compared to day-3 (HFEA eSET OR:1.29, p < 0.001; MET OR:1.37, p < 0.001; ABM: eSET OR:1.50, p < 0.001; MET OR:1.35, p < 0.001). mLBR was significantly higher following day-5 MET (HFEA OR:1.59, p < 0.001; ABM OR:1.27, p < 0.001) but only significantly higher following eSET in the French registry (HFEA OR:0.64, NS; ABM OR:1.40, p < 0.01). Limitations, reasons for caution Limitations included selection bias due to the retrospective design of the study, as a result, the populations include predominantly caucasian, European populations. The conclusions drawn relate to the practices from particular countries, so it is important to evaluate multiple, more diverse datasets. Wider implications of the findings It is encouraging that both registries agreed on the effects of age, whilst remaining factors were not generalizable and require further assessment. This study demonstrated that even large, national registries contain inherent biases and policymakers should not draw policies based on one dataset, especially when prospective trial is not feasible. Trial registration number N/A

O-256 Effect of an endometrial microbiota on pregnancy outcome of frozen embryo transfer (FET) cycles

Abstract Study question Is specific microbiota in the endometrial fluid (EF) associated with pregnancy outcome in a frozen embryo transfer (FET) cycles? Summary answer Lactobacillus had high abundance in the microbiota of endometrial fluid of patients with successful pregnancy after FET treatment. What is known already Recently, the relationship between endometrial microbiota and repeated implantation failure (RIF) has been reported. While Lactobacillus-dominated microbiota (LDM, defined as > = 90 % Lactobacillus species) in the EF of the receptive phase was reported to be associated with favorable reproductive outcome, non-LDM (< 90 % Lactobacillus species) was found to decrease implantation, clinical pregnancy, ongoing pregnancy, and live birth rates. However, it is still unclear the effect of the microbiota on pregnancy outcome of the patients with the assisted reproductive technology, especially a frozen embryo transfer (FET) treatment. Study design, size, duration We included 802 cycles with clinical results of pregnancy outcome after FET treatment at our clinic from December 2018 to January 2021. Endometrium fluid was collected before FET and microbiota was examined. We examined the relationship between endometrial microbiota and pregnancy outcome in 463 cycles in which the endometrial microbiome test results were available, pregnancy outcome was known, CD138 negative, and less than 38years (229 cycles were positive pregnancy and 234 cycles were negative pregnancy). Participants/materials, setting, methods Sampling was performed carefully avoiding contamination before FET treatment. Extracted genomic DNA was sequenced for the region of 16S ribosomal RNA using next-generation sequencer (Endometrial microbiome test, Varinos, Japan). The sequencing data was assigned to bacterial taxonomy and the background-contaminated bacteria were excluded from the microbiome profile. Lactobacillus abundance was calculated. After concordance of patient background, Lactobacillus abundance between successful and unsuccessful pregnancy group was compared. Main results and the role of chance We investigated the effect of microbiota in EF, especially Lactobacillus, for the infertile female with frozen embryo transfer (FET) as an assisted reproductive treatment with large amount of cycles (802 cycles). First, all of cycles were divided into a successful pregnancy group (305/802) and an unsuccessful pregnant group (497/802), and then the abundance of Lactobacillus was analyzed (the pregnancy ratio was 38.0%). The mean abundance of Lactobacillus in EF was significantly higher in the successful pregnant group than in the unsuccessful pregnant group (70.2% 40.5 versus 63.5% 43.1, p = 0.007). To further refine the conditions of each cycle, patient backgrounds of each cycle were compared. There was a significant difference in age and CD138 results between the successful pregnant group and unsuccessful pregnant group, so the analysis was performed with the limited cycles with CD138-negative and under 38 years (229/463 versus 234/463, respectively). However, even under this strict condition, the mean abundance of Lactobacillus was significantly higher in the pregnant group than in the unsuccessful pregnant group (72.1% 39.7 versus 61.1% 44.4, p = 0.003). These results suggest that the endometrial environment with high Lactobacillus abundance is more conducive to pregnancy in FET treatment. Limitations, reasons for caution The limitation of this study is that the design is not a randomized controlled one, although it is prospective. Wider implications of the findings The results of this study suggest that the endometrial microbiota at the time of embryo implantation, especially Lactobacillus genus, is highly relevant to pregnancy outcome, implying that disruption of the intrauterine microbiota may cause infertility. We need further studies to improve the endometrial microbiota of infertile women. Trial registration number not applicable

P-572 Hormonal Monitoring and Progesterone Adjustment in Frozen Embryo Transfer Cycles

Abstract Study question We investigated the effect of monitoring and follow-up of serum progesterone, estradiol & luteinizing hormone levels and progesterone supplement adjustments on pregnancy outcomes for FET in programmed HRT cycles. Summary answer Monitoring and follow-up of different hormones in patient’s serum in programmed frozen embryo transfer cycles didn’t have an impact on overall pregnancy rate. What is known already Frozen Embryo transfer treatment rapidly expanded in recent years. FET can effectively prevent IVF associated complications, such as ovarian hyperstimulation syndrome and multiple pregnancy. In addition, FET serves as a safe & cost-effective way to increase cumulative pregnancy rate. Though progesterone levels on the day of transfer have not yet been studied extensively in humans, tailoring the time of transferring a frozen embryo based on serial P4 values rather than cycle day number alone results in higher pregnancy rates.The value of measuring the serum E2 levels on the day of embryo transfer as an indicator for clinical pregnancy is still doubtful. Study design, size, duration Six Hundred FETs were performed in a randomized prospective manner in an infertility center in Alexandria starting from 2019 till 2021.These were further subdivided based on computer randomization into Group I (Non-intervention) including300 patients with only ultrasound monitoring & Group II (intervention) including 300 patients with monitoring of serum level of progesterone, estrogen & luteinizing hormone as well as progesterone supplement adjustments. Participants/materials, setting, methods Group I (Non-intervention) including300 patients with only ultrasound monitoring & Group II (intervention) including 300 patients with monitoring of serum level of progesterone, estrogen & luteinizing hormone as well as progesterone supplement adjustments which are further subdivided into three groups depending on progesterone level in patient’s serum early in the morning prior to embryo transfer. Group II A: P4 levels < 5ng/dl, Group II B: P4 levels 5-10ng/dl, Group II C: P4 levels >10ng Main results and the role of chance Live birth and overall pregnancy rate showed no significant difference among group monitored with ultrasound only compared to the hormonal monitored group. However, on the day of embryo transfer, serum P4 > 11.83 ng/ml represented a cut-off value above which there is marked increase in OPR/LBR and a sensitivity of 53.7% & specificity of 51.2% in predicting pregnancy. Also, serum E2 value more than 292 pg/ml at day 18 demonstrated an increase in OPR/LBR.In our study, the cut off value of LH level less than of 10.8 IU/L at day13 had a sensitivity of 58.3% & specificity of 50.4% in predicting pregnancy. Regarding pregnancy outcome, monitoring of LH levels didn’t generate useful data with no added prognostic value.Conclusion: Monitoring and follow-up of different hormones in patient’s serum in programmed frozen embryo transfer cycles didn’t have an impact on live birth, overall pregnancy & abortion rate when compared to ultrasound only in non-intervention group. Limitations, reasons for caution Patients in non-intervention group didn’t have a hormonal pre-transfer assessment so it’s not possible to determine if the progesterone levels are suboptimal or not when compared to intervention group. Also, we didn’t have a post transfer hormonal assessment in case group to determine whether the progesterone dose reached desired levels. Wider implications of the findings Kofinas et al. proposed in his study that in order to optimize pregnancy and rate, P4 levels in FET cycles should be sustained between 10 and 20 ng/ml. This was in accordance to our study where the cut off value of P4 on day of 11.83 ng/ml optimized pregnancy rates. Trial registration number NCT05189145

Should any use of artificial cycle regimen for frozen-thawed embryo transfer in women capable of ovulation be abandoned: yes, but what's next for FET cycle practice and research?

Over the past decade, the use of frozen-thawed embryo transfer (FET) treatment cycles has increased substantially. The artificial ('programmed') cycle regimen, which suppresses ovulation, is widely used for that purpose, also in ovulatory women or women capable of ovulation, under the assumption of equivalent efficacy in terms of pregnancy achievement as compared to a natural cycle or modified natural cycle. The advantage of the artificial cycle is the easy alignment of the time point of thawing and transferring embryos with organizational necessities of the IVF laboratory, the treating doctors and the patient. However, recent data indicate that pregnancy establishment under absence of a corpus luteum as a consequence of anovulation may cause relevant maternal and fetal risks. Herein, we argue that randomized controlled trials (RCTs) are not needed to aid in the clinical decision for or against routine artificial cycle regimen use in ovulatory women. We also argue that RCTs are unlikely to answer the most burning questions of interest in that context, mostly because of lack of power and precision in detecting rare but decisive adverse outcomes (e.g. pre-eclampsia risk or long-term neonatal health outcomes). We pinpoint that, instead, large-scale observational data are better suited for that purpose. Eventually, we propose that the existing understanding and evidence is sufficient already to discourage the use of artificial cycle regimens for FET in ovulatory women or women capable of ovulation, as these may cause a strong deviation from physiology, thereby putting patient and fetus at avoidable health risk, without any apparent health benefit.

Changes in serum heavy metals in polycystic ovary syndrome and their association with endocrine, lipid-metabolism, inflammatory characteristics and pregnancy outcomes

Polycystic ovary syndrome (PCOS) is the most common endocrine disorder among women of reproductive age. Heavy metals, a nonnegligible pollutant, have significant effects on human health, and they are related to reproductive dysfunction. This study aimed to investigate whether serum heavy metal levels were related to PCOS and the association of increased heavy metals with endocrine, lipid-metabolism and inflammatory characteristics and pregnancy outcomes. 80 subjects (40 with PCOS and 40 for control) were selected to analyze their clinical characteristics and serum heavy metal concentrations. Serum chromium (Cr), mercury (Hg) and copper (Cu) were significantly higher in PCOS patients. Cr was positively correlated with luteinizing hormone (LH) (ρ = 0.240 and P = 0.033) and negatively correlated with prolactin (PRL) (ρ = −0.248 and P = 0.032), Hg was positively correlated with LH (ρ = 0.446 and P < 0.001), testosterone (T) (ρ = 0.300 and P = 0.008), androstenedione (A) (ρ = 0.316 and P = 0.006), interleukin-6 (IL-6) (ρ = 0.344 and P = 0.002), and tumour necrosis factor-α (TNF-α) (ρ = 0.248 and P = 0.026), and Cu was found to be positively correlated with low-density lipids (LDL) (ρ = 0.264 and P = 0.021), IL-6 (ρ = 0.295 and P = 0.008), TNF-α (ρ = 0.336 and P = 0.002) and white blood cells (WBC) count (ρ = 0.300 and P = 0.008). Cu levels were significantly higher in nonpregnant patients than in pregnant patients who received in vitro fertilization and embryo transfer treatments. In conclusion, the present study shows that PCOS patients exhibited increased serum Cr, Hg and Cu levels, which were related to some sex hormones and lipid-metabolism disturbances and inflammatory cytokines releases, and elevated Cu levels were related to pregnancy failure, suggesting that these heavy metals may provide valuable information to predict PCOS.

Complex Uterine Cavity Abnormalities Increase the Risk of Miscarriage in In Vitro Fertilization/Intracytoplasmic Sperm Injection in Fresh Cycle-Assisted Pregnancies

Any abnormality of the uterine cavity can result in reduced endometrial receptivity, which negatively affects embryo implantation and leads to lower clinical pregnancy rates. The effects of improved uterine cavity environment on in vitro fertilization (IVF)/intracytoplasmic sperm injection (ICSI)-embryo transfer (ET) treatment outcome are unclear. This study aimed to investigate the impact of improved uterine cavity abnormalities on the pregnancy outcomes of infertile patients undergoing IVF/ICSI-ET. Retrospective study. Single-center. Women with infertility who underwent fresh cycles of IVF/ICSI-ET. We retrospectively analyzed the clinical data of 31 057 cycles of women with infertility undergoing IVF/ICSI-ET with hysteroscopy and treated at the First Affiliated Hospital of Zhengzhou University Reproductive Medicine Center from August 2009 to May 2018. According to the previous condition of their uterine cavity, patients were divided into the normal uterine cavity, single uterine cavity abnormality, and complex uterine cavity abnormality groups. Differences in general conditions and pregnancy-assisted outcomes were compared among the groups, which were screened according to propensity score matching. In 3005 cycles after propensity score matching screening, there were no statistically significant differences in the implantation and clinical pregnancy rates of patients with successfully treated uterine cavity abnormalities and lesions (p > .05). The miscarriage rate was significantly higher in the complex uterine cavity abnormality group than in the normal (p=.001) and single uterine cavity abnormality groups (p=.002). Logistic regression analysis showed that the female partner's age (adjusted odds ratio, 1.12; 95% confidence interval, 1.05-1.19; p=.001) and history of intrauterine adhesions (adjusted odds ratio, 1.44; 95% confidence interval, 1.12-1.85; p=.005) were independent risk factors for miscarriage. The age of the female partner and history of intrauterine adhesions increased the miscarriage rate in patients undergoing IVF/ICSI-ET.

PTEN expression in human cumulus cells is associated with embryo development competence.

Embryo quality determines the success of in vitro fertilization and embryo transfer (IVF-ET) treatment. Biomarkers for the evaluation of embryo quality have some limitations. Apoptosis in cumulus cells (CCs) is important for ovarian function. PTEN (phosphatase and tensin homolog) is a well known tumour suppressor gene that functions as a mediator of apoptosis and is crucial for mammalian reproduction. In the present study, we analyzed the expression level of PTEN in human CCs and aimed to investigate its association with embryo developmental competence in IVF treatment cycles. The PTEN mRNA level in CCs was measured using real-time fluorescence quantitative PCR. The association of the differential expression of PTEN with embryo quality was analyzed. Our data showed that PTEN mRNA levels were significantly decreased in CCs surrounding mature oocytes compared with immature oocytes. Similar changes were found in the analysis of fertilization and blastocyst formation. The speculation that the measurement of PTEN mRNA levels in human CCs would provide a useful tool for selecting oocytes with greater chances to implant into the uterus needs to be further verified through single-embryo transfer in the future. The proapoptotic mechanism of PTEN in human reproduction needs to be further studied.

Expression of vascular endothelial growth factor A (VEGFA), placental growth factor (PlGF) and insulin-like growth factor 1 (IGF-1) in serum from women undergoing frozen embryo transfer

VEGFA, PlGF and IGF-1 are three main angiogenic factors which play significant roles in embryo implantation. However, the relationship between serum expressions of VEGFA, PlGF and IGF-1 and pregnancy outcomes has not been fully illustrated. In this study, serum specimens were collected precisely on day 7 after the LH surge in a natural non-conception cycle from 38 infertile patients who underwent frozen embryo transfer (FET) treatment. ELISA was used to determine the concentrations of VEGFA, PlGF and IGF-1. Serum levels of VEGFA, PlGF and IGF-1 were compared between patients who conceived (n=25) and who did not (n=13). Correlation and linear regression analyses were used to investigate the correlations of serum angiogenic factors and β-hCG MoM levels in the pregnant group. The results demonstrated that no significant difference was found in serum VEGFA, PlGF or IGF-1 concentration between pregnant and non-pregnant women. Spearman correlation analysis revealed a positive correlation between IGF-1 concentration and β-hCG level in pregnant participants (rs = 0.490, p = 0.013). In conclusion, serum IGF-1 level correlated positively with β-hCG level in pregnant women, which may provide information on the prognostic value of IGF-1 in this group of women.

The impact of COVID-19 mitigation measures on fertility patients and clinics around the world

Research questionWhat is the impact of the response to COVID-19 on the management of fertility treatments and clinical practice around the world?DesignFertility clinic associates around the world were approached. They completed an online survey containing 33 questions focused on the country's response to the COVID-19 pandemic. Known fertility clinic associates that were contacted comprised scientific directors, medical directors and laboratory managers.ResultsThere were 43 individual country responses from Asia (13), Africa (3), Europe (17), North America (3), Oceania (2) and South America (5). In nine countries, clinics followed their government body recommendations, in 22 countries there was a combination of recommendations, in 3 countries changes were made by clinic initiative, and 9 countries did not specify. In 34 countries IVF/intracytoplasmic sperm injection (ICSI) and frozen embryo transfer (FET) treatments had an average delay of 56 days (IVF/ICSI) (minimum 0, maximum 160) and 57 days (FET) (minimum 0, maximum 166 days). During the shutdown, the number of freeze-all cycles increased in 22 countries. Only 23 countries reported patients having to undergo a SARS-CoV-2 test, and 20 countries did not report any COVID-19 testing in their clinic. Additional support counselling was offered in 28 countries, partner restrictions at clinics were reported in 41 countries and time between patients’ appointments was increased in 39 countries.ConclusionsThe implications of COVID-19 mitigation measures proved the need for government societies to introduce a set protocol that includes requirements such as increased patient counselling and additional guidelines for prioritizing couples who need care most urgently.

Excluding embryos with two novel mutations in FREM2 gene by the next-generation sequencing-based single nucleotide polymorphism haplotyping.

Fraser syndrome is a rare autosomal recessive malformation disorder. It is characterized by cryptophthalmos, syndactyly, urinary tract abnormalities and ambiguous genitalia. This condition is due to homozygous or heterozygous mutations in the FRAS1, FREM1, FREM2, and GRIP1 genes. In the present study, we recruited a Chinese family with Fraser syndrome. Two novel mutations c.7542_7543insG and c.2689C>T in the FREM2 gene were detected in this Fraser syndrome family by PCR-based sequencing. The next-generation sequencing-based single nucleotide polymorphism haplotyping method was applied to exclude these two mutations in 9 blastocysts obtained from the patient. After obtaining consent and informing the risk, the patient received in vitro fertilization and embryo transfer treatment with an embryo carrying a heterozygous mutation. Finally, she delivered a healthy baby without any complications on March 17, 2019. In conclusion, we first reported two novel mutations in the FREM2 gene associated with the risk of Fraser syndrome. Moreover, we described a next-generation sequencing-based single nucleotide polymorphism haplotyping method to select the ‘right’ embryos from patients with Fraser syndrome for in vitro fertilization and embryo transfer treatment.

P–630 Progesterone levels using pessaries of 400 mg of vaginal progesterone (Cyclogest®) in artificial cycles for frozen embryo transfer

Abstract Study question Does the use of pessaries of 400 mg of micronized progesterone provide comparable results as pessaries of 200 mg x2, in terms of progesterone levels? Summary answer The administration of pessaries of Cyclogest® 400 mg reduces the probability of presenting suboptimal level of progesterone on the day of the embryo transfer. What is known already The endometrial preparation for frozen embryo transfer (FET) in Artificial Cycle (AC) with vaginally-administered progesterone, is one of the most common IVF procedures nowadays. Now, it has been shown that suboptimal progesterone levels on the day of the embryo transfer compromise the results of FET treatments. Recently, a new preparation of 400 mg vaginal pessaries has been introduced in the market of European countries. Efficacy of this new preparation has been studied in “fresh” IVF cycles but we lack the comparative studies in AC making it necessary to further investigate this area. Study design, size, duration Non-inferiority retrospective case-control trial based on 347 embryo transfer treatments with endometrial preparation in AC carried out at Instituto Bernabeu between January 2019 and July 2020. 153 patients received 1 pessary of 400 mg every 12 hours (group A) and 194 received 2 pessaries of 200 mg every 12 hours (group B). Sampled size calculation resulted in 182 patients required to detect a minimum difference of 2 ng/ml so sample was powered for the purpose. Participants/materials, setting, methods Patients receiving embryos in AC preparation were included. All embryo transfers were performed at blastocyst stage after 5 days of progesterone administration. Progesterone levels were assessed the day of the embryo transfer by an electrochemiluminescence immunoassay.Primary outcome was the incidence of suboptimal progesterone levels according with the cutoff value stablished in the literature at 8.8 ng/mL. Secondary outcomes were pregnancy rates (PR), clinical pregnancy rates (CPR), ongoing pregnancy rates (OPR) and miscarriage rates (MR). Main results and the role of chance Incidence of suboptimal levels of progesterone was significantly lower in the group of 400 mg (9,8% in Group A vs 19,7% in the Group B, p = 0.011). Given that there was an imbalance between groups in the body weight (66.9 +/- 14 vs. 61.9 +/- 13.165 kg, p &amp;lt; 0.001) and BMI (24.63 +/- 4.861 vs. 22.54 +/- 3.092, p &amp;lt; 0.001), we decided to perform a binary logistic regression setting patient’s weight and BMI as confounding variables. The result confirms a higher risk of suboptimal progesterone levels (&amp;lt;8.8) with the 2x200 mg regimen (OR: 2.52 95%CI: 1.28–4.96; p = 0.007). Mean progesterone levels were similar in both groups (13,8035 ng/mL +/- 4.62159 vs. 13.9799 ng/mL +/- 7.73243 respectively, p = 0.146). No differences were observed in clinical outcomes: PR (52.3% vs. 53.1%, p = 0.881), BM (14.7% vs. 17.6%, p = 0.597), CM (20% vs. 18.6%, p = 0.819) and OPR (33.1% vs. 33.7%, p = 0.912). The subjective medical decision to administer additional progesterone from the day of the embryo transfer onwards (taking values other than 8.8 ng / mL as a reference), was significantly lower in the group of 400 mg (24,3% vs 37,3%, p = 0.009 ). Limitations, reasons for caution The inherent limitations of a retrospective analysis. The study was not powered to detect differences in clinical outcomes. Therefore, results other than progesterone levels should be interpreted with caution. Wider implications of the findings: A single pessary of 400 mg minimizes the necessity of additional medication (usually subcutaneous progesterone). Presentation of 400 mg is superior to 2x200 providing adequate progesterone levels and patient comfort. Dose finding and pharmacokinetics studies of the vaginal administration will be necessary for the future to optimize FET under AC. Trial registration number NCT04722471

An exploration of contributing factors to the oocyte utility rate of high responders in an in vitro fertilization cycle

Objective: To figure out the clinical factors contributing to the oocytes utility rate (OUR) of high responders in in vitro fertilization (IVF)-embryo transfer treatment. Methods: OUR was defined by the number of usable embryos for transfer and (or) cryopreservation divided by the number of oocytes retrieved in a freeze-all cycle. The cycles with ≥15 eggs were included from January 2013 to December 2019. Those with OUR at the top 10% (Group A) and the bottom 10% (Group B) were picked and compared for patients' characteristics, parameters relating to ovary stimulation and pregnant outcomes. Multifactorial logistic regression was applied to reveal the risk factors affecting OUR in them. Results: A total of 43 patients were included in Group A (OUR: 77.4%, 601/776) and 47 for Group B (OUR: 11.9%, 104/874). Previous IVF/intracytoplasmic sperm injection (ICSI; OR=0.10, 95%CI: 0.01-0.81) and endometriosis (OR=0.16, 95%CI: 0.03-0.84) were negative factors for OUR (P<0.05); dual suppression protocol (OR=3.74, 95%CI: 1.06-26.86) and longer days of stimulation (OR=3.24, 95%CI: 1.25-8.42) were protective factors in terms of ovarian stimulation (P<0.05), on contrary to that, any decline of estradiol during the stimulation led to poorer OUR (OR=0.16, 95%CI: 0.04-0.64). Although two groups had similar quantities of eggs and metaphase of meiosis Ⅱ (MⅡ) oocytes, distinguished cumulative clinical pregnancy rate and cumulative live birth rate were seen in Group A and group B respectively [95.3% (41/43) vs 40.4% (19/47) and 90.7% (39/43) vs 31.9% (15/47), all P<0.01]. Conclusions: High responders with attempted IVF/ICSI and endometriosis should be considered as risk factors for OUR. Over ovarian stimulation, dual suppression and a slightly longer stimulating duration could be tried; besides, estradiol decline should be prevented for a better OUR.

Exogenous progesterone for LH surge prevention is redundant in ovarian stimulation protocols

During ovarian stimulation for IVF–embryo transfer treatment, a premature LH surge may lead to progesterone elevation that disrupts endometrial maturation and affects the probability of pregnancy following fresh embryo transfer. Preventing this LH surge and progesterone elevation using gonadotrophin-releasing hormone (GnRH) analogues is considered a standard practice. The same policy applies to cycles in which the ‘freeze-all’ protocol has been selected from the outset (e.g. donors), but the need for this has not been discussed. Moreover, in ‘freeze-all’ cycles, exogenous progesterone administration tends to replace GnRH antagonists, without reducing efficacy after embryo transfer in frozen-thawed cycles. Nevertheless, as exogenous progesterone is expected to have the same impact on the endometrium as endogenous progesterone, it is clear that, unlike in fresh cycles, in ‘freeze-all’ cycles an endogenous LH surge prevention does not seem necessary. Therefore, both GnRH antagonists and exogenous progesterone appear to be redundant in ‘freeze-all’ cycles, and in this context the indications for the use of GnRH analogues in ovarian stimulation protocols need to be revisited.

PTEN Expression in Human Granulosa Cells Is Associated with Ovarian Responses and Clinical Outcomes in IVF.

The ovarian reserve determines the success of in vitro fertilization (IVF) and embryo transfer treatment. It predicts the ovarian response in controlled ovarian hyperstimulation cycles. Apoptosis in granulosa cells surrounding oocytes is important for ovarian function and has been closely associated with follicular atresia. PTEN (encoding phosphatase and tensin homolog) is a well-known tumor suppressor gene that functions as a mediator of apoptosis and is crucial for mammal reproduction. In the present study, we analyzed the expression level of PTEN in human granulosa cells and aimed to investigate its association with the ovarian response and clinical outcomes in IVF. Apoptosis in granulosa cells were analyzed using Annexin V-Allophycocyanin staining after PTEN short hairpin RNA lentivirus transfection. Real-time fluorescent quantitative PCR analysis showed that the PTEN transcript level was significantly higher in poor responders and significantly lower in high responders, compared with that in normal responders. However, PTEN expression in the pregnancy group decreased slightly, but not significantly, compared with that in the non-pregnancy group. The apoptosis rate of granulosa cells declined significantly after 24-h transfection of the PTEN-shRNA lentivirus. These results suggest a fundamental role of PTEN in the regulation of follicular development, and that it might be involved in the pathogenesis of follicular dysplasia and ovarian dysfunction.

Relationship Between Anti-Müllerian Hormone and In Vitro Fertilization-Embryo Transfer in Clinical Pregnancy

ObjectivesTo retrospectively analyze the correlation between anti-Müllerian hormone (AMH) and the number of oocytes obtained by controlled ovarian hyperstimulation (COH) in women of different ages and explore the factors affecting in vitro fertilization and embryo transfer (IVF-ET) in clinical pregnancy of infertile women to provide evidence for infertile women to choose assisted reproduction strategies.MethodsInfertile women who received IVF-ET or intracytoplasmic sperm injection and embryo transfer (ICSI-ET) treatment in the reproductive center of XX hospital between October 2018 and September 2019 were included. Patient data on medical records, age, body mass index (BMI), years of infertility, basic follicle-stimulating hormone (FSH), basic luteinizing hormone (LH), basic estradiol (E2), anti-Müllerian hormone level (AMH), antral follicle count (AFC), gonadotropins (Gn) medication days, Gn dosage, endometrial thickness on transplantation day, the number of retrieved oocytes, the number of mature oocytes obtained, the number of embryos transferred, clinical pregnancy status, etc., were collected.ResultsA total of 314 patients were enrolled in this study, with an average age of 31.0 ± 4.5 years. The infertility period ranged from 0–21 years. The AMH level showed a downward trend with increasing age. Overall, the AMH level of women of all ages was positively correlated with the number of retrieved oocytes (r = 0.335, p < 0.001). The AMH level of women between 22 and 28 years old was positively correlated with the number of retrieved oocytes (r = 0.164, p < 0.061) but it was not statistically significant. Similarly, the AMH level of women aged 29–35 and 36–43 was positively correlated with the number of retrieved oocytes (r = 0.356, p < 0.001; r = 0.461, p < 0.001). The average age of the pregnant group (30.6 ± 4.4 years) was lower than that of the non-pregnant group (32.2 ± 4.6 years) (p < 0.001). The number of oocytes obtained (9.8 ± 4.5) and the number of embryos transferred (1.9 ± 0.4) in the pregnant group was significantly higher than that in the non-pregnant group (9.2 ± 4.5; 1.7 ± 0.5); the difference was statistically significant. The multivariate logistic regression model showed that age (OR = 0.574 95% CI: 0.350–0.940), AMH (OR = 1.430 95% CI: 1.130–1.820) and the number of oocytes obtained (OR = 1.360 95% CI: 1.030–1.790) were factors affecting clinical pregnancy.ConclusionWe found that the level of AMH in infertile women decreased with age and the number of oocytes obtained in infertile women was positively correlated with AMH. Moreover, the number of oocytes and embryo transferred in the pregnant group was significantly higher than those in the non-pregnant group. Furthermore, age, AMH and the number of oocytes affected the clinical pregnancy.

Bitki ve Hayvan Biyoteknolojisi; Hücresel Tarım ve Nano-Teknoloji

Tarımsal biyoteknoloji araştırmacılara, tarımı ve yetiştiriciliği yapılan bütün organizlamarın genetiğini anlama ve manipüle etme imkanı sağlayan bir alandır. Tarımsal biyoteknolojinin başlangıcında fermantasyon gibi yöntemler sık kullanılırken, bugün modern tarımsal biyoteknoloji besinlerin kalitesini, miktarını, içeriğini arttırmaya ve tat gibi farklı özellikleri değiştirmeye imkan sağlamaktadır. Bitki biyoteknolojisi alanındaki çalışmalar çoğunlukla bitkilerde verim ve kaliteyi arttırmanın yanında biyotik ve abiyotik stres faktörlerine karşı dayanıklı bitkiler geliştirmeye odaklanırken, hayvan biyoteknolojisi ise hayvansal ürünlerin kalitesini arttırma, suni dölleme, embriyo transferi, hayvan hastalıklarının daha ucuz ve kolay bir şekilde teşhis ve tedavi yöntemlerinin geliştirilmesi konularını ele almaktadır. Bu çalışmada bitkisel ve hayvansal üretim alanında yeni uygulama alanı bulan hücresel üretim ve nano-biyoteknoloji uygulamaları irdelenmiştir.

Female cigarette smoking and successful fertility treatment: A Danish cohort study.

Despite smoking being a well-established risk factor for adverse pregnancy and neonatal outcomes, a substantial proportion of women of reproductive age smoke. Previously, meta-analyses have indicated a significantly negative impact of female smoking on outcomes of assisted reproduction, yet most of the included studies have several, essential methodological limitations. We aimed to investigate whether female cigarette smoking may affect the chance of achieving a clinical pregnancy and live birth among women and couples receiving medically assisted reproduction treatment. A cohort study with longitudinally and repeatedly collected exposure information from 1 January 2010 to 31 August 2015, including data on 1708 women and potential partners initiating either intrauterine insemination, in vitro fertilization (IVF)/intracytoplasmic sperm injection (ICSI) or frozen embryo transfer treatment cycles at the public Fertility Clinic, Aarhus University Hospital, Denmark. Smoking was assessed from self-reported questionnaires completed before treatment. Outcomes were a clinical pregnancy and a live birth. Information on these was obtained from the Danish national health registries, allowing complete follow-up. To evaluate associations between female occasional/daily cigarette smoking and successful medically assisted reproduction treatments, a modified Poisson regression with robust standard errors was used. Female occasional/daily cigarette smoking was not associated with the chance of achieving a clinical pregnancy or a live birth in all intrauterine insemination or IVF/ICSI treatment cycles. When compared with nonsmokers, the adjusted relative risk for obtaining a live birth for those reporting smoking was 1.22 (0.70-2.12) among women initiating 1456 intrauterine insemination treatment cycles. Among women initiating 2788 IVF/ICSI treatment cycles, those reporting occasional/daily smoking had a relative risk for obtaining a live birth of 1.15 (0.82-1.60) when compared with nonsmokers. Occasionally/daily cigarette smoking women had similar chance of achieving a clinical pregnancy or a live birth as the nonsmokers when receiving medically assisted reproduction treatments. However, tobacco use before and during pregnancy remains a major cause of reduced fertility as well as maternal, fetal, and infant morbidity and mortality, and should strongly be discouraged.