GnRH Agonist Trigger Research Articles

P–273 Effects of ovulation induction with GnRH Agonist (GnRHa) on oocyte and embryo quality at the mitochondrial level: A retrospective and experimental study

Abstract Study question Does the GnRHa trigger improve oocyte and embryo quality in patients younger than 40, and do mtUPPR have a role? Summary answer GnRHa trigger improves oocyte nuclear/cytoplasmic maturation, blastocyst utilization and downregulates HSP60 levels and upregulates ATF5 levels compared to hCG trigger. GnRHa trigger suppresses mitochondrial stress. What is known already hCG has been used for decades to achieve final oocyte maturation and, thereby, correct oocyte retrieval timing in connection with ovarian hyperstimulation protocols. As an alternative to hCG, a GnRH agonist has been used to trigger the endogenous release of LH (and FSH) in a fashion resembling the mid-cycle surge of gonadotrophins. GnRHa is as effective as hCG for the induction of ovulation. It has been very well known that the GnRHa trigger improves oocyte nuclear maturation, embryo quality, and implantation rate, but the underlying mechanism remains unknown. Study design, size, duration 3054 women younger than 40; oocytes retrieved more than 10 (up to 20) analyzed. Male infertility was excluded. Ovulation triggered either by hCG (n = 1368) or GnRHa (1668). Female mice were divided into three groups as control, hCG-treated and GnRHa-treated group. Superovulation was performed by FSH + hCG or GnRHa. Oocytes were collected 13 hours after hCG/GnRHa injection. ATF5, BiP, and HSP60 levels were analyzed by Western blot. Statistical analysis was performed using Student’s t-test. Participants/materials, setting, methods This study has two parts. i) RCT and ii) Experimental. In the experimental part, three months old female BALB/C mice (25–30 g) were used and divided into three groups (n = 20/group) as control, hCG-treated and GnRHa-treated group. Superovulation was performed by administering an injection of 5 IU FSH (i.p.) and hCG (i.p.) or GnRHa (20 mg/kg) i.m. Oocytes were collected 13 hours after hCG/GnRHa injection. ATF5, BiP, and HSP60 levels were analyzed by Western blot. Main results and the role of chance The mean age (34.8 vs. 35.2 years), total gonadotropin dose (2176 vs. 2230 IU), and the number of oocytes picked up (14.9 vs. 13.4) were not statistically different among GnRHa and hCG group, respectively. No LH rise or any OHSS was noticed in any groups. Oocyte maturation (79.8% vs. 75.9%), oocyte diameter (as a marker of cytoplasmic maturity) (10198 µm2 and 9474 µm2), fertilization rate (78% vs. 72%), and embryo utilization rate (52% vs. 47.2%) were significantly higher in GnRHa group compared to hCG group, respectively. HSP60 level (activated by mtUPR) was statistically higher in the hCG group compared to the GnRHa group (55% vs. 22%, p < 0.05 respectively). On the other hand, the ATF5 level was significantly higher in the GnRHa group than the hCG group (p < 0.0001). Limitations, reasons for caution The limitation is that this is a proof-of-concept study to reveal the mechanism of good embryo quality with GnRHa trigger. Wider implications of the findings: This application offers convenience and simplifies the IVF protocol with a better oocyte and embryo quality while reducing Ovarian Hyperstimulation Syndrome (OHSS) risk during IVF care Trial registration number Not applicable

P–464 What is the optimal ovarian stimulation (OS) protocol for women who undergo planned oocyte cryopreservation (POC)?

Abstract Study question When repeated cycles of OS for planned oocyte cryopreservation using a standard GnRH antagonist protocol are required, can OS protocol modifications improve oocyte yield? Summary answer Compared to repeating a standard GnRH antagonist protocol, switching to a long GnRH agonist protocol for POC results in a higher number of cryopreserved oocytes. What is known already The total number of cryopreserved oocytes is a key parameter of POC programs because of its association with livebirth. A substantial proportion of women embarking on POC will undergo repeated cycles of OS to reach their desired target number of vitrified oocytes. According to recent guidelines, the GnRH antagonist protocol with GnRH agonist triggering is considered the first choice protocol for POC, because of its safety profile and convenience. However, in women with normal ovarian reserve, the long GnRH agonist protocol results in a higher number of oocytes retrieved. Evidence regarding the optimal protocol for POC is limited. Study design, size, duration This is a single-centre, retrospective cohort study including 283 women who had a first cycle for POC using a standard GnRH antagonist protocol and who requested a second OS cycle to increase their total number of vitrified oocytes for later use. The choice of protocol for the second cycle was left at the discretion of the reproductive medicine specialist. All OS cycles took place between January 2009 and December 2019 in a tertiary referral hospital. Participants/materials, setting, methods After ovarian reserve testing, the first cycle OS was performed using rFSH or HPhMG in a GnRH antagonist protocol. For the second cycle, a GnRH antagonist protocol with or without antagonist pretreatment, or a long GnRH agonist protocol was prescribed. The primary outcome was the number of mature oocytes (MII) vitrified per cycle. Cycle characteristics were compared. Data were assessed by generalized estimating equation (GEE) regression analysis adjusting for covariates. Main results and the role of chance In total, 226 (79.9%) women had a GnRH antagonist protocol and 57 (20.1%) had a long GnRH agonist protocol in their second OS cycle for POC. Overall, mean age was 36.6±2.4 years. The median (CI) number of mature oocytes vitrified after the second OS cycle was significantly higher than that after the first cycle [8 (5–11) vs. 7 (4–10), p < 0.001]. According to GEE multivariate regression, adjusting for relevant confounders, switching from a GnRH antagonist protocol in the first cycle to a long GnRH agonist protocol in the second cycle was the only significant predictor of the number of vitrified oocytes after the subsequent cycle (coefficient 1.59, CI 0.29–2.89, p-value = 0.017). Age, AFC, initial dose and type of gonadotropins did not predict the number of vitrified oocytes. None of the women developed moderate or severe OHSS. Similarly, of 174 women who underwent their first OS cycle with a standard GnRH antagonist protocol, 133 women (76.4%) had the same protocol for their second cycle and 41 women (23.6%) an additional three-day course of GnRH antagonist pretreatment. According to GEE multivariate regression, this protocol modification did not result in more mature oocytes available for vitrification (coefficient –0.25, CI –1.86–1.36, p-value = 0.76). Limitations, reasons for caution These data should be interpreted with caution because of the retrospective design and limited sample. Although more oocytes were obtained with a long GnRH agonist protocol we have no data on livebirth in women returning to use their oocytes to support the choice for a specific OS protocol for POC. Wider implications of the findings: Although oocyte yield in the context of POC is an important parameter that may be modulated by the choice of OS protocol, the ultimate outcome measure of a successful POC program is livebirth after oocyte vitrification. Future research of oocyte parameters reflecting oocyte quality is paramount. Trial registration number Not applicable

O-161 Cumulative live birth rate after a freeze-all approach in women with polycystic ovaries: does the PCOS phenotype have an impact?

Abstract Study question Do cumulative live birth rates (CLBR) differ between PCOS phenotypes when a freeze-all strategy is used to prevent OHSS after ovarian stimulation (OS)? Summary answer When conventional-dose OS resulted in high response, a CLBR of ∼ 70% was observed after “freeze-all” in women with PCOS, irrespective of their phenotype. What is known already Previous observational studies have shown that CLBR in women with PCOS who undergo assisted reproductive technologies (ART) may depend on their phenotype. When OS was performed with caution to avoid ovarian hyperresponse, CLBR was lower in women with a hyperandrogenic PCOS phenotype. However, when women with PCOS do exhibit hyperresponse and a freeze-all strategy is used, the impact of the PCOS phenotype on the clinical outcome of the ART cycle is unclear. Study design, size, duration This is a single-centre, retrospective cohort study including 422 women with polycystic ovary syndrome (PCOS) as defined by Rotterdam criteria or PCO-like ovarian morphology-only (PCOM) in whom a freeze-all strategy was applied after GnRH agonist triggering in the context of hyperresponse defined as ³19 follicles of ³11mm in their first or second IVF-ICSI cycle between January 2015 and December 2019 in a tertiary referral hospital. Participants/materials, setting, methods PCOS phenotype was based on hyperandrogenism (H), ovulatory dysfunction (O) and PCO-like ovarian morphology (P). Ovarian stimulation was performed with rFSH or HPhMG, adjusted to BMI. The primary outcome was cumulative live birth rate (CLBR) resulting from the transfer of all cryopreserved embryos from the same IVF-ICSI cycle. Patient and cycle characteristics and laboratory and clinical data were analysed. Data were analysed by multivariate logistic regression adjusting for covariates. Main results and the role of chance In total, 91/422 (21.6%) patients had PCOS phenotype A (HOP); 33 (7.8%) had phenotype C (HP), 161/422 (38.2%) had phenotype D (OP) and 137/422 (32.5%) had PCOM (n = 137). BMI, AMH and AFC were significantly different between phenotype groups (p < 0.001), and highest in PCOS phenotype A. The type of gonadotropin used, as well as the mean daily and total stimulation dose were comparable for all groups. The mean number of retrieved oocytes was comparable among groups (22.4±10.8 for phenotype A, 21.4±7.1 for phenotype C, 20.4±7.8 for phenotype D and 22.2±9.7 for PCOM; p = 0.46). The mean number of embryos available for vitrification differed significantly (4.4±3.7, 5.7±3.4, 5.7±3.4 and 5.2±3.6, respectively; p = 0.005). Following the first frozen embryo transfer, LBR was comparable among groups (41.5%, 43.3%, 49.3% and 38.5%, respectively; p = 0.31). Unadjusted CLBR was also similar (69.2%, 69.7%, 79.5% and 67.9%, respectively; p = 0.11). The multivariate logistic regression model adjusting for age, BMI, number of oocytes and embryo stage (cleavage vs. blastocyst stage) confirmed that the PCOS/PCOM phenotype did not have any impact on CLBR (OR 0.80, CI 0.28-2.29 (phenotype C); OR 1.40, CI 0.67-2.90 (phenotype D); OR 0.65, CI 0.31-1.34 (PCOM); p = 0.1, with phenotype A as reference). Limitations, reasons for caution These data should be interpreted with caution as the retrospective nature of the study holds the possibility of unmeasured confounding factors. The results cannot be generalised to all ART cycles in women with polycystic ovaries as they pertain to those cycles where OS leads to hyperresponse. Wider implications of the findings In subfertile women with PCOS eligible for ART, hyperresponse after OS confers excellent cumulative live birth rates when a freeze-all strategy is used, eliminating unfavourable clinical outcomes that had previously been observed in hyperandrogenic PCOS women after mild OS targeting normal ovarian response and fresh embryo transfer. Trial registration number not applicable

O-171 Altered meiotic spindle morphology and composition in in vitro matured oocytes

Abstract Study question How does the meiotic spindle tubulin PTMs of MII oocytes matured in vitro compare to that of MII oocytes matured in vivo? Summary answer MII cultured in vitro present detyrosinated tubulin in the spindle microtubules, while MII oocytes matured in vivo do not. What is known already A functional spindle is required for chromosomal segregation during meiosis, but the role of tubulin post-translational modifications (PTMs) in spindle meiotic dynamics remains poorly characterized. In contrast with GVs matured in vitro within the cumulus oophorous, in vitro maturation of denuded GVs to the MII stage (GV-MII) is associated with spindle abnormalities, chromosome misalignment and compromised developmental potential. Although aneuploidy rates in GV-MII are not higher than in vivo matured MII, disorganized chromosomes may contribute to compromised developmental potential. However, to date, spindle PTMs morphology of GV-MII has not been compared to that of in vivo cultured MII oocytes. Study design, size, duration GV (n = 125), and MII oocytes (n = 24) were retrieved from hormonally stimulated women, aged 20 to 35 years old. GVs were matured to the MII stage in vitro in G-2 PLUS medium for 30h; the maturation rate was 68,2%; the 46 GV-MII oocytes obtained were vitrified, stored, and warmed before fixing and subjecting to immunofluorescent analysis. In vivo matured MII oocytes donated to research were used as controls. Participants/materials, setting, methods Women were stimulated using a GnRH antagonist protocol, with GnRH agonist trigger. Trigger criterion was ≥2 follicles ≥18mm; oocytes were harvested 36h later. Spindle microtubules were incubated with antibodies against alpha tubulin and tubulin PTMs (acetylation, tyrosination, polyglutamylation, Δ2-tubulin, and detyrosination); chromosomes were stained with Hoechst 33342 and samples subjected to confocal immunofluorescence microscopy (ZEISS LSM780), with ImageJ software analysis. Differences in spindle morphometric parameters were assessed by non-parametric Kruskal–Wallis and Fisher’s exact tests. Main results and the role of chance Qualitatively, Δ2-tubulin, tyrosination and polyglutamylation were similar for both groups. Acetylation was also present in both groups, albeit in different patterns: while in vivo matured MII oocytes showed acetylation at the poles, GV-MII showed a symmetrical distribution of signal intensity, but discontinuous signal on individual microtubule tracts, suggesting apparent islands of acetylation. In contrast, detyrosination was detected in in vivo matured MII oocytes but was absent from GV-MII. Regarding spindle pole morphology, of the four possible phenotypes described in the literature (double flattened and double focused; flattened-focused, focused-flattened, with the first word characterizing the cortex side of the spindle), we observed double flat shaped spindle poles in 86% of GV-MII oocytes (25/29) as opposed to 40.5% (15/37) for the in vivo matured MII oocytes (p = 0.0004, Fisher’s exact test). Further morphometric analysis of the spindle size (maximum projection, major and minor axis length) and the metaphase plate position (proximal to distal ratio, angle) revealed decreased spindle size in GV-MII oocytes (p = 0.019, non parametric Kruskal- Wallis test). Limitations, reasons for caution Oocytes retrieved from hyperstimulation cycles could be intrinsically impaired since they failed to mature in vivo. Our conclusions should not be extrapolated to IVM in non-stimulated cycles, as in this model, the cumulus oophorus is a major factor in oocyte maturation and correlation with spindle dynamics has been inferred. Wider implications of the findings The metaphase II spindle stability compared to the mitotic or metaphase I meiotic one justifies the presence of PTMs such as acetylation and glutamylation, which are found in stable, long-lived microtubules. The significance of the absence of detyrosinated microtubules in the MII-GV group remains to be determined Trial registration number not applicable

P–690 Clinical predictors of a high oocyte maturation rate in IVF treatment cycles

Abstract Study question Which clinical parameters predict a high oocyte maturation rate in patients undergoing IVF treatment? Summary answer Time between oocyte collection and insemination demonstrated significant association with oocyte maturation and represents a parameter that could be optimised in IVF cycles. What is known already Oocyte maturation is an important factor determining IVF outcomes and can be a rate-limiting step for patients undergoing treatment. A number of clinical and laboratory variables may affect this process, including the choice of trigger prior to oocyte collection, and certain laboratory procedures. Identification of which of these are predictors of maturation in individual centres enables local protocols to be optimised. Study design, size, duration This is a retrospective study of 714 oocyte collections from 661 women between January 2020 to November 2020 treated in a large, single centre in the UK. Subsequent fertilisation on fresh oocytes consisted of 371 IVF and 343 ICSI cycles. Participants/materials, setting, methods Patient and treatment data was collected by clinical staff at time of treatment. Either GnRH agonist, hCG or double trigger were administered 36 hours before collection. Prior to ICSI, oocyte maturation was assessed by visualisation of polar body (PB) extrusion. After IVF, the number of 2PNs plus unfertilised oocytes with PB extrusion were assessed. Univariate analyses consisted of Mann-Whitney test, t-test, Fisher’s Exact test or ANOVA. Potential predictors were investigated by logistic regression. Main results and the role of chance The end point was maturation rate, defined as high (greater or equal to 70%) or low (less than 70%). Factors predictive of a high rate included insemination more than 4 hours after collection. Oocytes inseminated over 4 hours post-collection displayed significantly higher maturation rates than oocytes inseminated less than 2 hours after collection (69% and 61% respectively; P = 0.01). Oocytes inseminated between 2–4 hours also had higher maturation than those inseminated less than 2 hours post-collection, but this did not reach significance (67% and 61%, respectively; P = 0.06). Further, oocytes fertilised by ICSI had significantly higher maturation than conventional IVF (77% and 67%, respectively, P < 0.001). No significant difference in oocyte maturation between triggers was observed. Similarly, neither age, AMH, a diagnosis of PCOS or number of oocytes collected predicted oocyte maturation in univariate analysis. Logistic regression analysis showed only time between oocyte collection and insemination (aOR 2.12; 95% CI 1.03–4.38; P = 0.04) to be a significant independent predictor. Limitations, reasons for caution Varying means of data collection across clinics and between clinical staff inevitably leads to provision of incomplete data and should be taken into consideration alongside interpretation. Prescription bias of specific triggers to certain patient demographics should be noted. Wider implications of the findings: Collectively, these results suggest that greater time between oocyte collection and insemination could be recommended to IVF clinics that wish to optimise their oocyte maturation. Triggering final maturation with GnRH agonist versus hCG or dual trigger did not have a significant effect on oocyte maturation when adjusted for confounders. Trial registration number Not applicable

P-777 Comparison of GnRH-a trigger and GnRH-a plus low-dose HCG trigger for high ovarian responders in IVF/ICSI: A retrospective study based on propensity score matching

Abstract Study question Does GnRH agonist trigger for high responders during IVF/ICSI cycles improve the number of good-quality embryos, the incidence of moderate-to-severe OHSS, and pregnancy outcome compared to GnRH-a plus low-dose HCG? Summary answer GnRH-a trigger alone can effectively reduce the incidence of moderate-to-severe OHSS in women with high ovarian responses without affecting embryo quality. What is known already Previous studies have shown conflicting results on the different trigger protocol in high responders in IVF/ICSI outcomes, and as for women with high ovarian response, there is little known about the effects of GnRH-a plus low-dose HCG versus GnRH-a alone on oocytes maturation, the rate of good quality embryos, the incidence of moderate-to-severe OHSS, and pregnancy outcome during IVF/ICSI cycles. Study design, size, duration A retrospective analysis was conducted on patients with high ovarian response who received IVF/ICSI treatment with a flexible GnRH antagonist regimen, at the Center of Reproductive Medicine, Chengdu Jinjiang Hospital for Maternal and Child Health Care, from January 1 2017 to December 31 2018. Using 1:1 propensity score matching, 513 cases entered each group (a total of 1,026 females). Participants/materials, setting, methods The high responders were included and assigned to groups A (0.2 mg triptorelin) and B (0.2 mg triptorelin plus 2000 IU HCG) for final oocyte maturation. Their basic clinical characteristics, information about controlled ovarian stimulation cycle, embryologic data, and pregnancy outcome in FET were retrospectively compared. The main outcome measures of the study were the rate of good-quality embryos, the number of available embryos, the incidence of moderate-to-severe OHSS, and the cumulative live-birth rate. Main results and the role of chance Using 1:1 propensity score matching, 513 females were included in each group. No significant differences in baseline clinical data were found between the two groups, including age at diagnosis, spouse’s age, the duration of infertility, the infertility type, and the cause of infertility, BMI, anti-Müllerian hormone (AMH) levels, and the antral follicle count (AFC) (p > 0.05). None significant differences were found in the total doses of gonadotropin (Gn), the duration of ovarian stimulation, serum P and LH levels on the trigger day, the number of oocytes retrieved, the rate of 2PN embryos, and the rate of good-quality embryos (p > 0.05). The serum E2 level on the trigger day in group A was significantly higher than that in group B (p < 0.001). Women in group A had a lower incidence rate of moderate-to-severe OHSS than individuals in group B (p < 0.001). There was a non-significant difference in the cumulative live-birth rate between the two groups (p > 0.05). Limitations, reasons for caution As this is a retrospective study that uses data initially collected for other purposes, limitations may exist in the selection, implementation, and measurement biases that cannot be avoided. However, our study underlies the need for further prospective, multi-center joint-controlled studies to validate these findings. Wider implications of the findings This study demonstrates that GnRH-a alone can reduce the incidence of moderate-to-severe OHSS without harming embyro quality in women with high ovarian response. These findings need further prospective validations in hyperresponsive populations by multi-center, large-sample, randomized controlled studies. Trial registration number N/A

P–777 Comparison of GnRH-a trigger and GnRH-a plus low-dose HCG trigger for high ovarian responders in IVF/ICSI: A retrospective study based on propensity score matching

Abstract Study question Does GnRH agonist trigger for high responders during IVF/ICSI cycles improve the number of good-quality embryos, the incidence of moderate-to-severe OHSS, and pregnancy outcome compared to GnRH-a plus low-dose HCG? Summary answer GnRH-a trigger alone can effectively reduce the incidence of moderate-to-severe OHSS in women with high ovarian responses without affecting embryo quality. What is known already Previous studies have shown conflicting results on the different trigger protocol in high responders in IVF/ICSI outcomes, and as for women with high ovarian response, there is little known about the effects of GnRH-a plus low-dose HCG versus GnRH-a alone on oocytes maturation, the rate of good quality embryos, the incidence of moderate-to-severe OHSS, and pregnancy outcome during IVF/ICSI cycles. Study design, size, duration A retrospective analysis was conducted on patients with high ovarian response who received IVF/ICSI treatment with a flexible GnRH antagonist regimen, at the Center of Reproductive Medicine, Chengdu Jinjiang Hospital for Maternal and Child Health Care, from January 1 2017 to December 31 2018. Using 1:1 propensity score matching, 513 cases entered each group (a total of 1,026 females). Participants/materials, setting, methods The high responders were included and assigned to groups A (0.2 mg triptorelin) and B (0.2 mg triptorelin plus 2000 IU HCG) for final oocyte maturation. Their basic clinical characteristics, information about controlled ovarian stimulation cycle, embryologic data, and pregnancy outcome in FET were retrospectively compared. The main outcome measures of the study were the rate of good-quality embryos, the number of available embryos, the incidence of moderate-to-severe OHSS, and the cumulative live-birth rate. Main results and the role of chance Using 1:1 propensity score matching, 513 females were included in each group. No significant differences in baseline clinical data were found between the two groups, including age at diagnosis, spouse’s age, the duration of infertility, the infertility type, and the cause of infertility, BMI, anti-Müllerian hormone (AMH) levels, and the antral follicle count (AFC) (p > 0.05). None significant differences were found in the total doses of gonadotropin (Gn), the duration of ovarian stimulation, serum P and LH levels on the trigger day, the number of oocytes retrieved, the rate of 2PN embryos, and the rate of good-quality embryos (p > 0.05). The serum E2 level on the trigger day in group A was significantly higher than that in group B (p < 0.001). Women in group A had a lower incidence rate of moderate-to-severe OHSS than individuals in group B (p < 0.001). There was a non-significant difference in the cumulative live-birth rate between the two groups (p > 0.05). Limitations, reasons for caution As this is a retrospective study that uses data initially collected for other purposes, limitations may exist in the selection, implementation, and measurement biases that cannot be avoided. However, our study underlies the need for further prospective, multi-center joint-controlled studies to validate these findings. Wider implications of the findings: This study demonstrates that GnRH-a alone can reduce the incidence of moderate-to-severe OHSS without harming embyro quality in women with high ovarian response. These findings need further prospective validations in hyperresponsive populations by multi-center, large-sample, randomized controlled studies. Trial registration number N/A

P–667 Dydrogesterone supplementation in cycles triggered with lone GnRH agonist for final oocyte maturation resulted in an acceptable pregnancy rate

Abstract Study question Can luteal oral Dydrogesterone (Duphaston) supplementation in an antagonist cycle after a lone GnRH agonist trigger rescue the luteal phase, allowing the possibility to peruse with fresh embryo transfer? Summary answer Functionality of the luteal phase in an antagonist cycle after a lone GnRH agonist trigger can be restored by adding Duphaston to conventional luteal support. What is known already Ovarian hyperstimulation syndrome (OHSS) is dramatically reduced when using antagonist cycle with lone GnRH agonist trigger before ovum pick up. This trigger induces short luteinizing hormone (LH) and follicle-stimulating hormone (FSH) peaks, associated with reduced progesterone and estrogen levels during the luteal phase. They cause an inadequate luteal phase and a significantly reduced implantation rate leading to a freeze all practice in those cycles. Study design, size, duration A retrospective cohort study. The study group (n = 123) included women that underwent in vitro fertilization cycles from January 2017 to May 2020. Patients received a GnRH-antagonist with a lone GnRH-agonist trigger due to imminent OSHH. The control group (n = 374) included patients under 35 years old that, during the same time period, underwent a standard antagonist protocol with a dual trigger of a GnRH-agonist and hCG. Participants/materials, setting, methods Study patients were given Dydrogesterone (Duphaston) in addition to micronized progesterone vaginal pills (Utrogestan) for luteal support (Duphaston group). Controls were treated conventionally with Utrogestan for luteal phase support (hCG group). The outcomes measured were pregnancy rate and OHSS events. Main results and the role of chance Our study was the first to evaluate the addition of Duphaston to standard luteal phase support in an antagonist cycle triggered by a lone GnRH agonist before a fresh embryo transfer. The mean number of oocytes retrieved and estradiol plasma levels were significantly higher in the Duphaston group than in the hCG group (16.9 ±7.7 vs. 10.8 ± 5.3 and 11658 ± 5280 pmol/L vs. 6048 ± 3059 pmol/L, respectively). The fertilization rate was comparable between the two groups. The mean number of embryos transferred and the clinical pregnancy rate were also comparable between groups (1.5 ± 0.6 vs 1.5 ± 0.5 and 46.3% vs 40.9%, respectively). No OHSS event was reported in either group. Limitations, reasons for caution This retrospective study may carry an inherent selection and information bias, derived from medical record coding. An additional limitation was the choice of physician for the lone GnRH trigger, which may have introduced a selection bias and another potential caveat was the relatively small sample size of our study groups. Wider implications of the findings: The addition of Duphaston to conventional luteal support could effectively salvage the luteal phase without increasing the risk for OHSS. This enables, to peruse in those cycle, with fresh embryo transfer, avoiding the need to freeze all the embryos and postponed embryo transfer. Leading to lower psychological burden and costs. Trial registration number 0632–20-HMO

Comparison of hCG Versus Gonadotropin-Releasing Hormone Agonist to Induce Oocyte Maturation in Assisted Reproductive Technology Cycles: A Retrospective Cohort Study

OBJECTIVE: To compare some cycle characteristics and outcomes using a protocol consisting of a GnRH agonist trigger or hCG trigger after cotreatment with GnRH antagonist.STUDY DESIGN: Thirty-three patients under 35 years of age with polycystic ovarian syndrome, polycystic ovarian morphology, or previous high response who underwent ovulation trigger by GnRH agonist trigger and 132 patients under 35 years of age with the polycystic ovarian syndrome, polycystic ovarian morphology, or previous high response who underwent ovulation trigger by hCG for IVF treatment. Patients were non-randomly assigned to an ovarian stimulation protocol consisting of either GnRH agonist trigger after cotreatment with GnRH antagonist (study group) or hCG trigger after antagonist protocol (control group).RESULTS: The positive pregnancy test was obtained in 70 women in the control group whereas in 13 cases in the study group (p=0.161). No case in the study group needed hospitalization whereas there were 15 cases in the control group who were required to be hospitalized due to ovarian hyperstimulation related symptoms (p=0.04).CONCLUSIONS: The use of a protocol consisting of a GnRH agonist trigger after GnRH antagonist cotreatment and freeze-all strategy reduces the risk of ovarian hyperstimulation syndrome in high-risk patients undergoing IVF without affecting pregnancy rates.

Does the timing of cabergoline administration impact rates of ovarian hyperstimulation syndrome?

ObjectiveDoes the timing of cabergoline administration impact the rate of mild/moderate ovarian hyperstimulation syndrome in women with a GnRH agonist trigger?MethodsWe conducted a retrospective cohort analysis of 285 in-vitro fertilization patients at risk of OHSS who received a GnRH agonist trigger from 2011 to 2019 at McGill University Health Centre. Group 1 (Trig, n=101) began taking cabergoline 0.5 mg orally for 7 days at the time of GnRH agonist trigger, while Group 2 (Retriev, n=184) started taking cabergoline on the day of oocyte retrieval. The rates of OHSS were then compared between the groups using analysis of variance and chi-square analysis, where appropriate.ResultsThe baseline demographic characteristics of the two groups were similar. Trig appeared to be at a slightly higher risk of OHSS based on a significantly higher antral follicle count (20.2±4.2 vs. 19.0±4.3; P=0.02), higher number of stimulated follicles >10 mm at trigger (25.7±7.0 vs. 22.8±8.3, P=0.003), and higher peak serum E2 level (17,325±2,542 vs. 14,822±3,098; P=0.0001). The Trig group had lower rates of mild and moderate OHSS (24% vs. 36%; P=0.045). Neither group had any patients who developed severe OHSS. Trig had fewer patients presenting with pelvic free fluid (13% vs. 23%; P=0.03), lower hematocrit (37.8±4.8% vs. 40.5±4.2%; P=0.0001), higher albumin concentrations (30.4±2.7 vs. 29.5±2.0; P=0.01), and lower potassium concentrations (3.9±0.5 vs. 4.2±0.7; P=0.0002).ConclusionCabergoline at the time of trigger as compared to the time of collection should be investigated to assess its role in reducing the rates of mild/moderate OHSS.

Does gonadotropin-releasing hormone agonist cause luteolysis by inducing apoptosis of the human granulosa-luteal cells?

To evaluates the effect of different modes of final follicular maturation triggering on the degree of apoptosis of granulosa cells (GCs) and the potential effect on progesterone secretion. Thirty patients undergoing controlled ovarian hyperstimulation for IVF who received hCG, GnRH agonist, or dual trigger for final follicular maturation were included in the study. Granulosa cells were obtained at the time of oocyte retrieval. The proportion of apoptotic cells was evaluated via TUNEL and immunohistochemistry. The proportion of apoptotic cells was significantly higher in the GnRH agonist-alone group compared to hCG-alone and the dual trigger groups (13.5 ± 1.5% vs. 7.8% ± 1.8 vs. 10.1% ± 2, respectively, P < 0.01). Moreover, the expression of active-caspase-3 was also significantly increased in the GnRH agonist-alone group compared with the hCG-alone and the dual trigger groups (15.5% ± 2.9 vs. 8.4% ± 1.6 vs. 12.7% ± 2.6, respectively, P < 0.01). The progesterone levels measured in the granulosa-luteal cell culture medium after 24 h of incubation were similar between the three groups. The levels of apoptosis are increased after GnRH agonist/dual trigger. The increased apoptosis might be one of the culprit of the subsequent premature demise of the corpus luteum post GnRH agonist trigger.

How far is too far? Does time interval between GnRH antagonist and GnRH agonist trigger in GnRH antagonist cycles matter?

Research questionWhat is a suitable time interval between the last GnRH antagonist exposure and GnRH agonist (GnRHa) triggering for final follicular maturation? DesignA retrospective cohort study including 413 patients undergoing GnRH antagonist cycles in which GnRHa trigger was used, either solely or as a dual trigger. The primary outcome measure was the follicle/mature oocyte ratio. Cycles were analysed according to the time interval between the last GnRH antagonist exposure and the GnRHa triggering: Group 1 included patients with a 12–14 h interval; Group 2: 7–10 h interval; Group 3: 5–6 h interval and Group 4: 2–4 h interval. LH concentration was measured 11–13 h post-GnRHa injection. ResultsMedian LH value was 65 IU/l. There was a weak but significant correlation between basal LH and the LH surge (R2 = 0.137, P < 0.001). Although square root LH values differed significantly between study groups (P < 0.001; higher in Groups 2 and 3), the follicle/mature oocyte ratio was not different across the four antagonist–agonist interval groups and no correlation was detected between the post-trigger LH concentration and the follicle/oocyte ratio (R2 = 0.011). In a model integrating age, day 3 FSH concentration, maximal oestradiol and body mass index along with the study groups, none of these factors was significantly related to the follicle/mature oocyte outcome ratio. Insufficient surge (LH < 15 IU/l) occurred in 14 (3.4%) cases. Rates of insufficient LH surge did not differ significantly between the groups (2.4%, 3.2%, 3.4% and 7.1% in Groups 1 to 4, respectively; P = 0.5). ConclusionsLH concentrations post-GnRHa trigger differ in regard to antagonist–agonist intervals, but the follicle/mature oocyte ratio achieved was not affected.

Severe ovarian hyperstimulation syndrome with GnRH agonist trigger during the COVID pandemic: lessons learned from an unusual case

Background: Injectable gonadotropins stimulate multi-follicular recruitment and allows retrieval of multiple oocytes for assisted reproduction. The widespread utilization of gonadotropin releasing hormone agonist (GnRHa) to induce oocyte maturation for oocyte retrieval has nearly eliminated the risk of severe ovarian hyperstimulation syndrome (OHSS), and only a few cases have been reported in the literature. The rarity of severe OHSS may lead to the mistaken conclusion that gonadotropin stimulation can be safely administered with limited monitoring, even in high-risk patients. We present an unusual case of a woman with limited monitoring due to the COVID pandemic who developed severe OHSS before GnRH agonist trigger and oocyte. Case Presentation: A 29-year-old nulliparous woman with polycystic ovarian syndrome (PCOS) initiated ovarian stimulation for oocyte retrieval. She had a robust initial response, and developed worsening abdominal pain, bloating, nausea, vomiting, and decreased appetite before retrieval. GnRH agonist was given to “trigger ovulation and retrieval scheduled due to the low reported incidence of severe OHSS. Symptoms progressed, and on the morning of retrieval, ultrasound demonstrated bilaterally enlarged ovaries &gt;10cm and 48 oocytes were retrieved for a planned cryo-all cycle. She was hospitalized on the day of retrieval for severe OHSS and had two large-volume paracenteses. She was stable and discharged home by day 5, and symptoms markedly improved with the onset of menses. She has an ongoing pregnancy from her first frozen embryo transfer. Conclusion: We add a rare case of severe OHSS with a GnRHa trigger and cryo-all protocol with the onset of symptoms before GnRH agonist administration. Although rare, severe OHSS may still occur with a GnRHa trigger, and caution is needed when an initial robust response is identified. Here we also provide an opportunity to review the important patient risk factors for the development of OHSS and measures to reduce the risk in excessive responders.

Optimising Follicular Development, Pituitary Suppression, Triggering and Luteal Phase Support During Assisted Reproductive Technology: A Delphi Consensus.

BackgroundA Delphi consensus was conducted to evaluate global expert opinions on key aspects of assisted reproductive technology (ART) treatment.MethodsTen experts plus the Scientific Coordinator discussed and amended statements plus supporting references proposed by the Scientific Coordinator. The statements were distributed via an online survey to 35 experts, who voted on their level of agreement or disagreement with each statement. Consensus was reached if the proportion of participants agreeing or disagreeing with a statement was >66%.ResultsEighteen statements were developed. All statements reached consensus and the most relevant are summarised here. (1) Follicular development and stimulation with gonadotropins (n = 9 statements): Recombinant human follicle stimulating hormone (r-hFSH) alone is sufficient for follicular development in normogonadotropic patients aged <35 years. Oocyte number and live birth rate are strongly correlated; there is a positive linear correlation with cumulative live birth rate. Different r-hFSH preparations have identical polypeptide chains but different glycosylation patterns, affecting the biospecific activity of r-hFSH. r-hFSH plus recombinant human LH (r-hFSH:r-hLH) demonstrates improved pregnancy rates and cost efficacy versus human menopausal gonadotropin (hMG) in patients with severe FSH and LH deficiency. (2) Pituitary suppression (n = 2 statements): Gonadotropin releasing hormone (GnRH) antagonists are associated with lower rates of any grade ovarian hyperstimulation syndrome (OHSS) and cycle cancellation versus GnRH agonists. (3) Final oocyte maturation triggering (n=4 statements): Human chorionic gonadotropin (hCG) represents the gold standard in fresh cycles. The efficacy of hCG triggering for frozen transfers in modified natural cycles is controversial compared with LH peak monitoring. Current evidence supports significantly higher pregnancy rates with hCG + GnRH agonist versus hCG alone, but further evidence is needed. GnRH agonist trigger, in GnRH antagonist protocol, is recommended for final oocyte maturation in women at risk of OHSS. (4) Luteal-phase support (n = 3 statements): Vaginal progesterone therapy represents the gold standard for luteal-phase support.ConclusionsThis Delphi consensus provides a real-world clinical perspective on the specific approaches during the key steps of ART treatment from a diverse group of international experts. Additional guidance from clinicians on ART strategies could complement guidelines and policies, and may help to further improve treatment outcomes.

Superovulation with gonadotropin-releasing hormone agonist or chorionic gonadotropin for ovulation trigger differentially affects leukocyte populations in the peri-implantation mouse uterus

To investigate the effect of superovulation with human chorionic gonadotropin (hCG) or gonadotropin-releasing hormone agonist (GnRHa) trigger on leukocyte density and expression of leukocyte-specific genes in the peri-implantation period in the mouse uterus. Laboratory research. University laboratory facility. Female mice were mated to fertile male mice in one of three protocols: (1) natural mating or mating following injection with pregnant mare serum gonadotropin followed by trigger with (2) GnRHa or (3) hCG. Female mice were killed prior to implantation, 3 days after ovulation (E3.5), and the ovaries and uterine tissue were collected. Total RNA was isolated and assayed using quantitative reverse transcription polymerase chain reaction, and the uterine tissue was stained for histologic analysis of immune cell markers. Endometrial leukocyte (CD45) and vessel density (CD31) by immunohistochemical staining; expression of leukocyte markers CD11b, CD335, and CD22, by quantitative reverse transcription polymerase chain reaction in the whole uterine tissue. Superovulation decreased (compared with controls) the endometrial leukocyte density, based on the number of cells staining for CD45, and endometrial vessel density, based on the number of cells staining for CD31. Leukocyte density was additionally decreased in the GnRHa trigger group compared with that in the hCG trigger group. Superovulation with hCG and GnRHa triggers decreased the uterine expression of the B-cell marker CD22 compared with controls. The expression of the natural killer cell marker CD11b was decreased by the hCG trigger but not by the GnRHa. Abundance of mRNA encoding the CD335 natural killer cell marker was not affected by superovulation or trigger agent. In mice, superovulation with the GnRHa trigger compared with that with the hCG trigger differentially alters key immunologic factors in the uterine peri-implantation. These altered immunologic factors have roles in angiogenesis that may assist in elucidating the effects of assisted reproductive technologies on implantation efficiency and fetal growth and development.

Ovarian stimulation for oocyte donation: a systematic review and meta-analysis.

Since its introduction in the 1980s, oocyte donation (OD) has been largely integrated into ART. Lately, both demand and the indications for OD have increased greatly. Oocyte donors are healthy and potentially fertile women undergoing voluntarily ovarian stimulation (OS). Selection of the optimal type of stimulation is of paramount importance in order to achieve the most favourable outcomes for the oocyte recipients, but most importantly for the safety of the oocyte donors. This is the first systematic review (SR) with the objective to summarize the current evidence on OS in oocyte donors. The scope of this SR was to evaluate the OD programme by assessing four different aspects: how to assess the ovarian response prior to stimulation; how to plan the OS (gonadotrophins; LH suppression; ovulation trigger; when to start OS); how to control for the risk of ovarian hyperstimulation syndrome (OHSS) and other complications; and the differences between the use of fresh versus vitrified donated oocytes. A systematic literature search was conducted in May 2020, according to PRISMA guidelines in the databases PubMed and Embase, using a string that combined synonyms for oocytes, donation, banking, freezing, complications and reproductive outcomes. Studies reporting on the safety and/or efficacy of OS in oocyte donors were identified. The quality of the included studies was assessed using ROBINS-I and ROB2. Meta-analysis was performed where appropriate. Data were combined to calculate mean differences (MD) for continuous variables and odd ratios (OR) for binary data with their corresponding 95% CIs. Heterogeneity between the included studies was assessed using I2 and tau statistics. In total, 57 manuscripts were selected for the review, out of 191 citations identified. Antral follicle count and anti-Müllerian hormone levels correlate with ovarian response to OS in OD but have limited value to discriminate donors who are likely to show either impaired or excessive response. Five randomized controlled trials compared different type of gonadotrophins as part of OS in oocyte donors; owing to high heterogeneity, meta-analysis was precluded. When comparing different types of LH control, namely GnRH antagonist versus agonist, the studies showed no differences in ovarian response. Use of progesterone primed ovarian stimulation protocols has been evaluated in seven studies: the evidence has shown little or no difference, compared to GnRH antagonist protocols, in mean number of retrieved oocytes (MD 0.23, [95% CI 0.58-1.05], n = 2147; 6 studies; I2 = 13%, P = 0.33) and in clinical pregnancy rates among recipients (OR 0.87 [95% CI 0.60-1.26], n = 2260, I2 = 72%, P < 0.01). There is insufficient evidence on long-term safety for babies born. GnRH agonist triggering is the gold standard and should be used in all oocyte donors, given the excellent oocyte retrieval rates, the practical elimination of OHSS and no differences in pregnancy rates in recipients (four studies, OR 0.86, 95%CI 0.58-1.26; I2 = 0%). OS in OD is a safe procedure with a low rate of hospitalization after oocyte retrieval. The use of a levonorgestrel intrauterine device or a progestin contraceptive pill during OS does not impact the number of oocytes retrieved or the clinical pregnancy rate in recipients. Ultrasound monitoring seems enough for an adequate follow up of the stimulation cycle in OD. Use of fresh versus vitrified donated oocytes yielded similar pregnancy outcomes. This update will be helpful in the clinical management of OS in OD based on the most recent knowledge and recommendations, and possibly in the management of women under 35 years undergoing oocyte vitrification for social freezing, owing to the population similarities. More clinical research is needed on OS protocols that are specifically designed for OD, especially in term of the long-term safety for newborns, effective contraception during OS, and treatment satisfaction.

The exogenous progesterone-free luteal phase: two pilot randomized controlled trials in IVF patients

Research questionIs the reproductive outcome similar after gonadotrophin-releasing hormone agonist (GnRHa) trigger followed by luteal human chorionic gonadotrophin (HCG) boluses compared with HCG trigger and a standard luteal phase support (LPS)? DesignTwo open-label pilot randomized controlled trials (RCT) with 250 patients from 2014 to 2019, with a primary outcome of ongoing pregnancy per embryo transfer. Patients with ≤13 follicles on the trigger day were randomized (RCT 1) to: Group A (n = 65): GnRHa trigger followed by a bolus of 1500 IU HCG s.c. on the oocyte retrieval day (ORD) and 1000 IU HCG s.c. 4 days later, and no vaginal LPS; or Group B (n = 65): 6500 IU HCG trigger, followed by a standard vaginal progesterone LPS. Patients with 14–25 follicles on the trigger day were randomized (RCT 2) to Group C (n = 60): GnRHa trigger followed by 1000 IU HCG s.c. on ORD and 500 IU HCG s.c. 4 days later, and no vaginal LPS; or Group D (n = 60): 6500 IU HCG trigger and a standard vaginal LPS. ResultsIn RCT 1, the ongoing pregnancy rate was 44% (22/50) in the GnRHa group versus 46% (25/54) in the HCG trigger group (RR 0.95, 95% CI 0.62–1.45). No ovarian hyperstimulation syndrome (OHSS) was seen in Groups A or B. In RCT 2, the ongoing pregnancy rate was 51% (25/49) in the GnRHa group versus 60% (31/52) in the HCG trigger group (RR 0.86, 95% CI 0.60–1.22). The OHSS rates were 3.3% and 6.7%, respectively. ConclusionsAlthough a larger-scale study is needed before standard clinical implementation, the present study supports that the exogenous progesterone-free LPS is efficacious, simple and patient-friendly.

Luteal Support with very Low Daily Dose of Human Chorionic Gonadotropin after Fresh Embryo Transfer as an Alternative to Cycle Segmentation for High Responders Patients Undergoing Gonadotropin-Releasing Hormone Agonist-Triggered IVF.

The segmentation of the in vitro fertilization (IVF) cycle, consisting of the freezing of all embryos and the postponement of embryo transfer (ET), has become popular in recent years, with the main purpose of preventing ovarian hyperstimulation syndrome (OHSS) in patients with high response to controlled ovarian stimulation (COS). Indeed cycle segmentation (CS), especially when coupled to a GnRH-agonist trigger, was shown to reduce the incidence of OHSS in high-risk patients. However, CS increases the economic costs and the work amount for IVF laboratories. An alternative strategy is to perform a fresh ET in association with intensive luteal phase pharmacological support, able to overcome the negative effects of the GnRH-agonist trigger on the luteal phase and on endometrial receptivity. In order to compare these two strategies, we performed a retrospective, real-life cohort study including 240 non-polycystic ovarian syndrome (PCO) women with expected high responsiveness to COS (AMH >2.5 ng/mL), who received either fresh ET plus 100 IU daily human chorionic gonadotropin (hCG) as luteal support (FRESH group, n = 133), or cycle segmentation with freezing of all embryos and postponed ET (CS group, n = 107). The primary outcomes were: implantation rate (IR), live birth rate (LBR) after the first ET, and incidence of OHSS. Overall, significantly higher IR and LBR were observed in the CS group than in the FRESH group (42.9% vs. 27.8%, p < 0.05 and 32.7% vs. 19.5%, p < 0.05, respectively); the superiority of CS strategy was particularly evident when 16–19 oocytes were retrieved (LBR 42.2% vs. 9.5%, p = 0.01). Mild OHSS appeared with the same incidence in the two groups, whereas moderate and severe OHSS forms were observed only in the FRESH group (1.5% and 0.8%, respectively). In conclusion, in non-PCO women, high responders submitted to COS with the GnRH-antagonist protocol and GnRH-agonist trigger, CS strategy was associated with higher IR and LBR than the strategy including fresh ET followed by luteal phase support with a low daily hCG dose. CS appears to be advisable, especially when >15 oocytes are retrieved.

Suboptimal response to GnRH agonist trigger: causes and practical management.

GnRH agonist products are used extensively worldwide to trigger ovulation and final oocyte maturation in in vitro fertilization cycles. The purpose of this article is to outline possible causes for a suboptimal response to the GnRH agonist trigger. Risk factors for such a suboptimal response include prolonged hormonal contraceptive use, previous GnRHa-induced pituitary downregulation, a hypogonadotropic/hypogonadal condition, patient error, environmental conditions that may damage the GnRHa product used, GnRH and luteinizing hormone (LH) receptors polymorphisms, low baseline LH and low endogenous serum LH levels on trigger day as well as low BMI. The induction of an adequate LH surge can be ascertained by an LH urine test 12 h post trigger. In most cases, GnRHa trigger elicits effective LH+follicle stimulating hormone surges, resulting in mature, fertilizable oocytes. Clinical awareness to conditions that may predispose to a suboptimal response to the GnRHa trigger may prevent failed oocyte retrial.

Ideal lag time from ovulation to oocyte aspiration using a GnRH agonist trigger

BackgroundIn ART, oocyte maturation (M2) and ovulation is stimulated by a hormonal trigger. For maturation to occur, sufficient “lag time” must elapse between the trigger and aspiration, ranging from 32 to 38 hours. Premature aspiration can result in poor yields; late aspiration risks spontaneous ovulation. AimOur study examines optimal lag time using a GnRH antagonist protocol and GnRH agonist trigger for ICSI. Methods and MaterialsWe analyzed data from 220 women undergoing GnRH antagonist protocol using a GnRH agonist trigger for ICSI at our clinic between 02/2012−03/2018. Patients were divided into 4 groups based on lag time: 34.00–34.99 hours (n = 32), 35.00–35.99 hours (n = 113), 36.00–36.99 hours (n = 57) and 37.00 h or more (n = 18).Analyses were performed with the Kruskal-Wallis test, Chi-Square, and Spearman’s rho correlation. ResultsA positive correlation was found for the number of M2 oocytes aspirated and lag time (ρ = 0.138, p = 0.04) and for the total number of oocytes aspirated and lag time, (ρ = 0.174, p = 0.01). No correlation was found between the proportion of M2 oocytes aspirated and lag time (p = 0.217).The third group (36 h) had significantly more M2 oocytes aspirated than the second group (35 h) (12.4 ± 7.1 vs 9.4 ± 6.2; p = 0.039). The four groups did not differ for the proportion of mature M2 oocytes (H = 2.453, p = 0.484). The four groups differed in the frequency of live births per fresh embryos transferred (χ2 = 9.364, p = 0.025). ConclusionOur study identified a positive correlation between lag time and both the number of M2 oocytes and the total number of oocytes aspirated-factors which lead to an increased rate of successful pregnancies. Further research is necessary.