P4 Levels Research Articles

Is it possible to replace eCG by GnRH in the hormonal protocol for timed artificial insemination in ewes during the breeding season?

Hormonal protocols based on progestogens and equine chorionic gonadotrophin (eCG) are efficient for estrus and ovulation synchronization in ewes. Although eCG is indispensable during seasonal anestrus, it may not be necessary during the breeding season. Thus, we tested the hypothesis that GnRH is effective in replacing eCG during the breeding season allowing satisfactory ovulation rate, luteal function and conception rates after timed artificial insemination (TAI). Ewes (n = 134) with a minimum body condition score of 2.5 (0–5 scale) were treated with intravaginal devices (IVD) containing 60 mg of medroxyprogesterone acetate (MPA) for seven days and received 0.26 mg of sodium cloprostenol at the time of IVD removal. In Exp. 1, at IVD removal, ewes (n = 29) were allocated to three groups: eCG (200 IU at IVD removal; n = 10); eCG+GnRH (200 IU eCG at IVD removal and 4 µg of buserelin 36 h later; n = 10); or GnRH (buserelin 36 h after IVD removal; n = 9). Blood samples were collected 2, 6 and 12 days after TAI moment (54 h after IVD removal), for progesterone (P4) analysis. In Exp 2, the ewes were allocated to eCG (n = 10) or GnRH (n = 10) groups, as above described, and ovulation moment was evaluated 54, 66 and 78 h after IVD removal. In Exp 3, TAI was performed in ewes from eCG (n = 45) and GnRH (n = 40) groups using 100 × 106 motile spermatozoa from a pool of semen collected from four rams. In Exp. 1, based on P4 levels, we confirmed that all the ewes ovulated (29/29) and there was no significant effect of group (P = 0.89) or group x day (P = 0.18) on P4 concentration, being observed a significant effect of day (P = 0.0001). In Exp. 2, the maximum DF diameter (P = 0.26) and ovulation moment (P = 0.69) did not differ between groups. In Exp. 3, pregnancy rate was significantly lower (P = 0.02) in GnRH (22.5 %; 9/40) compared to eCG (46.7 %; 21/45). The results indicate that, although ovulation and luteal function were not altered after eCG, eCG+GnRH or GnRH treatment, GnRH alone before TAI cannot be used to replace eCG treatment during the breeding season.

The natural menstrual cycle revisited – can natural cycle be trusted

BackgroundThe serum progesterone (P4) level during the luteal phase (LP) plays a crucial role in the initiation and maintenance of pregnancy. However, it is unclear whether the natural cycle consistently provides the best endocrine profile and whether mid-luteal serum P4 levels are always sufficient to support implantation and early pregnancy. The question has become more relevant in relation to fertility treatment, as more frozen embryo transfer cycles are performed in the natural cycle. Moreover, can serum hormone levels and covariates measured during the follicular phase (FP), such as Follicle Stimulation Hormone (FSH), Luteinizing Hormone (LH), Estradiol (E2), Anti-Mullerian Hormone (AMH) and Antral Follicle Count (AFC), be used to predict P4 levels during the luteal phase (LP)?ResultsThis observational prospective cohort study analysed 26 healthy women with a cycle length between 21–35 days and a body mass index (BMI) < 30 kg/m2. Blood sampling started on the fifth day of the menstrual cycle and continued every fifth day until the next cycle. The procedure was repeated for a total of three cycles. The study found that only ten women had a P4 level greater than 30 nmol/L on cycle day 20 or 25 in all three cycles. In total, only 45 cycles out of 77 cycles had serum P4 levels ≥ 30 nmol/L.The E2 level ≥ 345 pmol/L on cycle day 10 proved to be predictive of a P4 level of ≥ 30 nmol/L on either day 20 or day 25 with a sensitivity of 57% and a specificity of 89%. No other covariates, including the FSH level cycle day 5, LH levels during the follicular phase, age, weight, AFC and AMH cycle day 5 correlated with LP P4 levels.ConclusionsA significant correlation between FP E2 levels cycle day 5 (> 131pmol/L) and cycle day 10 (> 345pmol/L) and a LP P4 level ≥ 30 nmol/l was found; thus, the FP E2 level is a predictor of corpus luteum competence. Our findings highlight the existence of suboptimal P4 levels during the LP and a significant inter-individual and intra-cycle variation in P4 levels during the LP in regular menstruating women.

Triclosan affects steroidogenesis in mouse primary astrocytes in vitro with engagement of Sirtuin 1 and 3

Triclosan (TCS) is a widely used antimicrobial, antifungal, and antiviral agent. To date, it has been reported that TCS can enter the human body and disrupt hormonal homeostasis. Therefore, the aim of our paper was to evaluate the impact of TCS on astrocytes, i.e. a crucial population of cells responsible for steroid hormone production. Our data showed that, in mouse primary astrocyte cultures, TCS can act as an endocrine disrupting chemical through destabilization of the production or secretion of progesterone (P4), testosterone (T), and estradiol (E2). TCS affects the mRNA expression of enzymes involved in neurosteroidogenesis, such as Cyp17a1, 17β-Hsd, and Cyp19a1. Our data showed that a partial PPARγ agonist (honokiol) prevented changes in Cyp17a1 mRNA expression caused by TCS. Similarly, honokiol inhibited TCS-stimulated P4 release. However, rosiglitazone (classic PPARγ agonist) or GW9662 (PPARγ antagonist) had a much stronger effect. Therefore, we believe that the changes observed in the P4, T, and E2 levels are a result of dysregulation of the activity of the aforementioned enzymes, whose expression can be affected by TCS through a Pparγ-dependent pathway. TCS was found to decrease the aryl hydrocarbon receptor (AhR) and Sirtuin 3 protein levels, which may be the result of the activation of the these proteins. Since our study showed dysregulation of the production or secretion of neurosteroids in astrocytes, it can be concluded that TCS reaching the brain may contribute to the development of neurodegenerative diseases in which an abnormal amount of neurosteroids is observed.

P-429 Measuring serum progesterone levels on the day of a fresh blastocyst transfer is debatable as there is no evidence for correlation with ongoing pregnancy results

Abstract Study question Does serum progesterone (P4) level on the day of a fresh blastocyst transfer relate to ongoing pregnancy rate (OPR)? Summary answer OPR after a fresh single blastocyst transfer seems not to be associated with serum P4 level on the day of transfer. What is known already Several studies have suggested an optimal serum P4 threshold for embryo transfers in frozen embryo transfer cycles. However, for fresh embryo transfers, data are scarce and conflicting. Study design, size, duration A retrospective single-center cohort study was performed on 598 complete data cycles, retained out of 631 consecutive fresh single blastocyst transfer cycles. Data were collected from patients undergoing a fresh IVF/ICSI cycle with autologous oocytes at a tertiary fertility center between June 2022 and August 2023. Participants/materials, setting, methods A total of 482 unique patients underwent one or more single fresh blastocyst transfers using micronized vaginal progesterone (MVP) 800mg BID for luteal phase support starting day after oocyte retrieval. Serum P4 levels were determined on day of transfer. The primary endpoint was OPR, positive hCG-test and early pregnancy loss (EPL) were secondary endpoints. Unadjusted and adjusted generalized estimating equations (GEE) assessed P4 level association with the endpoints, using an exchangeable working correlation matrix. Main results and the role of chance No association was identified between P4 levels on day of transfer and OPR after accounting for maternal age, blastocyst quality, P4 and estradiol level at last ultrasound, BMI and endometrial thickness. Data were grouped according to 10/50/90 and 25/50/75 percentiles, resulting in groups &amp;lt; = or &amp;gt; 27ng/ml and &amp;lt; = or &amp;gt; 40ng/ml, with omission of 90th and 75th percentile groups due to right-skewed distribution. With serum P4 levels higher than 40ng/ml, the probability of a positive hCG-test significantly increased (OR 1.79, CI 1.11-2.91; p = 0.018). No significant difference was observed in patients with serum P4 &amp;lt; = or &amp;gt; 27ng/ml (OR 1.23, CI 0.66-2.30; p = 0.506). Regarding OPR, no significant differences were observed when comparing patients with serum P4 &amp;lt; = or &amp;gt; 27/ng/ml or &amp;lt; = or &amp;gt; 40ng/ml (OR 0.88, CI 0.42-18.85; p = 0.736 and OR 1.41, CI 0.83-2.37; p = 0.203, respectively). Moreover, EPL did not significantly differ between serum P4 level groups (OR 1.93, CI 0.61-6.13; p = 0.265 and OR 1.2, CI 0.51-2.84; p = 0.674). Age (p &amp;lt; 0.001) and embryo quality (p &amp;lt; 0.001) are significantly related with OPR. Additionally, late follicular progesterone levels and OPR are significantly associated (OR 2.39, CI 1.16-4.95 in patients with P4 &amp;lt; = or &amp;gt; 27/ng/ml and OR 2.21, CI 1.07-4.55 in patients with &amp;lt; =40ng/ml and &amp;gt;40ng/ml). Limitations, reasons for caution Limitations include the retrospective design and the lack of live birth outcome data. Progesterone levels exceeding 60 ng/ml were not further analyzed to their precise value but grouped as 60 plus which did not allow to identify potential detrimental effects of very high P4 levels on treatment outcome. Wider implications of the findings Our data suggest that measuring P4 on the day of a single blastocyst transfer with MVP luteal phase support may not be indicative of OPR or the risk of EPL. This raises questions regarding the necessity for routine P4 measurements as well as the development of rescue strategies. Trial registration number ONZ-2023-0628

P-418 What is the optimal midluteal phase circulating progesterone level during a fresh IVF cycle? The answer differs from what one expects in a natural cycle

Abstract Study question Do midluteal phase circulating progesterone (P4) levels relate to the fresh embryo transfer clinical outcome and is rescue P4 useful in such cases? Summary answer Live birth rate (LBR) was significantly lower when P4 levels were below 20ng/ml and did not seem to benefit from rescue intramuscular P4. What is known already Preceding literature on the predictive value of midluteal P4 for LBR has shown conflicting results and frequently included small sample sets and heterogeneous ovarian stimulation protocols. The findings reported ranged from the lack of an association between P4 serum levels and clinical outcome to worse success rates seen either in the groups with the highest or the lowest P4 in midluteal phase. Furthermore, while rescue P4 has been empirically proposed, it has lacked formal assessment thus far. Study design, size, duration In total, 652 patients who underwent a fresh single blastocyst transfer in an IVF cycle in our center between 2014 and 2021 were included in this retrospective cohort study. All were GnRH-antagonist cycles using hCG trigger and luteal support with micronized vaginal progesterone 200mg twice daily. Rescue intramuscular progesterone was systematically administered whenever low midluteal P4 (below 1 standard deviation of the center’s mean) levels were detected. Participants/materials, setting, methods The cycles were subdivided into three groups according to the midluteal P4 levels: &amp;lt;10ng/ml, 10-20ng/ml and &amp;gt;20ng/ml. The primary outcome assessed was LBR, with secondary outcomes including clinical pregnancy (CPR) and pregnancy loss rates. Maternal age and body mass index (BMI), year of treatment, ovarian response, endometrial thickness, cause of infertility and embryo quality were controlled for using multivariable logistic regression. Main results and the role of chance The mean age ± standard deviation of patients was 36.8±3.8 years, while mean weight and BMI were 63.2±11.5kg and 23.3±3.9kg/m2, respectively. LBR in the group with P4&amp;gt;20ng/ml (46.5%) was significantly higher than in the groups with low and intermediate P4 serum levels (23.1% and 28.1%, respectively, p &amp;lt; 0.001). The same was true for CPR (54.5%, 30.8% and 37.9%, respectively, p &amp;lt; 0.001). Conversely, the rate of pregnancy loss in the group with P4&amp;gt;20ng/ml (23.8%) was significantly lower than in the groups with low and intermediate P4 levels (42.6% and 37.5%, respectively, p = 0.018). The multivariable logistic regression confirmed that, after controlling for the confounding variables described above, P4&amp;gt;20ng/ml remained significantly associated with a higher LBR (OR: 2.285; 95% CI: 1.506-3.469) and a lower pregnancy loss rate (OR: 0.488; 95% CI: 0.273-0.873), irrespective of the use of rescue IM P4. Finally, a multivariable linear regression analysis of the patients baseline characteristics suggested that the only parameter predictive of midluteal P4 levels was the female patients’ weight, with an inverse relationship (adjusted regression coefficient: -0,012; 95% CI: -0,016/-0,009). Limitations, reasons for caution As the pituitary suppression, ovulation trigger and luteal support protocols used in the cycles included in the study were homogeneous (in order to maximize the internal validity of the results), this limits the extrapolibility of the results to other stimulation protocols. Furthermore, we caution that this is a retrospective study. Wider implications of the findings The usefulness of both midluteal P4 serum measurements and rescue P4 should continue to be assessed. The reasons behind the need for higher levels of P4 in a fresh cycle, when compared to natural cycles, must also be investigated. Luteal support may require personalization according to female body weight. Trial registration number Not Applicable

P-653 Impact of dydrogesterone use in cycles with low progesterone levels on the day of frozen embryo transfer

Abstract Study question Assessing if Frozen Embryo Transfer (FET) outcomes in hormonal replacement treatment with dydrogesterone (DYD) for low progesterone levels match those with normal progesterone (P4) values Summary answer The live birth rate showed no significant difference between patients with low and normal serum P4 levels in FET, utilizing DYD as rescue therapy What is known already The number of IVF/ICSI cycles and frozen embryo transfers (FET) is increasing. The preparation for FET can occur in natural or artificial cycles, showing no significant differences in clinical pregnancy and live birth rates. Artificial cycles involve exogenous estradiol supplementation, starting after menses, and progesterone (P4) addition post-endometrial proliferation. Numerous studies consistently indicate that diminished P4 levels are correlated with inferior outcomes in hormonal replacement treatment, showing a potential decrease of up to 30% in ongoing pregnancy rates. Despite a suggested rescue strategy, evidence supporting its efficacy using an oral route is insufficient. Study design, size, duration This was a retrospective cohort study, of single embryo transfers (SET) from FET cycles conducted between July 2019 and March 2022, after artificial endometrial preparation with valerate-estradiol and micronized vaginal P4. In cases of low serum P4 (up to 12,5ng/dl) on the planned transfer day, 10 mg of dydrogesterone (DYD) three times daily was added. Solely blastocyst single embryo transfers (SET) were included. The primary endpoint was the live birth rate&amp;gt; 24 weeks. Participants/materials, setting, methods 535 FET-SET cycles were analyzed. Patients with an endometrial layer thinner than 7 mm, endocavitary pathology or serum P4 values&amp;lt; 4ng/dl were excluded. The explanatory variables evaluated were: relevant female characteristics, previous IVF/ICSI cycle and relevant FET cycle parameters (endometrial thickness, serum P4 value on the FET day, days in culture before blastocyst freezing) and use of DYD as rescue therapy. Statistical analysis was performed using IBM®SPSS® statistics v22.0 and R4.2.2 for Windows. Main results and the role of chance Out of the 535 FET cycles analyzed, 136(25.4%) underwent treatment with DYD. There were 337 pregnancies (63%), 207 live births (38.6%) and 130 miscarriages (38.5%). Serum P4 varied from 4.2-32.3 ng/dl, with an average of 14.93 ng/dl (SD 4.52). The best model to explain serum P4 distribution considered a gamma distribution for P4. Female age on the FET day, female ethnicity and female weight seemed to explain P4 variation. A sensitivity analysis on the P4 values was performed. The data were divided into 5 groups according to P4 levels and the rate of live birth per quintile was evaluated. There were no statistically significant differences between the groups. Patients with low P4 which had DYD added had no statistical differences regarding deliveries, 40% vs 36% (p = 0,354). The multivariable model revealed that older females at pick up, a thinner endometrial measure and transferring a day 6 blastocyst were negativey associated with live birth (p &amp;lt; 0,05), but not P4 levels or medication with DYD. A sub-group analyses was performed according to the median value of P4 (9.6 ng/ml) of those to whom DYD was added. No statistically significant differences were identified for biochemical pregnancy 59% vs 60% (p = 1) and live birth rate 34%vs38% (p = 0,653). Limitations, reasons for caution This retrospective study lacks an untreated control group, causing inherent heterogeneity. It’s unclear if there’s an upper limit for P4 levels to restrict DYD supplementation or a minimum level rendering this strategy unsuitable. Monitoring DYD’s fetal/neonatal safety is imperative. Wider implications of the findings The supplementation with oral DYD in hormonal replacement FET cycles is an easy and cheap strategy in patients with for low P4 levels. The duration of DYD administration should be further evaluated. Trial registration number not applicable

P-297 Intensive luteal phase support regimens for endometriosis and adenomyosis patients in hormone replacement therapy frozen embryo transfer cycles – subcutaneous or intramuscular progesterone treatment?

Abstract Study question Does subcutaneous (SC) progesterone treatment provide progesterone levels comparable to intramuscular (IM) progesterone when using a cut-off level of 118nmol/l(37.1ng/ml) during intensive luteal phase support? Summary answer As part of an intensive luteal phase support regimen SC progesterone provides higher P4 levels compared to the use of IM progesterone. What is known already Low serum progesterone (P4) levels in HRT-FET cycles negatively impact reproductive outcomes. This effect is particularly pronounced in patients with endometriosis and adenomyosis, due to reduced progesterone actions and progesterone resistance at the receptor level. Therefore, higher serum P4 levels are required to maintain an optimal reproductive outcome during HRT-FET. A P4 cut-off of 118nmol/l (37.1ng/ml) or above has been suggested, a level four times higher than that of the non-endometriosis patient. To achieve high P4 levels, intensive luteal phase support (LPS) regimen has been suggested. This includes vaginal progesterone administration and additional treatment via IM, SC or rectal administration. Study design, size, duration This cohort study includes 380 HRT-FET cycles in endometriosis/adenomyosis patients. Patients were treated from January 2016 until August 2019 with one type of intensive LPS regimen (n = 262) and from January 2000 to January 2023 with two different LPS regimens depending on whether serum P4 levels were lower or higher than 118nmol/l on the day of blastocyst transfer (BT) (n = 118). P4 was measured in a standardised manner. Participants/materials, setting, methods Patients received treatment in a public university affiliated fertility clinic. Endometrial priming involved the use of estradiol followed by vaginal progesterone. From the fourth day of progesterone, one of the following intensive LPS regimens were added: A) 50mg IM progesterone, B) 25mg SC progesterone BD, or C) 25mg SC progesterone BD plus 400mg rectal rescue progesterone BD if P4&amp;lt;118nmol/l. Blastocyst transfer (BT) and P4 measurement were performed on the sixth day of progesterone administration. Main results and the role of chance The overall pregnancy rate, live birth rate (LBR) and total pregnancy loss rates for the whole cohort were 63%, 41% and 36%, respectively. For the three groups A, B and C the mean serum P4 levels on the day of BT were 103.1 ±44.4nmol/l, 170.5 ±62.9nmol/l and 90.1 ±21.5nmol/l. In Group A (IM) 67% of patients (176/262) had P4 levels &amp;lt;118 nmol/l whereas only 36% (43/118; p &amp;lt; 0.001) had P4 &amp;lt;118 nmol/l in the groups of patients treated with SC progesterone (Group B and C). The unadjusted LBR was not significantly different between groups, 39%, 39%, 51%, (p = 0.32). However, a sub-group analysis showed that comparing patients with serum P4 levels &amp;lt;118nmol/l on the BT day in Group A with patients with P4 levels &amp;lt;118nmol/l in Group C a significantly higher LBR was found if intensive LPS included both SC and rectally administered progesterone compared to IM progesterone, only: 51% (44/86) vs 34% (59/176), p = 0.03. The adjusted odds ratio for live birth was 2.04 (95% CI [1.00, 4.16], p = 0.05) after adjusting for age, vitrification day 5 or 6, blastocyst quality and numbers of blastocysts transferred, if intensive LPS included SC and rectal rescue compared to IM, only in patients with P4 levels &amp;lt;118nmol/l. Limitations, reasons for caution This cohort study was conducted over two different time periods. Single embryo transfer was performed only during the later period. The bio-pharmacological profiles of SC and IM progesterone differ, which can influence serum P4 levels. However, in the present study, all blood samples were collected in a standardised manner. Wider implications of the findings This study is the first to report reproductive outcomes following different intensive LPS regimens in a cohort of endometriosis/adenomyosis patients. To increase reproductive outcomes, individualisation of the HRT-FET cycle may be beneficial and, importantly, rectal progesterone administration can be used as a rescue regimen in addition to SC progesterone. Trial registration number Due to the retrospective study design no ethical approval was needed.

P-626 Progesterone rescue is effective for clinical pregnancy outcome in women presenting low circulating progesterone levels at frozen embryo transfer day

Abstract Study question Is subcutaneous progesterone (prolutex® 25mg daily) addition effective for clinical pregnancy outcome in women presenting low progesterone(P4) levels at frozen embryo transfer(FET) day? Summary answer Subcutaneous progesterone rescue improved ongoing pregnancy rate and clinical pregnancy rate in women with low P4 results at FET day. What is known already FET has been increased, because preimplantation genetic testing for aneuploidies is performed widely, and FET presents a significantly low risk of ovarian hyperstimulation syndrome.In FET, there is no established optimal protocol for preparing endometrium, but hormonal replacement therapy (HRT) is widely used due to its convenient schedule adjustment. HRT protocol induce anovulation, functioning corpus luteum is lacking and exogenous progesterone administration is very important. Therefore, women showing low P4 levels around FET day have lower pregnancy rate. However, there are currently no data on the effectiveness of progesterone rescue. Study design, size, duration Primary outcomes from a randomized cohort retrospective study investigating subcutaneous progesterone (prolutex® 25mg) daily addition for whom had low P4 levels (&amp;lt;12ng/mL) at FET day. The study was conducted at one infertility center in Korea from January 2022 to December 2023 and pregnancy outcomes of FET transfers registered until Oct. 2023,. 194 patients fulfilled the inclusion criteria. Participants/materials, setting, methods One hundred ninety four women divided into two groups based on progesterone levels, with 12ng/ml as the threshold. Under 12ng/mL group were randomized to subgroup with and without addition of subcutaneous progesterone. In total 278 FET cycles, the effect of progesterone rescue on cumulative clinical pregnancy rate (CPR), Ongoing pregnancy rate (OPR) was reported as primary outcomes. Main results and the role of chance In total 278 cycles, there were 137 cycles with adequate serum P4 levels, while 141 cycles presented with low P4 levels in which randomly rescue subcutaneous progesterone administration added from next day of embryo transfer. When comparing the groups, there was no statistical difference in maternal age, BMI, smokers, duration of infertility, type of infertility, indication of in vitro fertilization(IVF), endometrial preparation method. In low P4 group, with and without subcutaneous progesterone addition subgroups included 51 and 90 cycles, retrospectively. CPR were 52.9% versus 35.6% (P = 0.044; chi-squared test) with a statistically significant higher rate of OPR in progesterone rescue group (45.1% versus 28.9%, p = 0.052). When comparing the group with adequate P4 levels and the group with low P4 levels without progesterone rescue, OPR were 18.2% versus 28.9% (p = 0.06, chi-squared test), CPR was significantly higher in low P4 levels group (23.4% versus 35.6%,p=0.046, chi-squared test). Adequate P4 level group had more IVF attempts (4.6 versus 2.2, p &amp;lt; 0.001, chi-squared test). There is tendency to use more luteal phase support medication with repeated attempt (serum P4 concentration on ET day 39.3 ng/mL versus 8.7 ng/mL, p &amp;lt; 0.001, t-test) Limitations, reasons for caution The main limitation of the study was retrospective, and had small sample size. A control group without progesterone rescue in low P4 levels did not receive any intervention. Wider implications of the findings The progesterone rescue for women presenting low P4 on FET day improved clinical outcome and excessively raising progesterone medication may have adverse effects. Therefore rather than using excessive progesterone administration from luteal support start day, progesterone rescue might be effective when low P4 levels presenting at FET day. Trial registration number not applicable

P-437 Are serum progesterone levels predictive for reproductive outcomes in artificial cycles if individualized luteal phase support is only administered when progesterone is below 5 ng/ml?

Abstract Study question Do serum progesterone levels predict reproductive outcomes in artificial cycles when individualized luteal phase support is administered only if progesterone falls below 5 ng/ml? Summary answer The low 5 ng/ml progesterone threshold appears to be sufficient for the patient not to require additional progesterone injections when progesterone is above 5 ng/ml. What is known already Serum progesterone (P4) level is negatively associated with ongoing pregnancy rate (OPR) and miscarriage rate in artificially prepared frozen embryo transfer (FET) cycles using standard micronized vaginal progesterone (MVP) for luteal phase support. Individualized luteal support is considered a successful rescue strategy in cycles with low serum progesterone levels eliminating the effect of P4 levels on outcome. Cut-off levels for this rescue strategy range from 8.8 to 10 ng/ml. Study design, size, duration A retrospective single-centre cohort study including a total of 1,016 vitrified-warmed single blastocyst transfers performed at a tertiary fertility centre between October 2021 and December 2022. Blastocysts of ≥ 3CC Gardner score were eligible for vitrification on day 5 or day 6 (Vit-kit or Vit-kit NX, FuijiFilm). Embryos were warmed and subsequently transferred on the same day which was day 5 in the cycle, regardless if the embryo was vitrified on day 5 or day 6. Participants/materials, setting, methods We included 595 patients undergoing an artificially prepared embryo transfer (ET) cycle using MVP 800mg BID for luteal phase support. Serum P4 levels were determined on the day of ET with the initiation of additional injections of subcutaneous P4 from the day after ET in patients with P4 &amp;lt; 5ng/ml. Primary endpoint was OPR. Multivariable logistic regression was performed to correct for confounders. Generalized estimating equations (GEE) models were fitted for primary and secondary outcomes. Main results and the role of chance OPR was comparable for the 3 groups: 9/40 (22.5%) for patients with P4 &amp;lt;5ng/ml, 64/275 (23.3%) for patients with P4 5-10 ng/ml and 181/701 (25.8%) for patients with P4 &amp;gt;10ng/ml (p = 0.377). No significant association between P4 category and likelihood for OPR was found after adjusting for confounders. Early clinical miscarriage rate was significantly different between the 3 groups: 5/40 (12.5%) in patients with P4 &amp;lt;5ng/ml, 26/275 (9.5%) with P4 5-10ng/ml and 37/701 (5.3%) with P4 &amp;gt;10ng/ml (P = 0.005). Limitations, reasons for caution Higher clinical miscarriage rate in cycles with a P4 level under 10 ng/ml questions the efficacy of rescue progesterone in cycles with a low P4. The patient population also exists of a subgroup of PGT-a tested cycles which was not included as a potential confounder in this analysis. Wider implications of the findings The 5 ng/ml P4 threshold appears to be sufficient for the patient not to require additional P4 injections when P4 is above 5 ng/ml. Consequently, a significantly lower proportion of cycles would need to be supplemented with rescue progesterone; 3.8% versus 31.3% for the 10 ng/mL cut off. Trial registration number ONZ-2023-0223

P-672 Is there a sweet spot for follicular size and/or estradiol level for the initiation of micronized progesterone to optimize natural proliferative phase (NPP) cryo-cycle outcome?

Abstract Study question Is there an optimal follicular size and/or estradiol level for the initiation of micronized progesterone to optimize outcomes in NPP cycles potentially restricting scheduling flexibility? Summary answer All follicular sizes &amp;gt;12 mm and estradiol levels &amp;gt;120 pg./mL do not significantly correlate with ongoing pregnancy rates (OPR) or pregnancy loss (PL). What is known already Planning of a frozen embryo transfer (FET) is feasible within either a natural (NC) or artificial cycle (AC). While AC-FETs offer flexibility in planning, concerns have raised due to studies indicating a higher risk for obstetric complications. Conversely, NC-FETs show a protective effect against these complications but are usually less flexible in planning and ask for more monitoring. Recently, the natural proliferative phase (NPP) protocol was described, in which luteal phase support is initiated as soon as the endometrium reaches 7mm. NPP-FETs were associated with ongoing pregnancy rates comparable to AC- and NC-FETs. Study design, size, duration A retrospective single-centre study including a total of 300 consecutive single frozen embryo transfers (FET), performed at a tertiary fertility centre between March 2023 and December 2023. All cycles were single blastocyst transfer cycles retained from IVF/ICSI cycles with autologous oocytes. Participants/materials, setting, methods A total of 227 patients underwent one or more frozen embryo transfer cycles. Administration of MVP 800mg BID was initiated after fulfilling the following criteria: follicular size &amp;gt; 12mm, estradiol level ≥120ng/mL and endometrium thickness ≥7mm. Blastocyst transfers were scheduled on the fifth day of MVP administration. Serum progesterone (P4) levels were determined on the day of transfer. Associations between treatment outcome, patient and cycle characteristics were explored using generalized estimating equations analyses. Main results and the role of chance The follicular size and estradiol level on the day of MVP initiation was not significantly associated with pregnancy rate (PR), ongoing pregnancy rate (OPR) or pregnancy loss (PL) (p = 0.386 and p = 0.260; p = 0.830 and p = 0.467; p = 0.978 and p = 0.313 respectively). Mean number of visits needed prior to the initiation of the MVP for scheduling the embryo transfer was 2.4 (SD 1.3). PR, but not OPR, was significantly associated with both the duration of the follicular phase as well as the number of visits (p = 0.008 and p = 0.006 respectively). For 91.3 % (274/300) of patients, embryo transfer could be scheduled on a weekday. 49% (146/300) of patients had a positive hCG test after the fresh blastocyst transfer in the NPP cycle. Ongoing pregnancy rate was 35 % (104/300), whereas pregnancy loss (early clinical miscarriage and biochemical pregnancy) was 23% (33/146). The OPR was significantly related to the age of the patient (p = 0.029). Both the PR as the OPR were significantly related the blastocyst quality (p = 0.008 and 0.009 respectively). The incidence of pregnancy loss was only significantly associated with blastocyst quality (p = 0.027). Mean serum P4 level at day of transfer was 19.4 ng/ml (SD 6.1) with a P4 level above 10ng/ml in 97% of cycles. Limitations, reasons for caution The retrospective character is a limitation of this study as well as the absence of live birth rate as a clinical outcome variable. Wider implications of the findings Flexibility about the required follicular sizes and estradiol levels open the perspective for exploring more flexible planning criteria such as lower thresholds for endometrial thickness. Further studies are needed to investigate whether ovulation occurs with formation of functional corpora lutea to effectively protect against hypertensive disorders. Trial registration number ONZ-2024-0015

P-412 Elevated progesterone levels after hormone replacement therapy (HRT) for frozen embryo transfer (FET) does not impair IVF outcomes, a retrospective study

Abstract Study question Does high serum progesterone (P) level on the day of FET after hormone replacement therapy affect IVF outcomes? Summary answer The supraphysiological serum progesterone levels on the day of FET after HRT do not impair reproductive outcomes. What is known already A proper synchronization between embryo and the endometrium is still required by controlling the timing and the dosage of exposure to exogenous hormones. Previous observational studies have highlighted the negative effects of serum hormone levels at the minimum threshold during FET cycles. However, based on existing research, the relationship between progesterone levels on FET day and pregnancy outcomes as well as the certain dose of progesterone ideally to achieve pregnancy and a live birth are debatable topics Study design, size, duration This is an observational, retrospective and cohort study including 7539 FET cycles that were performed under artificial endometrial preparation with HRT using exogenous estradiol and progesterone (vaginal, subcutaneous and intramuscular) between January 2017 and December 2022. Both euploid FET from autologous oocyte ICSI-PGT-A cycles (n = 1822) or FET from ICSI cycles using donated oocytes (n = 5724) were considered. Participants/materials, setting, methods Women were divided into three groups according to P4 level on the day of FET:P&amp;lt;20 ng/ml (n = 6623),P≥20-40 ng/ml (n = 770) and P &amp;gt; 40 ng/ml (n = 146).All ET were performed at the blastocyst stage.The primary outcome was live birth rate (LBR). Secondary outcomes evaluated were biochemical, clinical, and ongoing pregnancy, and miscarriage rate (MR) calculated per FET.Categorical variables were compared between groups with Fisher’s exact test. Logistic regression models adjusted (AOR) with several confounders. Main results and the role of chance There were no significant differences in baseline characteristics of the study population. The mean of P4 level was 15,66 ng/ml. Female age was P &amp;lt; 20:38.7 ±3.1, P ≥ 20-40: 39.0 ±3.0, and P &amp;gt; 40: 38.1 ±2.9 (p = 0.53) years old. The group with the highest progesterone levels always found lower biochemical pregnancy even no statistically significant [64.6% (P &amp;lt; 20) vs 63.7% (P ≥ 20-&amp;lt;40) vs 61.1% (P &amp;gt; 40); p = 0.62]; in clinical pregnancy [56.3% (P &amp;lt; 20) vs 56.8% (P ≥ 20-&amp;lt;40) vs 51.4% (P &amp;gt; 40); p = 0.5] and in ongoing [47.0% (P &amp;lt; 20) vs 47.0% (P ≥ 20-&amp;lt;40) vs 41.8% (P &amp;gt; 40); p = 0.47] pregnancy rates. The MR was also higher in the group with higher progesterone, although not significantly: 16.2% (P &amp;lt; 20) vs 15.0% (P ≥ 20-&amp;lt;40) vs 18.0% (P &amp;gt; 40) (p = 0.60). No statistically significant differences were reported for live birth rate, 41.8% (P &amp;lt; 20) vs 40.3% (P ≥ 20-&amp;lt;40) vs 34.2% (P &amp;gt; 40) (p = 0.2). The multivariate analysis showed an increased risk of miscarriage when P &amp;gt; 40ng/ml, aOR= 1.14 (0.75-1.74) (p = 0.55) whereas no such association was found with P4 ≥20-40 ng/ml, aOR=0.92 (0.74-1.15) (p = 0.46). Additionally, the probability of live birth is similar when the patient was P &amp;gt; 40 (aOR= 0.77 (0.51-1,15) (p = 0.20)) than when P ≥ 20-&amp;lt;40 (aOR=0.94 (0.79-1,11) (p = 0.47) compared with the references group. Limitations, reasons for caution The retrospective design and different female factors for IVF were included in the study leading to a possible biased population although the adjusted analysis. Different routes of progesterone administration were considered in the study. Currently, we don’t report the timing between the last P4 administration and dosage on the FET. Wider implications of the findings This information is useful for clinicians who monitor progesterone levels prior to FET to avoid reaching high levels that would undermine the chances of achieving a newborn. Trial registration number Not Applicable

O-147 Role of serum progesterone levels and subcutaneous progesterone supplementation in endometriosis patients undergoing artificial cycle frozen embryo transfer

Abstract Study question Is there any impact of serum progesterone levels and subcutaneous progesterone supplementation on live birth rate in endometriosis patients undergoing Artificial Cycle-Frozen Embryo Transfer (AC-FET)? Summary answer Endometriosis patients have comparable progesterone levels before transfer to those without endometriosis. Progesterone supplementation results in similar live birth rate (LBR) to the general population. What is known already Progesterone (P4) resistance is a hallmark of uterine alterations in endometriosis. This disease could disrupt balance between progesterone and estrogen signaling pathways, resulting in estrogen dominance and progesterone resistance at the receptor level, which warrants a deeper exploration for adjusting dosages when performing AC-FET. A recent retrospective study highlights the potential significance of a progesterone threshold when using vaginal and intramuscular progesterone systematically in AC-FET, correlating with a significantly higher LBR for endometriosis-patients. However, no current studies have compared LBR according to P4 the day before blastocyst-transfer and progesterone supplementation in endometriosis versus non-endometriosis-patients undergoing AC-FET with standard vaginal treatment. Study design, size, duration This is a retrospective cohort study including 985 cycles undergoing autologous AC-FET with standard vaginal progesterone treatment (600 mg/day) at a university-affiliated fertility center from January 2019 to December 2022. 168 cycles were from patients with confirmed endometriosis. LBR was evaluated in both groups based on P4 levels the day before transfer. Progesterone supplementation via SC injection (25 mg/day) was administered when levels were deemed low (&amp;lt;10.6 ng/mL based on our previously published research). Participants/materials, setting, methods Endometrial preparation involved oral estrogens at 6 mg daily from the begging of menstrual cycle. Micronized vaginal progesterone (600 mg/day) was added when thickness reached 7 mm. P4 was measured before embryo transfer; supplementation (25 mg SC injection) was added if &amp;lt; 10.6 ng/mL. FET occurred 6 days after initiating luteal phase support. Treatment continued until the pregnancy test. If pregnant, treatment ceased at 10 weeks of amenorrhea. Main results and the role of chance Comparisons between endometriosis and non-endometriosis groups revealed similar baseline characteristics regarding age, BMI, parity, number of embryos transferred, and embryo quality. The number of cycles with levels ≥10.6 ng/mL before transfer were comparable between patients with and without endometriosis (aOR 0.98, 95% CI 0.65-1.47) according to a multivariable logistic regression analysis adjusting for age, BMI, and prior progesterone levels &amp;lt;10.6ng/mL the day before transfer. Unadjusted findings demonstrated equivalent clinical pregnancy, miscarriage, and live birth rates in endometriosis compared to non-endometriosis patients. A subsequent multivariable logistic regression analysis to analyze LBR was conducted, considering four groups: Group 1: Endometriosis with progesterone &amp;lt;10.6ng/mL and SC supplementation (51 patients) Group 2: Endometriosis with progesterone ≥10.6ng/mL (117 patients) Group 3: Non-endometriosis with progesterone &amp;lt;10.6ng/mL and SC supplementation (274 patients) Group 4: Non-endometriosis with progesterone ≥10.6ng/mL (543 patients) Adjusted for age, BMI, and embryo quality, Group 1 was considered as the reference. Results showed comparable live birth rates between the reference group and Group 2 (aOR 0.79, 95% CI 0.36-1.74), Group 3 (aOR 0.91, 95% CI 0.45-1.8), and Group 4 (aOR 1.9, 95% CI 0.71-2.74). Subcutaneous supplementation for endometriosis patients with P4&amp;lt;10.6ng/mL the day before embryo transfer led to similar LBR compared to the other groups. Limitations, reasons for caution The main limitation of this study is the single-center retrospective approach. While the total number of endometriosis patients is 168, the representation of endometriosis patients in group 1 may be a study limitation. Furthermore, all endometriosis stages were analyzed together although this may be clinically distinct entities. Wider implications of the findings Endometriosis patients have the same risk to have P4&amp;lt;10.6ng/mL the day before FET as non-endometriosis patients. Vaginal progesterone, with subcutaneous supplementation if necessary, appears user-friendly, leading to a comparable live birth rate to that of patients with normal progesterone levels before transfer, regardless of endometriosis status. Trial registration number not applicable

O-303 What is the ideal dose of daily FSH after corifollitropin-alfa? A multicentre-randomized control trial

Abstract Study question Does the mean serum trigger-progesterone levels show significant variation based on the administered rFSH dose from day 8 onwards during ovarian stimulation using corifollitropin-alfa? Summary answer Lower rFSH doses from day 8 of ovarian stimulation onwards are linked to reduced serum progesterone (P4) levels without impact on cumulus-oocyte complexes. What is known already Progesterone elevation during ovulation trigger prompts a non-elective freeze-all strategy, given the lower live birth rates in fresh embryo transfers. Ongoing research explores methods to mitigate late follicular phase P4 rise to allow a fresh embryo transfer. Granulosa cells’ progesterone production is an FSH-driven and dose-dependent process, and FSH dose is furthermore pivotal for a good ovarian response. Balancing FSH dose is crucial for both optimal ovarian response and preventing undesirable P4 elevation while maintaining an adequate number of retrieved oocytes and allowing a fresh embryo transfer. Study design, size, duration This study is an interim-analysis of a randomized-controlled multi-center clinical-trial comparing three ovarian-stimulation-protocols: (A) corifollitropin alfa (CFA) followed by rFSH 50 IU/day; (B) CFA followed by rFSH 150 IU/day; (C) CFA followed by rFSH 250 IU/day.The primary outcome is the serum-progesterone-level on the day of hCG among the study groups, secondary outcomes included number of oocytes, clinical pregnancy rates and freeze-all cycles. 167 women (of 261) were randomized between July 2019-November 2023, with a 1:1allocation. Participants/materials, setting, methods Young (&amp;lt;40 years old), normo-ovulatory cycle (25-35 days), with AFC&amp;gt;7 and &amp;lt;20, and a BMI&amp;lt;29 Kg/m2 patients were recruited in the fertility clinic of three different tertiary university hospitals. On day 8 of ovarian stimulation (OS) following CFA administration, patients who needed additional rFSH were randomized into 3 groups (A-B-C). The OS was performed in a GnRH-antagonist protocol and the ovulation triggered with hCG. A fresh embryo-transfer was performed on day 5 following oocyte retrieval. Main results and the role of chance Overall, 167 patients were randomized, 60 in group A, 55 in group B and 52 in group C. The mean age and BMI were comparable between the groups. Mean AMH was respectively 2.2, 2.1 and 2.2 ng/mL, for group A, B and C (p = 0.95). Group A resulted in a significantly lower P4 level at trigger as compared with group B and C (respectively, 0.4ng/mL, 0.9 ng/mL and 1.4 ng/mL; p &amp;lt; 0.001). The mean number of retrieved oocytes was 10.7, 11 and 12.1 for group A, B and C, respectively (p = 0.47), while the mean number of mature oocytes was 7.3, 6.9 and 7.8 for group A, B and C, respectively (p = 0.7). A freeze-all approach was decided for 27%, 29% and 46% for group A, B and C respectively (p = 0.057). No significant differences were found for fresh-embryo-transfer outcomes (chemical pregnancy rates: 33%, 31% and 29%, respectively for A, B and C (p = 0.12).Of interest, comparisons between serum FSH at trigger showed a mean of 12.5, 17 and 20.5 IU/L for group A, B and C, respectively (p &amp;lt; 0.001) while the serum estradiol levels on day of trigger were comparable among the groups (1659, 2092 and 2098 ng/L for group A, B and C, respectively (p = 0.091). Limitations, reasons for caution This is an interim analysis planned to detect the mean difference in COC between groups. Therefore, the mean differences of the primary outcome should be interpreted as exploratory findings which warrant future investigation. Wider implications of the findings Administering a lower rFSH dose from day 8 onward during ovarian stimulation results in comparable numbers of COC but also leads to significantly lower serum P4 levels. Consequently, this finding has the potential to reduce the overall consumption of rFSH, thereby possibly increasing the likelihood of a fresh embryo transfer. Trial registration number NCT03686852

P-599 Oestradiol/Progesterone ratio Impact on Frozen Embryo Transfer (FET) and pregnancy outcomes

Abstract Study question Are oestradiol (E2) levels and oestradiol/progesterone (P4) ratio related to higher implantation rates and reduced risk of miscarriage? Summary answer We concluded that higher E2 levels and an increased E2/P4 ratio on the day before FET were associated with lower implantation rates. What is known already A progesterone threshold &amp;lt; 9.2 ng/ml on the day of FET has been related to lower rates of clinical pregnancy. Progesterone is responsible for increasing endometrial vascularization and exerting an immunomodulator effect that allows blastocyst implantation. How oestradiol affects implantation is not well described yet. An appropriate luteal phase support is essential to synchronize the endometrium, ensure proper implantation and reduce the risk of abortion. However, no studies exist regarding E2 levels and E2/P4 ratios related to pregnancy outcomes in the first weeks of pregnancy. Study design, size, duration This is a prospective cohort study. From February 2023 to July 2023, 35 patients who underwent IVF in a private ART medical center were enrolled. Only euploid embryos after PGT- A were transferred and only the first single FET per patient was included. All embryos were transferred in blastocyst stage. Mosaic embryos, fresh cycles, patients with no PGT-A, miscarriages beyond 10 weeks and chemical pregnancies (no evidence of gestational sac) were excluded. Participants/materials, setting, methods 35 patients who underwent IVF-ICSI were retrospectively analyzed. Serum oestradiol, serum progesterone and E2/P4 ratios were measured the day before embryo transfer and later in three moments: 15 days after positive BHCG test, when gestational sac was visualized; 10-15 days after visualization of the gestational sac, when positive embryo heart rate was verified, and in any moment of vaginal bleeding until the 10th week of pregnancy. Main results and the role of chance Our study supported the hypothesis that E2/P4 ratio before FET may be an important factor to predict the risk of threatened abortion considering that all patients had P4 levels &amp;gt; 10 ng/ml before FET. Multiple logistic regression revealed that E2 and E2/p4 ratio on the day before FET are variables related to the risk of threat of abortion. The resulting adjusted OR for E2 on the day before FET in patients who suffered a threatened abortion vs patients who did not experience threat of abortion was 1,025 (95% CI: 1,001-1,049), p value 0,041. The OR for E2/p4 ratio before FET in patients with treat of abortion vs those without threat of abortion was 1,235 (95% CI: 1,035-1,472), p value 0,019. ROC curves showed a significant predictive value for E2 levels &amp;gt;220 pg/ml [AUC of 0,88 (95% CI: 0,73-1,0), p value 0,009] and E2/P4 ratio &amp;gt; 23 [AUC 0,91 (95% CI: 0,78-1,0), p &amp;lt; 0,05], the day before FET indicating a higher risk of threat of miscarriage and loss of pregnancy. Multiple logistic regression did not find significant statistical differences, either E2 or E2/P4 ratio, between the two groups of patients in subsequent determinations at 4 and 6 weeks after transfer. Limitations, reasons for caution The main limitation of this study is the sample size with the lack of further serial measurements of oestradiol and p4 in patients that presented threat of abortion/ miscarriage before the 10th week of pregnancy. Wider implications of the findings E2/P4 ratio the day before FET seems to be the most important factor to decrease threat of abortion in patients with P4&amp;gt; 10 ng/ml. Our study may hold predictive value regarding endometrial receptivity and provide clinicians a guide to individualise FET protocols and to prevent early pregnancy losses Trial registration number N/A

P-285 Serum progesterone level may not impact ongoing pregnancy rate (OPR) in patients with endometriosis undergoing frozen-warmed-blastocyst transfer employing hormone replacement treatment (HRT) with subcutaneous progesterone

Abstract Study question Do serum progesterone (P4) levels on frozen embryo transfer (FET)-1 day impact OPRs in patients with endometriosis undergoing HRT-FET with subcutaneous progesterone (sc P)? Summary answer In HRT cycles employing sc P, serum P4 levels on FET-1 day do not affect OPRs in patients with or without endometriosis. What is known already In HRT-FET cycles employing vaginal P, reproductive outcomes are improved by serum P4 monitoring and employing a rescue protocol, if needed. However, there is paucity of data for the need of serum P4 monitoring in HRT cycles employing sc P. Endometriosis is associated with P4 resistance. There is only one study evaluating the impact of serum P4 levels on reproductive outcomes in patients with endometriosis; in this study lacking a control group and employing vaginal P gel 90 mg/12 hours supplemented by daily 50 mg intramuscular P, a threshold of 37.1 ng/ml has been reported for improved live birth rates. Study design, size, duration Retrospective cohort study. 451 consecutive patients undergoing their first HRT- vitrified warmed blastocyst transfer during November 2021-June 2023 at Anatolia IVF Centre were included. 44 patients had endometriosis with surgical and/or ultrasound findings and the remaining 407 patients did not harbor endometriosis. The presence of adenomyosis was an exclusion criterion. Following 10-12 days of 6 mg/day estradiol valereate use, sc P 25 mg/12hours (Prolutex, Ibsa) was administered. The primary outcome measure was OPR. Participants/materials, setting, methods Serum P4 levels were measured 4-5 hours after morning administration of sc P on FET-1 day. The impact of serum P4 on OPRs was assessed at various percentiles adjusting for female age, body mass index, prior birth, preimplantation genetic testing for aneuploidy, number/grade of blastocysts transferred. Multivariate regression models were performed in the whole cohort incorporating polynomial P4 terms and endometriosis interactions; optimal model for OPR prediction was selected using the lowest Akaike Information Criteria. Main results and the role of chance Patients with (27/44, 61.4%) and without (214/407, 52.6%) endometriosis had comparable OPRs. In the endometriosis group, the 10th, 50th, and 90th percentiles of serum P4 were 15.6, 25.1, and 35.4 ng/ml, respectively, compared to 14.9, 23.5, and 35.6 ng/ml in the non-endometriosis group. In the endometriosis group, the OPRs were 60.0%, 66.7%, 56.3%, and 60.0% for the &amp;lt;10p, 10-50p, 50-90p, &amp;gt;90p groups, respectively. These figures were 46.3%, 50.3%, 56.8%, and 51.2%, in the non-endometriosis group, respectively. In the endometriosis group, taking the 50-90p group as the reference, the adjusted odds ratios (aORs) for OPR of &amp;lt; 10p, 10-50p, and &amp;gt;90p groups were 1.24 [0.03, 50.45], 1.77 [0.31, 10.24], and 2.08 [0.05, 80.23], respectively. In the non-endometriosis group, the respective aORs were 0.85 [0.40, 1.79], 0.80 [0.49, 1.29], and 0.72 [0.34, 1.50]. In the multivariate regression analysis to determine the optimal model, neither serum P4 levels (aOR for linear term: 1.11 [0.98, 1.26]; for quadratic term: 0.998 [0.996, 1.00]) nor the diagnosis of endometriosis (OR: 1.48 [0.74, 2.98]) significantly predicted ongoing pregnancy after adjusting for confounders. Additionally, the optimal model did not include the interaction term, suggesting that endometriosis does not significantly modify the impact of serum P4 levels on OPRs. Limitations, reasons for caution The study’s limitations include retrospective study design, a limited sample size of the endometriosis cohort, absence of laparoscopy for endometriosis exclusion in the non-endometriosis group, and lack of data on endometriosis phenotypes. Wider implications of the findings In HRT-FET cycles employing sc P 25 mg/12hours neither serum P4 nor the presence of endometriosis have an impact on OPRs. Endometriosis may not modify the impact of serum P4 on OPRs suggesting a lack of P resistance in such cycles or the overcoming of P resistance by this regimen. Trial registration number not applicable

Uterine tone: A neglected criterion for the selection of bovine embryo transfer recipients.

This study evaluated the relationship between CL features assessed by ultrasound (luteal tissue area and blood flow, BF) or rectal palpation (size), uterine tone (UT), plasma progesterone (P4) concentration on Day 7 (D7) and subsequent pregnancy outcomes in bovine embryo recipients. A total of 163 cows and heifers were included in this study. The expected day of ovulation after the synchronization protocol was designated as D0. On D7, ovaries and uterus were examined by ultrasonography and rectal palpation, and subjective scores (1-3 scale) were assigned for CL size, area and BF, and for UT. Blood samples were collected for further P4 analysis. Each embryo recipient then received a grade I frozen-thawed invivo-produced blastocyst, which was transferred to the uterine horn ipsilateral to the CL. Pregnancy diagnosis was performed on D35, and the results were retrospectively compared with the assigned scores for CL and UT. We observed a significant (p < .02) interaction between CL size and UT, with a progressive increase in the likelihood of pregnancy for recipients bearing a large CL among those with turgid UT. Ultrasound scoring of the CL using B-mode and Doppler-mode did not significantly predict pregnancy rates on D35 (p < .6 and p < .5, respectively). However, logistic regression analysis revealed a trend towards a quadratic effect (p < .08 and p < .06) indicating that the probability of pregnancy varied according to the area of luteal tissue and P4 concentrations, respectively. No significant (p > .05) association was found between the probability of pregnancy and the BF area of the CL. In summary, UT before embryo transfer may reflect successful recipient synchronization. Elevated P4 levels, assessed by CL size, may offset uterine contractility, mitigating adverse effects. Additionally, the CL area may be more important than its vascularization area when evaluating recipients D7 after ovulation.

NR4A1-mediated regulation of lipid droplets in progesterone synthesis in goat luteal cells†.

Nuclear receptor NR4A1 is a key factor in glycolipid metabolism and steroidogenesis, while lipid droplets serve as crucial dynamic organelles for lipid metabolism in luteal cells. To investigate the effects of NR4A1 on lipid droplet metabolism and progesterone (P4) synthesis in goat corpus luteum in vitro, luteal cells from the middle-cyclic corpus luteum were isolated and treated with Cytosporone B (CSNB, an agonist) or siRNA of NR4A1. Results showed that both low (1μM) and high (50μM) concentrations of CSNB promoted lipid droplet accumulation, while NR4A1 knockdown reduced lipid droplet content. CSNB increased while siNR4A1 decreased total cholesterol content; however, CSNB and siNR4A1 did not change triglyceride content. CSNB increased the expression of perilipins at mRNA and protein levels, also increased LDLR, SCARB1, SREBFs, and HMGCR mRNA abundance. Treatment with siNR4A1 revealed opposite results of CSNB, except for HMCGR and SREBF2. For steroidogenesis, 1μM CSNB increased, but 50μM CSNB inhibited P4 synthesis, NR4A1 knockdown also reduced the P4 level. Further analysis demonstrated that 1μM CSNB increased the protein levels of StAR, HSD3B, and P-HSL, while 50μM CSNB decreased StAR, HSD3B, and CYP11A1 protein levels. Moreover, 50μM CSNB impaired active mitochondria, reduced the BCL2, and increased DRP1, Caspase 3, and cleaved-Caspase 3 protein levels. siNR4A1 consistently downregulated the P-HSL/HSL ratio and the steroidogenic protein levels. In conclusion, NR4A1-mediated lipid droplets are involved in the regulation of progesterone synthesis in goat luteal cells.

The Effect of Prolactin on Gene Expression and the Secretion of Reproductive Hormones in Ewes during the Estrus Cycle.

Prolactin (PRL) plays an important role in animal follicle development and ovulation. However, its regulatory effects on the different stages of the estrus cycle in ewes are unclear. In this study, bromocriptine (BCR, PRL inhibitor) was used to study the effect of PRL on the secretion of reproductive hormones and gene expressions in order to explore its regulatory effects on the sexual cycle of ewes. Eighty healthy ewes with the same parity and similar weights were randomly assigned to the control group (C, n = 40) and the treatment group (T, n = 40, fed bromocriptine). After estrus synchronization, thirty-one ewes with overt signs of estrus were selected from each group. Six blood samples were randomly obtained by jugular venipuncture to measure the concentration of PRL, estrogen (E2), progesterone (P4), luteinizing hormone (LH), follicle-stimulating hormone (FSH), and gonadotropin-releasing hormone (GnRH) in the proestrus, estrus, metestrus, and diestrus. At the same time, we collected the ovaries of the six ewes in vivo after anesthesia in order to detect follicle and corpus luteum (CL) counts and measure the expression of hormone-receptor and apoptosis-related genes. The results show that PRL inhibition had no significant effects on the length of the estrus cycle (p > 0.05). In proestrus, the number of large and small follicles, the levels of E2, FSH, and GnRH, and the expressions of ER, FSHR, and the apoptotic gene Caspase-3 were increased (p < 0.05); and the number of middle follicles and the expression of anti-apoptotic gene Bcl-2 were decreased (p < 0.05) in the T group. In estrus, the number of large follicles, the levels of E2 and GnRH, and the expressions of the StAR, CYP19A1, and Bcl-2 genes were increased (p < 0.05), and the number of middle follicles was decreased (p < 0.05) in the T group. In metestrus, the number of small follicles and the expression of LHR (p < 0.05) and the pro-apoptotic gene Bax were increased (p < 0.05); the number of middle follicles was decreased (p < 0.05) in the T group. In diestrus, the number of large follicles, middle follicles, and CL, the level of P4, and the expressions of PR, 3β-HSD, StAR, Caspase-3, and Bax were increased (p < 0.05); the number of small follicles and the expression of Bcl-2 were decreased (p < 0.05) in the T group. In summary, PRL inhibition can affect the secretion of reproductive hormones, the follicle count, and the gene expression during the estrus cycle. These results provide a basis for understanding the mechanisms underlying the regulation of the ewe estrus cycle by PRL.

Alteration in kisspeptin and reproductive hormones during different superovulation protocols in dromedary camel

This study explored the alteration in kisspeptin and reproductive hormones during different superovulation protocols (SOP) in dromedary camel. The kisspeptin and reproductive hormonal profile, ovarian response, and the quality and quantity of embryos in dromedary camel donors were evaluated. A total of thirty donor camels were divided into two groups: the 5dSOP group, which received diluent containing 400 mg pFSH dissolved in 20 ml and administered two times daily for 5 days at decreasing doses (2.5, 2, 1.5, and 1 ml); and the 3dSOP group, which received diluent containing 400 mg pFSH dissolved in 12 ml and administered two times daily for 3 days at decreasing doses (3 ml, 2 ml, and 1 ml). Ultrasonography was used to monitor the ovarian environment, recording daily follicle count and dimensions and the time taken for follicles to mature. On the sixth day after mating, a corpus luteum (CL) count was conducted. On the 8th day after mating, records of the quantity and quality of embryos collected were kept. Blood samples from the jugular vein were collected at the commencement of the superovulation protocol and at 8:00 a.m. for the following 48 h to measure the concentrations of follicle-stimulating hormone (FSH), luteinizing hormone (LH), estradiol (E2), kisspeptin (KP), and progesterone (P4). The findings indicated that the 3dSOP yielded superior results compared to the 5dSOP in terms of follicle quantity and size, as well as the quantity of CL and embryos. This improvement was attributed to significantly higher concentrations of reproductive hormones, including FSH, LH, E2, kisspeptin, and P4 (P ≤ 0.05), in the 3dSOP than in the 5dSOP. In conclusion, reducing the duration of superovulation protocols contributed to the proliferation of follicles with improved dimensions and counts, ultimately resulting in a greater quantity of embryos of superior quality. The levels of FSH, LH, E2, KP, and P4 were affected significantly by SOP and time of evaluation.

Peculiarities of the endocrine status of women with precancerous lesions of the vulva

The purpose of the study was to determine the hormonal profile of peripheral blood in women with precancerous lesions of the vulva.Material and methods. 309 women with precancerous lesions of the vulva and 60 gynecologically healthy women aged 25 to 70 years were included in the study. Patients with precancerous lesions were divided into 4 groups depending on the nosological unit: 87 women with severe vulvar dysplasia dependent on HPV (VHSIL), 154 individuals with differentiated vulvar dysplasia independent of HPV (dVIN), 36 patients with extramammary Paget's disease of the vulva (VPD) grade Ia and 32 women with melanoma in situ of the skin vulva In the groups with VHSIL, with dVIN, with melanoma in situ, there were women of premenopausal and postmenopausal age, therefore, for the comparison of hormonal data in these groups, subgroups were selected according to age: subgroups with persons younger than 50 years, subgroups of women aged 50 years and older. 60 conditionally healthy women of the control group also included 30 people under 50 years and 30 people 50 years and older. The levels of estradiol (E2), progesterone (P4), free testosterone (Tf), follicle-stimulating hormone (FSH), luteinizing hormone (LH), prolactin (PRL), anti-Müllerian hormone (AMH), insulin, thyroid-stimulating hormone (TSH), free triiodothyronine (T3f), free thyroxine (T4f), antibodies to thyroperoxidase (ATPO)in blood serumand HOMA indexwere determined. The results. There was no difference between serum LH and FSH levels in women with various precancerous diseases of the vulva under the age of 50 years. Individuals with dVIN were characterized by the highest level of PRL (24.44±1.55 ng/ml). Among women aged 50 years and older, the highest levels of LH, FSH and PRL were observed in persons with melanoma in situ - respectively 36.90±2.09 mIU/ml, 74.67±4.06 mIU/ml, 9.85±2.03 ng/ml; the level of FSH and PRL was higher, and the LH/FSH ratio was lower than in controls in all precancerous diseases of the vulva. No statistically significant difference was found between the basal levels of serum P4 and AMH in subgroups of women younger than 50 years and 50 years and older. Among both premenopausal and postmenopausal women, the highest average levels of E2 were recorded in women with melanoma in situ (104.61±10.22 pg/ml and 30.71±1.90 pg/ml), Tf and Tf index - in patients with dVIN (3.20±0.10 pg/ml and 1.58±0.05 pg/ml vs. 8.25±0.24% and 3.77±0.10%, respectively). Features of the thyroid status were the highest indicators of TSH and ATP in persons with dVIN both in premenopause (2.68±0.30 mIU/ml and 196.39±76.48 IU/ml) and in postmenopause (2.23±0 .09 mIU/ml and 123.46±16.07 IU/ml). The average level of 25(OH)D did not differ statistically significantly between women with various precancerous diseases of the vulva. The level of insulin and NOMA index among premenopausal women was the highest in women with melanoma in situ (14.33±1.55 mIU/ml), in postmenopausal women with VPD (14.90±1.02 mIU/ml), HOMA index – respectively, with dVIN (2.87±0.30) and with VPD (3.60±0.27). Insulin resistance was most common in premenopause among individuals with dVIN (53.33%), and in postmenopause - with VPD (72.22%).Conclusions. Features of the hormonal profile of people with precancerous diseases of the vulva under the age of 50 are the highest levels of PRL, TSH and ATPO in women with dVIN, the highest average levels of Tf and Tf index, insulin level and HOMA index – in patients with VPD, the highest average levels of E2– in women with melanoma in situ. Among postmenopausal patients, the highest levels of TSH and ATPO in women with dVIN, the highest average levels of Tf and Tf index, the level of insulin and NOMA index in patients with VPD, the highest average levels of LH, FSH, PRL and E2 in women with melanoma are characteristic features situ. The average level of 25(OH)D does not differ statistically significantly between women with various precancerous diseases of the vulva. Insulin resistance is most common in premenopausal individuals with dVIN, and in postmenopausal individuals with VPD.