The effect of frozen embryo transfer regimen on the association between serum progesterone and live birth: a multicentre prospective cohort study (ProFET).

STUDY QUESTION: What is the association between serum progesterone levels on the day of frozen embryo transfer (FET) and the probability of live birth in women undergoing different FET regimens? SUMMARY ANSWER: Overall, serum progesterone levels <7.8 ng/ml were associated with reduced odds of live birth, although the association between serum progesterone levels and the probability of live birth appeared to vary according to the route of progesterone administration. WHAT IS KNOWN ALREADY: Progesterone is essential for pregnancy success. A recent systematic review showed that in FET cycles using vaginal progesterone for endometrial preparation, lower serum progesterone levels (<10 ng/ml) were associated with a reduction in live birth rates and higher chance of miscarriage. However, there was uncertainty about the association between serum progesterone levels and treatment outcomes in natural cycle FET (NC-FET) and HRT-FET using non-vaginal routes of progesterone administration. STUDY DESIGN SIZE DURATION: This was a multicentre (n = 8) prospective cohort study conducted in the UK between January 2020 and February 2021. PARTICIPANTS/MATERIALS SETTING METHODS: We included women having NC-FET or HRT-FET treatment with progesterone administration by any available route. Women underwent venepuncture on the day of embryo transfer. Participants and clinical personnel were blinded to the serum progesterone levels. We conducted unadjusted and multivariable logistic regression analyses to investigate the association between serum progesterone levels on the day of FET and treatment outcomes according to the type of cycle and route of exogenous progesterone administration. Our primary outcome was the live birth rate per participant. MAIN RESULTS AND THE ROLE OF CHANCE: We studied a total of 402 women. The mean (SD) serum progesterone level was 14.9 (7.5) ng/ml. Overall, the mean adjusted probability of live birth increased non-linearly from 37.6% (95% CI 26.3-48.9%) to 45.5% (95% CI 32.1-58.9%) as serum progesterone rose between the 10th (7.8 ng/ml) and 90th (24.0 ng/ml) centiles. In comparison to participants whose serum progesterone level was ≥7.8 ng/ml, those with lower progesterone (<7.8 ng/ml, 10th centile) experienced fewer live births (28.2% versus 40.0%, adjusted odds ratio [aOR] 0.41, 95% CI 0.18-0.91, P = 0.028), lower odds of clinical pregnancy (30.8% versus 45.1%, aOR 0.36, 95% CI 0.16-0.79, P = 0.011) and a trend towards increased odds of miscarriage (42.1% versus 28.7%, aOR 2.58, 95% CI 0.88-7.62, P = 0.086). In women receiving vaginal progesterone, the mean adjusted probability of live birth increased as serum progesterone levels rose, whereas women having exclusively subcutaneous progesterone experienced a reduction in the mean probability of live birth as progesterone levels rose beyond 16.3 ng/ml. The combination of vaginal and subcutaneous routes appeared to exert little impact upon the mean probability of live birth in relation to serum progesterone levels. LIMITATIONS REASONS FOR CAUTION: The final sample size was smaller than originally planned, although our study was adequately powered to confidently identify a difference in live birth between optimal and inadequate progesterone levels. Furthermore, our cohort did not include women receiving oral or rectal progestogens. WIDER IMPLICATIONS OF THE FINDINGS: Our results corroborate existing evidence suggesting that lower serum progesterone levels hinder FET success. However, the relationship between serum progesterone and the probability of live birth appears to be non-linear in women receiving exclusively subcutaneous progesterone, suggesting that in this subgroup of women, high serum progesterone may also be detrimental to treatment success. STUDY FUNDING/COMPETING INTERESTS: This work was supported by CARE Fertility and a doctoral research fellowship (awarded to P.M.) by the Tommy's Charity and the University of Birmingham. M.J.P. is supported by the NIHR Birmingham Biomedical Research Centre. S.F. is a minor shareholder of CARE Fertility but has no financial or other interest with progesterone testing or manufacturing companies. P.L. reports personal fees from Pharmasure, outside the submitted work. G.P. reports personal fees from Besins Healthcare, outside the submitted work. M.W. reports personal fees from Ferring Pharmaceuticals, outside the submitted work. The remaining authors have no conflict of interest to declare. TRIAL REGISTRATION NUMBER: ClinicalTrials.gov: NCT04170517.

Serum luteal phase progesterone in women undergoing frozen embryo transfer in assisted conception: a systematic review and meta-analysis.

OBJECTIVE: To investigate the association between luteal serum progesterone levels and frozen embryo transfer (FET) outcomes. DESIGN: Systematic review and meta-analysis. SETTING: Not applicable. PATIENT(S): Women undergoing FET. INTERVENTION(S): We conducted electronic searches of MEDLINE, PubMed, CINAHL, EMBASE, the Cochrane Database of Systematic Reviews, Cochrane Central Register of Controlled Trials, Web of Science, ClinicalTrials.gov, and grey literature (not widely available) from inception to March 2021 to identify cohort studies in which the serum luteal progesterone level was measured around the time of FET. MAIN OUTCOME MEASURE(S): Ongoing pregnancy or live birth rate, clinical pregnancy rate, and miscarriage rate. RESULT(S): Among the studies analyzing serum progesterone level thresholds <10 ng/mL, a higher serum progesterone level was associated with increased rates of ongoing pregnancy or live birth (relative risk [RR] 1.47, 95% confidence interval [CI] 1.28 to 1.70), higher chance of clinical pregnancy (RR 1.31, 95% CI 1.16 to 1.49), and lower risk of miscarriage (RR 0.62, 95% CI 0.50 to 0.77) in cycles using exclusively vaginal progesterone and blastocyst embryos. There was uncertainty about whether progesterone thresholds ≥10 ng/mL were associated with FET outcomes in sensitivity analyses including all studies, owing to high interstudy heterogeneity and wide CIs. CONCLUSION(S): Our findings indicate that there may be a minimum clinically important luteal serum concentration of progesterone required to ensure an optimal endocrine milieu during embryo implantation and early pregnancy after FET treatment. Future clinical trials are required to assess whether administering higher-dose luteal phase support improves outcomes in women with a low serum progesterone level at the time of FET. PROSPERO NUMBER: CRD42019157071.

Serum FSH Levels in Coasting Programmes on the hCG Day and Their Clinical Outcomes in IVF ± ICSI Cycles.

Introduction. Coasting is the most commonly used strategy in prevention of severe OHSS. Serum FSH levels measurements during coasting may aid in optimizing the duration of coasting. Objective(s). To study live birth rates (LBRs), clinical pregnancy rates (CPRs), and optimal duration of coasting based on serum FSH levels on the hCG day. Materials and Methods. It is a retrospective study performed between 2005 and 2008 at Barts and The London Centre for Reproductive Medicine, NHS Trust, London, UK, on 349-coasted women undergoing controlled ovarian stimulation (COS) for IVF ± ICSI. The serum FSH level measurements on the hCG day during coasting programme were analysed to predict the LBR and CPR. Result(s). LBR and CPR were significantly higher when the FSH levels on the hCG day were >2.5 IU/L (LBR: 32.5%, P = 0.045 and CPR: 36.9%, P = 0.027) compared to FSH <2.5 IU/L. The optimal FSH cut-off level for LBR and CPR is 5.6 IU/L on the hCG day. The optimal cutoff for coasting is 4 days. Conclusion(s). Coasting may be continued as long as either serum FSH level is > 2.5 IU/L on the hCG day without compromising the LBR and CPR or to maximum of 4 days.

Can the fall in serum FSH during coasting in IVF/ICSI predict clinical outcomes?

This retrospective cohort study determined whether the total falls in serum FSH and oestradiol concentrations from start to end of coasting in IVF/intracytoplasmic sperm injection could predict clinical outcomes. Ninety-nine cycles, with gonadotrophin-releasing hormone-agonist down-regulation where coasting with serial serum oestradiol and FSH monitoring was adopted due to risk of severe ovarian hyperstimulation syndrome, were consecutively included. The primary clinical outcome was live-birth rate (LBR); other outcomes measured were number of oocytes retrieved and fertilization, implantation and clinical pregnancy rates. LBR for FSH fall>10 IU/l compared with 5-10 and<5 IU/l were 45.4% versus 22.0% and 25.0%, respectively. Mean serum FSH fall was similar with and without live birth (8.4 ± 6.2 versus 7.3 ± 5.0 IU/l) as were mean oestradiol and FSH concentrations on HCG administration, oestradiol fall, percentage fall in FSH/oestradiol and duration of coasting. None of the variables efficiently predicted live birth on regression analysis. The AUC of FSH fall was 0.53 at 11.0 IU/l. Basal FSH, starting and total gonadotrophin dose and duration of coasting were positively correlated with FSH fall. A potentially clinically important association between live birth and FSH fall during coasting was apparent, which requires further evaluation. The purpose of this retrospective cohort study was to determine whether the magnitude of fall in the serum FSH and oestradiol concentrations from start to end of coasting in IVF/intracytoplasmic sperm injection cycles could predict the clinical outcomes. Gonadotrophin-releasing hormone-agonist down-regulated cycles (n=99), where coasting with serial serum oestradiol and FSH monitoring was adopted due to risk of ovarian hyperstimulation, were consecutively included. Live birth was the primary clinical outcome measured; number of oocytes retrieved and fertilization, implantation and clinical pregnancy rates were the other outcomes examined. Live-birth rate tended to be high when FSH fall was >10 IU/l, compared with 5-10 IU/l and <5 IU/l, although not statistically significantly. Mean serum FSH fall were similar in live-birth and no-live-birth cycles (8.4 ± 6.2 versus 7.3 ± 5.0) as were mean oestradiol and FSH concentrations on hCG administration, oestradiol fall, percentage fall in FSH and oestradiol and duration of coasting. None of the variables efficiently predicted live birth. The area under the curve of FSH fall was 0.53. FSH fall of <11.0 IU/l was found to be more likely to predict negative outcome (specificity 84.72%) than predicting positive outcome when FSH fall was >11 IU/l (sensitivity 34.48%). Women's basal FSH, starting and total gonadotrophin dose of ovarian stimulation and duration of coasting had direct positive correlation with the magnitude of FSH fall. A potentially clinically important rise in live birth in association with greater FSH fall during coasting was apparent, which requires further evaluation.

Controlled ovarian hyperstimulation for low responders in in vitro fertilization/intracytoplasmic sperm injection: a low-dose flare protocol.

In this retrospective study of 652 anticipated low response women, the overall clinical outcomes (live birth rate and clinical pregnancy rate [PR]) of low-dose flare (LDF) protocol appeared lower than those of conventional down-regulation (DR) (LDF: 15.1% vs. DR: 20.6% and LDF: 10.3% vs. DR: 17.4%, respectively). The findings that LDF protocol improved the clinical outcome in older women, or when LDF followed an unsuccessful IVF/intracytoplasmic sperm injection (ICSI) cycle with DR (LDF: 19.4% vs. DR: 9.76% and LDF: 13.9% vs. DR: 4.2% respectively), need further evaluation through randomized trials.

Estradiol, progesterone, testosterone profiles in human follicular fluid and cultured granulosa cells from luteinized pre-ovulatory follicles.

BACKGROUND: The production of sex steroids by follicular cells is proposed to be influenced by the maturity of the incumbent oocyte. Thus steroid levels may reflect suitability of an oocyte for IVF. We examined follicular fluids and granulosa cell production of steroid from IVF patients in order to test the relationship between steroid levels and fertilization. METHODS: Follicular fluid and granulosa cells were extracted from 206 follicles of 35 women undergoing controlled ovarian stimulation. Follicular fluid was assayed for estradiol, progesterone and testosterone. Granulosa cells were cultured from individual follicles and their culture media assayed for production of these hormones after 24 hrs in vitro. Levels of steroids were correlated with follicular diameter, oocyte recovery and subsequent fertilization. RESULTS: Follicular fluid levels of progesterone were 6100 times higher than that of estradiol, and 16,900 times higher that of testosterone. Despite the size of follicle triggered after controlled luteinization, the levels of progesterone and testosterone were maintained at relatively constant levels (median 98.1 micromoles/L for progesterone, and 5.8 nanomoles/L for testosterone). However, estradiol levels were slightly lower in the larger follicles (follicular diameter 10-15 mm, median 25.3 nanomoles/L; follicles > = 15 mm, median 15.1 nanomoles/L; linear correlation r = -0.47, p < 0.0001). With respect to oocyte recovery, no steroid showed a significant association in follicular fluid levels. Similarly no difference in follicular fluid steroid levels was found for those oocytes that did or did not fertilize. Significant quantities of progesterone were produced by the granulosa cells but production was constant regardless of the size of follicle from which the cells originated. Estradiol levels were only detectable in 10 of 121 cultures examined, and testosterone in none. Interestingly, when an oocyte was present follicular estradiol levels correlated with progesterone levels. However, when absent, follicular estradiol levels correlated with testosterone levels but not with progesterone. CONCLUSIONS: The principle steroid product of luteinized pre-ovulatory granulosa is progesterone, a differentiation triggered by the gonadotropin surge. However, absolute steroid levels are associated with follicular size, not oocyte maturation/ability to fertilize.

High follicular fluid adenosine levels may be pivotal in the metabolism and recycling of adenosine nucleotides in the human follicle.

This study investigated the biochemical relationship between human follicular/oocyte maturity and the levels of follicular fluid purines. Intrafollicular levels of purine metabolites and creatinine are associated with oocyte presence, and the presence of such high levels of adenosine indicates a privileged site with no adenosine deaminase activity. Subgrouping according to oocyte recovery and fertilization revealed differences in correlation between the purine metabolites: Only where an oocyte was recovered and subsequently fertilized did follicular fluid adenosine, adenine, and hypoxanthine levels correlate with each other. Significantly, purines' correlation with levels of the terminal degradation product, uric acid, could only be seen in failed fertilization samples. Given the established metabolic pathways for adenosine triphosphate/adenosine diphosphate/adenosine monophosphate degradation, the results indicate maximization of 2 purine salvage pathways (from adenine and hypoxanthine) that pivot on the presence of high adenosine levels. Such optimized recovery may be necessary to build a store of salvaged adenosine phosphate for oocyte survival.

The correlation between basal serum follicle-stimulating hormone levels before embryo cryopreservation and the clinical outcome of frozen embryo transfers.

OBJECTIVE: To evaluate the correlation between basal serum FSH level before the fresh IVF/intracytoplasmic sperm injection (ICSI) cycle and the clinical outcome of the subsequent frozen embryo replacement cycles. DESIGN: Retrospective observational study. SETTING: University tertiary referral center, London, United Kingdom. PATIENT(S): Five hundred four consecutive frozen embryo transfer (FET) cycles where serum FSH levels were obtained, on days 1-4 of the cycle before the fresh IVF +/- ICSI cycles. INTERVENTION(S): Frozen-thawed embryo transfer. MAIN OUTCOME MEASURE(S): Clinical pregnancy (CP) and live birth (LB). RESULT(S): Basal serum FSH in 127 women (25.2%) who had a CP was significantly lower compared with that in women who did not have a CP. Multivariate regression analysis showed significant correlation between basal serum FSH levels and clinical pregnancy and a low significance to LB, but there was no statistical significant differences between women who had a CP and those who did not with regard to age, treatment protocol (natural or hormone treatment cycle), or the freeze-thaw interval. The LB rate was higher in natural cycles (n = 71; 21.2%) than in hormone treatment cycles (n = 28; 16.7%). Conceiving in the fresh cycle had a positive influence on the FET outcome. CONCLUSION(S): Basal serum FSH level before fresh IVF/ICSI cycle is inversely correlated to a CP outcome in FET cycles. A trend was present between FSH levels and LB, but this failed to reach statistical significance.

Human granulosa-lutein cell in vitro production of progesterone, inhibin A, inhibin B, and activin A are dependent on follicular size and not the presence of the oocyte.

OBJECTIVE: To investigate inhibin A, inhibin B, activin A, and P production by cultured granulosa cells (GCs) and what relationship this hormone production has to fertility. DESIGN: Luteinized GCs from individual follicles were cultured, and inhibin A, inhibin B, activin A, and P production were measured by ELISA at 24 and 72 hours. SETTING: Research laboratory and university hospital. PATIENT(S): Fifteen women who undertook an IVF-ICSI program, yielding 58 follicles. INTERVENTION(S): Individual follicular aspiration and preparation of GCs for culture. MAIN OUTCOME MEASURE(S): Inhibin A, inhibin B, activin A, and P production; oocyte retrieval; and fertility outcome. RESULT(S): Inhibin A, inhibin B, and P continued to be secreted by GCs in vitro, and activin A levels were detected only marginally in 56% of cultures. The rate of production also was dependent on the size of follicle from which the GCs originated but not on oocyte presence or ability to fertilize. Granulosa cell stimulation with hCG had no effect on inhibin A but increased P and decreased inhibin B production. CONCLUSION(S): A marked effect of luteal differentiation appears to be the inhibition of inhibin B production in response to hCG stimulation. Luteinized GC function, with respect to inhibins, activin A, and P production, was not influenced by the presence or absence of an oocyte and did not correlate with fertility outcome. However, follicle size did influence rates of local hormone production.

Does measuring early basal serum follicular luteinising [correction of lutinising] hormone assist in predicting in vitro fertilization (IVF)/intracytoplasmic sperm injection (ICSI) outcome?

BACKGROUND: The aim was to examine the correlation of early follicular serum lutinising hormone (LH) and the clinical outcome of assisted reproduction technique (ART). METHODS: An observational study included 1333 consecutive women undergoing in vitro fertilization (IVF) and intracytoplasmic sperm injection (ICSI). 964 women were having their first cycle of ART. Data were entered prospectively. All women had serum LH measured in the 6 months before the index cycle studied. No repeat cycles were included. The main outcomes measured were clinical pregnancy (CP) and live birth (LB) correlation to serum LH. Forward multivariate stepwise regression analysis was applied, and other statistical tests were used as appropriate. RESULTS: There was non significant correlation between basal serum LH and CP and LB in the polycystic ovary syndrome group (R2 = 0.02, F = 1.7 and P = 0.76) (R2 = 0.01, F = 2.6 and P = 0.77) respectively after adjusting for age, BMI, day of oocyte retrieval, starting dose, total dose of stimulation, type of gonadotrophin used, number of oocytes retrieved, fertilization rate and number of embryos transferred. Other aetiological causes group there was similarly non significant correlation between basal serum LH and CP (R2 = 0.05, F = 13.1 and P = 0.66), nor for LB (R2 = 0.007, F = 4.5 and P = 0.9). CONCLUSION: Early follicular serum LH measurements in the 6 months before IVF/ICSI treatment cycle did not correlate with the clinical pregnancy or the live birth rate.

Follicular fluid levels of inhibin A, inhibin B, and activin A levels reflect changes in follicle size but are not independent markers of the oocyte's ability to fertilize.

OBJECTIVE: To investigate the biochemical relationship between follicular/oocyte maturity and follicular inhibins and activin levels. DESIGN: Prospective study. SETTING: Research laboratory in university hospital. PATIENT(S): Thirty-five women undertook IVF/ICSI program. INTERVENTION(S): Individual follicular fluid aspirations, oocyte isolation, follicular fluid storage. MAIN OUTCOME MEASURE(S): Inhibin A, inhibin B, and activin A concentrations, oocyte retrieval, and fertility outcome. RESULT(S): Inhibin A, inhibin B, and activin A concentrations varied from 7.9 to 436 ng/mL, 9.7 to 786 ng/mL, and 1.7 to 267.9 ng/mL, respectively. There was no change of inhibin A concentrations, whereas inhibin B and activin A concentrations dropped dramatically as the follicles enlarged. Total follicular content of inhibin A and activin A increased, and inhibin B remained constant. Both inhibin A and inhibin B levels were significantly higher in those follicles from which an oocyte could be recovered, but they did not differ with respect to subsequent oocyte fertilization. CONCLUSION(S): Inhibin A is actively produced throughout follicular growth to retain a set concentration. In contrast, inhibin B appears not to be actively produced, and the concentration drops as follicles enlarge. Activin A concentrations also decrease, but there is some extra synthesis. Higher levels of inhibin A and B are associated with oocyte presence but not with fertilization rates.