Reproductive outcomes of dual trigger therapy with GnRH agonist and hCG versus hCG trigger in women with diminished ovarian reserve: a retrospective study.

BACKGROUND: Diminished ovarian reserve (DOR) is one of the obstacles affecting the reproductive outcomes of patients receiving assisted reproductive therapy. The purpose of this study was to investigate whether dual trigger, including gonadotropin-releasing hormone agonist (GnRHa) and human chorionic gonadotropin (hCG), can improve pregnancy outcomes in patients with DOR undergoing in vitro fertilization (IVF) cycles using mild stimulation protocols. METHODS: A total of 734 patients with DOR were included in this retrospective study. Patients were divided into a recombinant hCG trigger group and a dual trigger group (hCG combined with GnRHa) according to the different trigger drugs used. The main outcome measures included the number of oocytes retrieved, the fertilization rate, the number of transferable embryos, the implantation rate, the clinical pregnancy rate, the miscarriage rate, the live birth rate (LBR), and the cumulative live birth rate (CLBR). Generalized linear model and logistic regression analyses were performed for confounding factors. RESULTS: There were 337 cycles with a single hCG trigger and 397 cycles with dual trigger. The dual trigger group demonstrated significantly higher numbers of retrieved oocytes [3.60 vs. 2.39, adjusted ß = 0.538 (0.221-0.855)], fertilized oocytes [2.55 vs. 1.94, adjusted ß = 0.277 (0.031-0.523)] and transferable embryos [1.22 vs. 0.95, adjusted ß = 0.162 (-0.005-0.329)] than did the hCG trigger group, whereas no significant difference in the fertilization rate was observed between the two groups. Moreover, the embryo transfer cancellation rate (35.5% vs. 43.9%) was obviously lower in the dual trigger group. Among the fresh embryo transfer cycles, the implantation rate, clinical pregnancy rate, miscarriage rate and live birth rate were similar between the two groups. After controlling for potential confounding variables, the trigger method was identified as an independent factor affecting the number of oocytes retrieved but had no significant impact on the CLBR. CONCLUSIONS: Dual triggering of final oocyte maturation with hCG combined with GnRHa can significantly increase the number of oocytes retrieved in patients with DOR but has no improvement effect on the implantation rate, clinical pregnancy rate or LBR of fresh cycles or on the CLBR.

Effect of LH level on HCG trigger day on clinical outcomes in patients with diminished ovarian reserve undergoing GnRH-antagonist protocol.

RESEARCH QUESTION: Does luteinizing hormone (LH) levels on human chorionic gonadotropin (HCG) trigger day (LHHCG) affect the clinical outcomes of patients with diminished ovarian reserve (DOR) undergoing gonadotropin-releasing hormone antagonist (GnRH-ant) protocol? METHODS: Retrospective analysis fresh embryo transfer cycles of DOR patients who underwent GnRH-ant protocol from August 2019 to June 2023. The participants were divided into different groups according to LHHCG level and age. The clinical data and outcomes were compared between groups. RESULTS: In patients with DOR, the HCG positive rate (59.3% versus 39.8%, P = 0.005), embryo implantation rate (34.5% versus 19.7%, P = 0.002), clinical pregnancy rate (49.2% versus 28.4%, P = 0.003), live birth rate (41.5% versus 22.7%, P = 0.005) in LHHCG < 2.58 IU/L group were significantly higher than LHHCG ≥ 2.58 IU/L group. There was no significant correlation between LHHCG level and clinical pregnancy in POSEIDON group 3. In POSEIDON group 4, the HCG positive rate (52.8% versus 27.0%, P = 0.015), embryo implantation rate (29.2% versus 13.3%, P = 0.023), clinical pregnancy rate (45.3% versus 18.9%, P = 0.010) in LHHCG < 3.14 IU/L group were significantly higher than LHHCG ≥ 3.14 IU/L group. Logistic regression analysis indicated that LHHCG level was an independent influencing factor for clinical pregnancy in POSEIDON group 4 patients (OR = 3.831, 95% CI: 1.379-10.643, P < 0.05). CONCLUSIONS: LHHCG level is an independent factor affecting pregnancy outcome of fresh embryo transfer in DOR patients undergoing GnRH-ant protocol, especially for advanced-aged women. LHHCG had a high predictive value for POSEIDON group 4 patients, and LHHCG ≥ 3.14 IU/L predicts poor pregnancy outcomes.

Dietary acid load and risk of diminished ovarian reserve: a case-control study.

BACKGROUND: The epidemiologic evidence on the association between acid load potential of diet and the risk of diminished ovarian reserve (DOR) is scarce. We aim to explore the possible relationship between dietary acid load (DAL), markers of ovarian reserve and DOR risk in a case-control study. METHODS: 370 women (120 women with DOR and 250 women with normal ovarian reserve as controls), matched by age and BMI, were recruited. Dietary intake was obtained using a validated 80-item semi-quantitative food frequency questionnaire (FFQ). The DAL scores including the potential renal acid load (PRAL) and net endogenous acid production (NEAP) were calculated based on nutrients intake. NEAP and PRAL scores were categorized by quartiles based on the distribution of controls. Antral follicle count (AFC), serum antimullerian hormone (AMH) and anthropometric indices were measured. Logistic regression models were used to estimate multivariable odds ratio (OR) of DOR across quartiles of NEAP and PRAL scores. RESULTS: Following increase in PRAL and NEAP scores, serum AMH significantly decreased in women with DOR. Also, AFC count had a significant decrease following increase in PRAL score (P = 0.045). After adjustment for multiple confounding variables, participants in the top quartile of PRAL had increased OR for DOR (OR: 1.26; 95%CI: 1.08-1.42, P = 0.254). CONCLUSION: Diets with high acid-forming potential may negatively affect ovarian reserve in women with DOR. Also, high DAL may increase the risk of DOR. The association between DAL and markers of ovarian reserve should be explored in prospective studies and clinical trials.

Stratification of the risk of ovarian dysfunction by studying the complexity of intermediate and premutation alleles of the FMR1 gene.

FMR1 premutation female carriers are at risk of developing premature/primary ovarian insufficiency (POI) with an incomplete penetrance. In this study, we determined the CGG repeat size among 1095 women with diminished ovarian reserve (DOR) / POI and characterized the CGG/AGG substructure in 44 women carrying an abnormal FMR1 repeat expansion number, compared to a group of 25 pregnant women carrying an abnormal FMR1 CGG repeat size. Allelic complexity scores of the FMR1 gene were calculated and compared between the two groups. In the DOR/POI cohort, 2.1% of women presented with an intermediate repeat size and 1.9% with a premutation. Our results suggest that the risk of POI is highest in the mid-range of CGG repeats. We observed that the allelic score is significantly higher in POI women compared to the pregnant women group (p-value = 0.02). We suggest that a high allelic score due to more than 2 AGG interspersions in the context of an intermediate number of repetitions could favor POI. Larger studies are still needed to evaluate the relevance of this new tool for the determination of the individual risk of developing POI in women with abnormal number of CGG repeats.

Effectiveness of naturopathy for pregnancy in women with diminished ovarian reserve: feasibility randomized controlled trial.

RESEARCH QUESTION: Is conducting a randomized control trial (RCT) to assess the effectiveness of whole-system naturopathy in improving pregnancy rates among women with diminished ovarian reserve (DOR) feasible? DESIGN: A two-arm, parallel group, assessor-blinded feasibility RCT was conducted. Women with DOR, trying to conceive naturally or by ART, were randomly assigned to naturopathy plus usual care, or usual care alone for 16 weeks. Primary outcomes were feasibility (recruitment, adherence, retention rates), acceptability and safety. Secondary outcomes included ongoing pregnancy rates, live birth rates and health-related outcomes (mental health, quality of life, diet, exercise, sleep and weight). Statistical significance of the differences between the two groups (P-values) were exploratory. RESULTS: One hundred and fifteen women completed the screening survey between March and November 2022. Of these, 66 women were assessed for eligibility and 41 (62%) consented. Recruitment resulted in seven enrolments each month. All 41 participants (100%) adhered to the intervention, 38 (93%) completed end-point questionnaires, 32 (78%) found study participation to be acceptable and 18 out of 21 (86%) from the intervention group would recommend a naturopathic intervention to other women with DOR. The naturopathic treatment was associated with only mild and temporary adverse events. No between-group differences were observed for pregnancy and live birth rates. CONCLUSION: The evaluation of whole-system naturopathy through a RCT was feasible and the treatment was acceptable and well tolerated according to women with DOR. Outcomes from this study will help inform sample size calculations powered for fertility outcomes for future RCTs on this topic.

Metabolic signature of follicular fluid in infertility-related diseases: a narrative review.

Metabolomics offers new methods to identify biomarkers for oocyte and embryo quality, and for a better understanding of the physiopathology of infertility. This review investigated the latest findings regarding metabolome-derived biomarkers in follicular fluid of women with the most common types of infertility, and the potential impact on reproductive medicine outcomes. PubMed was searched for publications on metabolomics and human follicular fluid, and key biomarkers, kinetics and relationships with infertility diseases were identified. A reduced concentration of glucose and increased concentrations of lactate and pyruvate were found in follicular fluid of patients with endometriosis and diminished ovarian reserve, and the opposite was found in patients with polycystic ovary syndrome. These signatures may lead to the hypothesis of changed metabolite concentrations in patients with endometriosis and diminished ovarian reserve, and a metabolic pathway alteration with decreased aerobic glycolysis in patients with polycystic ovary syndrome. However, the pattern found in patients with endometriosis and low responders may also be expected in follicular fluid of fertile women. Larger studies are needed to confirm the results. An international database may help to highlight follicular fluid biomarkers in order to improve the selection of cryopreserved oocytes, and to enrich culture medium to restore normal metabolism and improve reproductive treatment outcomes.

Do women with severely diminished ovarian reserve undergoing modified natural-cycle in-vitro fertilization benefit from earlier trigger at smaller follicle size?

OBJECTIVE: To evaluate whether trigger and oocyte collection at a smaller follicle size decreases the risk of premature ovulation while maintaining the reproductive potential of oocytes in women with a severely diminished ovarian reserve undergoing modified natural-cycle in-vitro fertilization. METHODS: This was a retrospective cohort study including women who had at least one unsuccessful cycle (due to no response) of conventional ovarian stimulation with a high dosage of gonadotropins and subsequently underwent a modified natural cycle with a solitary growing follicle (i.e. only one follicle > 10 mm at the time of trigger). The association between follicle size at trigger and various cycle outcomes was tested using regression analyses. RESULTS: A total of 160 ovarian stimulation cycles from 110 patients were included in the analysis. Oocyte pick-up (OPU) was performed in 153 cycles and 7 cycles were canceled due to premature ovulation. Patients who received their trigger at smaller follicle sizes (≤ 15 mm) had significantly lower rates of premature ovulation and thus higher rates of OPU (98.9% vs 90.8%; odds ratio, 9.56 (95% CI, 1.58-182.9); P = 0.039) compared with those who received their trigger at larger follicle sizes (> 15 mm). On multivariable analysis, smaller follicle sizes at trigger (> 10 to 13 mm, > 13 to 15 mm, > 15 mm to 17 mm) were not associated significantly with a lower rate of cumulus-oocyte complex (COC) retrieval, metaphase-II (MII) oocytes or blastulation when compared to the > 17-mm group. On sensitivity analysis including only the first cycle of each couple, the maturity rate among those with COC retrieval was highest in follicle sizes > 15 to 17 mm (92.3%) and > 13 to 15 mm (91.7%), followed by > 10 to 13 mm (85.7%) and lowest in the > 17-mm group (58.8%). During the study period, five euploid blastocysts developed from 48 fertilized MII oocytes with follicle sizes of 12 mm (n = 3), 14 mm (n = 1) and 16 mm (n = 1) at trigger. Of those, four were transferred and resulted in two live births, both of which developed from follicles with a size at trigger of 12 mm. CONCLUSIONS: The ideal follicle size for triggering oocyte maturation may be smaller in women with a severely diminished ovarian reserve managed on a modified natural cycle when compared to conventional cut-offs. The risk of OPU cancellation was significantly higher in women triggered at follicle size > 15 mm and the yield of mature oocytes was not adversely affected in women triggered at follicle size > 13 to 15 mm compared with > 15 to 17 mm. Waiting for follicles to reach sizes > 17mm may be detrimental to achieving optimal outcome. © 2024 International Society of Ultrasound in Obstetrics and Gynecology.

Chitosan alleviates ovarian aging by enhancing macrophage phagocyte-mediated tissue homeostasis.

BACKGROUND: Age-related changes in the ovarian microenvironment are linked to impaired fertility in women. Macrophages play important roles in ovarian tissue homeostasis and immune surveillance. However, the impact of aging on ovarian macrophage function and ovarian homeostasis remains poorly understood. METHODS: Senescence-associated beta-galactosidase staining, immunohistochemistry, and TUNEL staining were used to assess senescence and apoptosis, respectively. Flow cytometry was employed to evaluate mitochondrial membrane potential (MMP) and apoptosis in granulosa cells lines (KGN), and macrophages phagocytosis. After a 2-month treatment with low molecular weight Chitosan (LMWC), ovarian tissues from mice were collected for comprehensive analysis. RESULTS: Compared with the liver and uterus, the ovary displayed accelerated aging in an age-dependent manner, which was accompanied by elevated levels of inflammatory factors and apoptotic cells, and impaired macrophage phagocytic activity. The aged KGN cells exhibited elevated reactive oxygen species (ROS) and apoptotic levels alongside decreased MMP. H2O2-induced aging macrophages showed reduced phagocytosis function. Moreover, there were excessive aging macrophages with impaired phagocytosis in the follicular fluid of patients with diminished ovarian reserve (DOR). Notably, LMWC administration alleviated ovarian aging by enhancing macrophage phagocytosis and promoting tissue homeostasis. CONCLUSIONS: Aging ovarian is characterized by an accumulation of aging and apoptotic granulosa cells, an inflammatory response and macrophage phagocytosis dysfunction. In turn, impaired phagocytosis of macrophage contributes to insufficient clearance of aging and apoptotic granulosa cells and the increased risk of DOR. Additionally, LMWC emerges as a potential therapeutic strategy for age-related ovarian dysfunction.

Effect of diminished ovarian reserve on the outcome of fresh embryo transfer in IVF/ICSI cycles among young women: A retrospective cohort study.

OBJECTIVE: This study aims to investigate the effect of diminished ovarian reserve (DOR) on the clinical outcomes and maternal and infant safety of in vitro fertilization/intracytoplasmic sperm injection (IVF/ICSI) procedures in young women aged ≤ 35 years. METHODS: A retrospective cohort study was performed to analyze the clinical data of 4,203 infertile women aged ≤ 35 years who underwent fresh embryo transfer (ET) in IVF/ICSI cycles. The data were collected from their initial visits to Fujian Maternity and Child Health Hospital between January 2015 and January 2022. Based on their ovarian reserve, the participants were categorized into two groups: DOR group (n = 1,027) and non-DOR group (n = 3,176). A propensity score matching (PSM) method was employed to ensure a relatively balanced distribution of covariates. The primary outcome assessed in this study was the live birth rate, while the secondary observation indicators included rates of high-quality embryo development, blastocyst formation, clinical pregnancy, and miscarriage, along with perinatal complications, neonatal birth weight, and the incidence of low birth weight (LBW). RESULTS: The DOR group showed notably lowered rates of blastocyst formation (59.8% vs. 64.1%), embryo implantation (29.8% vs.33.3%), clinical pregnancy (47.9% vs. 53.6%), and live birth (40.6% vs. 45.7%) compared to the non-DOR group (all P < 0.05). However, no statistically significant differences were observed in the high-quality embryo rate, miscarriage rate, perinatal complications, neonatal birth weight, or LBW incidence in infants between both groups (all P > 0.05). CONCLUSION: DOR has been found to reduce both clinical pregnancy and live birth rates in young females undergoing fresh ET in IVF/ICSI cycles. However, this reduction does not increase the risk of perinatal complications or LBW of infants through live birth cycles.

Live birth rates in in vitro fertilization cycles with five or fewer follicles.

PURPOSE: To evaluate live birth rates (LBRs) for in vitro fertilization (IVF) cycles with ≤5 follicles at trigger, with the goal of helping patients with low follicle counts decide whether to proceed to retrieval. METHODS: This is a retrospective cohort study from an urban, university-affiliated fertility center. All IVF cycles that yielded <10 oocytes between 2016 and 2020 were reviewed. Cycles were included if <5 follicles measuring >14 mm were verified on trigger day. The primary outcome was LBR per retrieval after fresh or frozen transfer. Secondary outcomes were number of oocytes, mature oocytes, 2-pronuclear zygotes (2-PNs), blastocysts for transfer/biopsy, and euploid blastocysts (if preimplantation genetic testing for aneuploidy (PGT-A) was used). RESULTS: 1502 cycles (900 with PGT-A) from 972 patients were included. Mean number of oocytes, mature oocytes, 2-PNs, blastocysts for transfer/biopsy, and euploid blastocysts differed by follicle number (p < 0.001). Across all age groups, there were differences in LBR associated with follicle number (p < 0.001). However, within age groups, not all results were significant. For example, for patients <35 years, LBR did not differ by follicle number and among patients 35-37 years; LBR with two or three follicles was lower than with five (p < 0.02). LBR for patients 35-40 years was <20% with 1-3 follicles and 25-40% with 4-5 follicles. LBR for patients >41 years was <5% with 1-3 follicles and <15% with 4-5 follicles. CONCLUSION: As expected, LBR is higher with more follicles. Providing patients with <5 follicles with specific data can help them weigh the emotional, physical, and financial costs of retrieval.

A phenomenological exploration of the mental health experiences of young women with diminished ovarian reserve.

Infertility is a reproductive disease affecting one in six individuals that renders an individual unable to conceive. One cause of infertility is diminished ovarian reserve (DOR), which reduces the quantity and/or quality of a female's oocyte pool. Although typically indicating normal ovarian aging during the late 30s and early 40s, DOR can also impact younger women, increasing their risk for psychological distress from an unexpected diagnosis of infertility. A phenomenological approach examined the mental health experiences and perceptions of infertility-related mental health care of young women with DOR. Women diagnosed with DOR by age 35 in the United States who experienced emotional distress during infertility were recruited from infertility-specific social media and via snowball sampling. Participants completed a demographic survey and semi-structured individual interview that was audio-recorded, transcribed verbatim, and analyzed using a phenomenological approach. Ten women ages 27-41 completed the study. On average, participants were 30 years of age at the time of DOR diagnosis (age range 25-35), primarily Caucasian (90%), and married (90%). Two main themes were found: (1) Young women with DOR feel like a "forgotten community" coping with an invisible disease; and (2) Not all fertility clinics are created equal. Participants perceived their diagnosis as devastating and hopeless and urged others to find a provider with ample experience treating patients with DOR. This study helped to understand how young women with DOR perceive their mental health and identified a significant need for advancing towards more holistic infertility healthcare that encompasses both physical and mental health.

Health needs, treatment decisions and experience of traditional complementary and integrative medicine use by women with diminished ovarian reserve: A cross-sectional survey.

BACKGROUND: Women with diminished ovarian reserve (DOR) have fewer eggs than would be expected at their age. It is estimated that 10% of women seeking fertility treatment are diagnosed with DOR. However, the success rate of medically assisted reproduction (MAR) is significantly lower in women with DOR, thus many seek additional approaches. AIM: To explore the health needs of women with DOR, treatment options and experience of treatment including traditional complementary integrative medicine (TCIM). METHODS: Anyone with a diagnosis of DOR, living in Australia or New Zealand, aged over 18 were invited to complete an online survey distributed via fertility support networks and social media platforms from April to December 2021. RESULTS: Data from 67 respondents were included. The main aspects of health that were impacted by DOR were fertility (91.0%) and mental health (52.2%). The main treatment recommended was MAR with most women either currently using MAR (38.8%) or having previously used MAR (37.3%). TCIM was widely used with 88.1% of women utilising supplements, 74.6% consulting with TCIM practitioners, and 65.7% adopting self-care practices. The main reasons for using TCIM were to improve fertility or support pregnancy, and to support general health and well-being. CONCLUSIONS: Women with DOR have additional health needs apart from infertility, most notably mental health support. The main form of treatment utilised is MAR, despite DOR being challenging for fertility clinicians. TCIM was widely used, and respondents perceived benefits related to improving fertility, supporting pregnancy, or improving well-being through use of acupuncture, meditation, naturopathy, massage, yoga.

The Protective Role of Magnoliae Flos in Preventing Ovotoxicity and Managing Ovarian Function: An In Vitro and In Vivo Study.

Magnoliae Flos (MF) is a medicinal herb widely employed in traditional medicine for relieving sinusitis, allergic rhinitis, headaches, and toothaches. Here, we investigated the potential preventive effects of MF extract (MFE) against 4-vinylcyclohexene diepoxide (VCD)-induced ovotoxicity in ovarian cells and a mouse model of premature ovarian insufficiency (POI). The cytoprotective effects of MFE were assessed using CHO-K1 or COV434 cells. In vivo, B6C3F1 female mice were intraperitoneally injected with VCD for two weeks to induce POI, while MFE was orally administered for four weeks, beginning one week before VCD administration. VCD led to a significant decline in the viabilities of CHO-K1 and COV434 cells and triggered excessive reactive oxygen species (ROS) production and apoptosis specifically in CHO-K1 cells. However, pretreatment with MFE effectively prevented VCD-induced cell death and ROS generation, while also activating the Akt signaling pathway. In vivo, MFE increased relative ovary weights, follicle numbers, and serum estradiol and anti-Müllerian hormone levels versus controls under conditions of ovary failure. Collectively, our results demonstrate that MFE has a preventive effect on VCD-induced ovotoxicity through Akt activation. These results suggest that MFE may have the potential to prevent and manage conditions such as POI and diminished ovarian reserve.

Diagnostic Value of Autoantibodies against Steroidogenic Enzymes and Hormones in Infertile Women with Premature Ovarian Insufficiency.

The objective of the study was to evaluate the profile and diagnostic significance of serum autoantibodies in infertile patients with premature ovarian insufficiency (POI). The pilot study included 26 patients of reproductive age with POI and diminished ovarian reserve who received complex treatment using new surgical technologies (Group 1) and 18 patients without POI (Group 2). The profile of serum autoantibodies, including anti-ovarian antibodies, antibodies against thyroid peroxidase (TPO), steroidogenic enzymes, and steroid and gonadotropic hormones, was studied using modified ELISAs and human recombinant steroidogenic enzymes (CYP11A1, CYP19A1, CYP21A2). Patients in Group 1 had higher levels of IgG autoantibodies against steroidogenic enzymes, estradiol, progesterone, and TPO than those in Group 2. Tests for IgG antibodies against CYP11A1, CYP19A1, and CYP21A2 exhibited high sensitivity (65.4-76.9%), specificity (83.3-89.9%), and AUC values (0.842-0.910) for POI, the highest in the first test. Three-antibodies panel screening showed higher diagnostic accuracy (84.1% versus 75-79.6%). The levels of these antibodies correlated with menstrual irregularities and a decrease in the antral follicle count. Thus, antibodies against CYP11A1, CYP19A1, and CYP21A2 have a high diagnostic value for POI. Three-antibody panel screening may improve the accuracy of POI diagnosis and be useful for identifying high-risk groups, early stages of the disease, and predicting POI progression.

Benefit from luteal phase progestin primed ovarian stimulation with clomiphene citrate supplementation in young women with diminished ovarian reserve: a retrospective study. / å¹´è½»åµå·¢å‚¨å¤‡åŠŸèƒ½å‡é€€æ‚£è€ åµæ³¡æœŸä¸Žé»„ä½“æœŸåº”ç”¨é«˜å­•æ¿€ç´ çŠ¶æ€è¶ ä¿ƒæŽ’åµæ–¹æ¡ˆæ•ˆæžœæ¯”è¾ƒ.

OBJECTIVES: To compare the pregnancy outcomes of luteal phase and follicular phase progestin-primed ovarian stimulation protocol with clomiphene citrate supplementation (LPPOS+CC and FPPOS+CC) in young women with diminished ovarian reserve (DOR). METHODS: A total of 483 women aged ≤35 years with DOR, who underwent in vitro fertilization (IVF)/intracytoplasmic sperm injection (ICSI)/embryo transfer (ET) with controlled ovarian stimulation using LPPOS+CC (n=257) or FPPOS+CC (n=226) protocols during June 2018 and December 2021 at the First Affiliated Hospital of Wenzhou Medical University, were included in this retrospective study. The baseline characteristics, superovulation results, laboratory related indicators between the two groups, and the pregnancy outcomes of women who achieved at least one high-quality cleavage-stage embryo or good-morphology blastocyst were compared between the two groups. RESULTS: No statistically significant differences were identified between the groups with respect to age, duration of infertility, proportion of secondary infertility, previous failed cycles, body mass index, anti-Müllerian hormone, antral follicle count, basal luteinizing hormone level, basal progesterone level, number of oocytes retrieved, oocyte maturation rate, high-quality cleavage-stage embryo cycle rate, the percentage of women with profound pituitary suppression, live birth rate and preterm birth rate (all P>0.05). The LH levels on the day of trigger [4.0 (2.7, 5.3) vs. 5.1 (3.2, 7.2) IU/L], the percentage of women with LH levels of >10 IU/L on the trigger day (3.13% vs. 10.67%), and the two pronucleus (2PN) rate of ICSI oocytes (72.16% vs. 79.56%) were significantly lower in the LPPOS+CC group than those in the FPPOS+CC group (P<0.05 or P<0.01). The duration of stimulation [11 (9, 12) vs. 9 (8, 11) d], the consumption of total gonadotropin [2213 (1650, 2700) vs. 2000 (1575, 2325) IU], the progesterone levels on the day of trigger [1.3 (0.8, 2.9) vs. 0.9 (0.6, 1.2) ng/mL], the clinical pregnancy rate [61.88% vs. 46.84%], and implantation rate [42.20% vs. 31.07%] in the LPPOS+CC group were significantly higher than those in the FPPOS+CC group (all P<0.01). CONCLUSIONS: Compared to FPPOS+CC, the LPPOS+CC protocol appears to have better pregnancy outcomes for young women with DOR undergoing IVF/ICSI-ET.

Increased serine synthesis in cumulus cells of young infertile women with diminished ovarian reserve.

STUDY QUESTION: What are the differences in gene expression of cumulus cells (CCs) between young women with diminished ovarian reserve (DOR) and those of similar age with normal ovarian reserve (NOR)? SUMMARY ANSWER: Gene expression and metabolome profiling analysis demonstrate that the de novo serine synthesis pathway (SSP) is increased in the CCs of young women with DOR. WHAT IS KNOWN ALREADY: The incidence of DOR has risen, tending to present at younger ages. Its mechanisms and aetiologies are still poorly understood. Abnormal metabolism is present in luteinized CCs of patients with DOR. Previous studies have revealed that mitochondrial dysfunction and impaired oxidative phosphorylation in CCs are related to DOR in women of advanced age. The pathogenic mechanisms likely differ between young women with DOR and cases associated with advanced maternal age. Several studies have examined amino acid metabolism in the follicle, with a focus on embryo development, but less information is available about CCs. The physiological significance of de novo serine synthesis in follicles and oocytes remains largely unknown. STUDY DESIGN, SIZE, DURATION: CC samples were obtained from 107 young infertile women (age <38 years) undergoing ICSI, from July 2017 to June 2019, including 54 patients with DOR and 53 patients with NOR. PARTICIPANTS/MATERIALS, SETTING, METHODS: Oocyte development data were analysed retrospectively. Comprehensive genome-wide transcriptomics of CCs was performed. Differentially expressed genes (DEGs) were identified. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses were performed to categorize the functions of the DEGs and identify significantly enriched pathways. The transcript and protein levels of key enzymes involved in serine synthesis were verified in additional samples using quantitative real-time PCR (qRT-PCR) (n = 10) and capillary western blotting (n = 36). Targeted metabolomics of amino acids in CC extracts was performed by ultrahigh-performance liquid MS (UHPLC-MS/MS). MAIN RESULTS AND THE ROLE OF CHANCE: The number of oocytes (2.4 ± 2.2 versus 12.1 ± 5.3) and metaphase II oocytes (2.1 ± 2.0 versus 9.9 ± 4.9) retrieved was significantly decreased in the DOR versus the NOR group, respectively (P < 0.0001). The rates of fertilization (80.7% versus 78.8%), viable embryos (73.7% versus 72.5%), and high-quality embryos (42.8% versus 49.0%) did not differ between the DOR and NOR groups, respectively (P > 0.05). A total of 95 DEGs were found by transcriptome sequencing. GO and KEGG analyses demonstrated that the DEGs were linked to amino acid metabolism and suggested significantly higher activity of the de novo SSP in the CCs of young women with DOR. Further qRT-PCR and capillary western blotting revealed that key enzymes (PHGDH, PSAT1, PSPH, and SHMT2) involved in de novo serine synthesis were upregulated, and UHPLC-MS/MS analysis showed increases in serine and glycine (a downstream product of serine) levels in the CCs of young patients with DOR. Our data clearly demonstrate that the de novo SSP, which diverts 3-phosphoglycerate from glycolysis to serine synthesis, was upregulated in young DOR CCs. LARGE SCALE DATA: N/A. LIMITATIONS, REASONS FOR CAUTION: Regarding the reproductive capacity of young patients DOR, the pregnancy outcomes were not analysed. The sample size was limited, and only women undergoing ICSI were examined since this was a prerequisite for the acquisition of CCs, which may cause selection bias. The exact mechanisms by which the SSP in CCs regulates ovarian reserve still require further study. WIDER IMPLICATIONS OF THE FINDINGS: Our research presents new evidence that alterations of the SSP in CCs of young infertile women are associated with DOR. We believe this is a significant contribution to the field, which should be key for understanding the cause and mechanisms of ovarian hypofunction in young women. STUDY FUNDING/COMPETING INTEREST(S): This work was supported by grants from the Ministry of Science and Technology of China (2018YFC1005001) and National Natural Science Foundation of China (31601197). There were no competing interests. TRIAL REGISTRATION NUMBER: N/A.

The effect of growth hormone on the metabolome of follicular fluid in patients with diminished ovarian reserve.

BACKGROUND: Increasing evidence supports that the co-treatment with growth hormone (GH) enhances ovarian response and oocyte quality during controlled ovarian stimulation (COS) in patients with diminished ovarian reserve (DOR). The composition of follicular fluid (FF) plays an essential role in oocyte development and mirrors the communication occurring between the oocyte and follicular microenvironment. However, the effect of GH on the FF metabolome remains unclear. METHODS: This prospective observational study recruited DOR patients undergoing in vitro fertilization (IVF) cycles with minimal stimulation protocol for COS. Each patient receiving GH co-treatment was matched to a patient without GH co-treatment by propensity score matching. The FF was collected after isolating oocytes and assayed by gas chromatograph-mass spectrometry (GC-MS) metabolomics. The Pearson correlation was performed to evaluate the relationship between the number of oocytes retrieved and the levels of differential metabolites. The KEGG database was used to map differential metabolites onto various metabolic pathways. RESULTS: One hundred thirty-four FF metabolites were identified by GC-MS metabolomics. Twenty-four metabolites, including glutathione, itaconic acid and S-adenosylmethionin (SAM) showed significant differences between the GH and control groups (p-value < 0.05 and q-value < 0.1). In addition, the number of oocytes retrieved was significantly higher in the GH group compared to the control group (3 vs 2, p = 0.04) and correlated with the levels of five differential metabolites. Among them, the levels of antioxidant metabolite itaconic acid were upregulated by GH administration, while SAM levels were downregulated. CONCLUSIONS: The co-treatment with GH during COS may improve oocyte development by altering FF metabolite profiles in DOR patients. However, given the downregulation of SAM, a regulator of genomic imprinting, the potential risk of imprinting disturbances should not be neglected.

Identification of compound heterozygous variants in MSH4 as a novel genetic cause of diminished ovarian reserve.

BACKGROUND: Diminished ovarian reserve (DOR) is a common cause of female infertility, with genetic factors being a significant contributor. However, due to high genetic heterogeneity, the etiology of DOR in many cases remains unknown. In this study, we analyzed the phenotype of a young woman with primary infertility and performed molecular genetic analysis to identify the genetic cause of her condition, thus providing important insights for genetic counseling and reproductive guidance. METHODS: We collected the patient's basic information, clinical data, as well as diagnostic and therapeutic history and performed whole-exome sequencing on her peripheral blood. Candidate pathogenic variants were validated by Sanger sequencing in family members, and the pathogenicity of variants was analyzed using ACMG guidelines. We used bioinformatics tools to predict variant effects on splicing and protein function, and performed in vitro experiments including minigene assay and expression analysis to evaluate their functional effects on HEK293T. RESULTS: We identified biallelic MSH4 variants, c.2374 A > G (p.Thr792Ala) and c.2222_2225delAAGA (p.Lys741Argfs*2) in the DOR patient. According to ACMG guidelines, the former was classified as likely pathogenic, while the latter was classified as pathogenic. The patient presented with poor oocyte quantity and quality, resulting in unsuccessful in vitro fertilization cycles. Bioinformatics and in vitro functional analysis showed that the c.2374 A > G variant altered the local conformation of the MutS_V domain without decreasing MSH4 protein expression, while the c.2222_2225delAAGA variant led to a reduction in MSH4 protein expression without impacting splicing. CONCLUSIONS: In this study, we present evidence of biallelic variants in MSH4 as a potential cause of DOR. Our findings indicate a correlation between MSH4 variants and reduced oocyte quality, as well as abnormal morphology of the first polar body, thereby expanding the phenotypic spectrum associated with MSH4 variants. Furthermore, Our study emphasizes the importance of utilizing whole-exome sequencing and functional analysis in diagnosing genetic causes, as well as providing effective genetic counseling and reproductive guidance for DOR patients.

Effect of PFOA exposure on diminished ovarian reserve and its metabolism.

Owing to its difficulty in degrading and ease of accumulation in the body, perfluorooctanoic acid (PFOA) has a detrimental effect on reproduction. This study aimed to examine the effect of PFOA concentration in follicular fluid during ovulation stimulation on embryo quality and the impact of PFOA exposure on the metabolic components of follicular fluid. This was a single-center prospective study that included 25 patients with diminished ovarian reserve (DOR), 25 with normal ovarian reserve (NOR), and 25 with polycystic ovary syndrome (PCOS). Follicular fluid samples were analyzed using ultra-high performance liquid chromatography-tandem mass spectrometry. We demonstrated that the PFOA levels of follicular fluid in the DOR group were higher than those in the NOR group and PCOS group (P < 0.05). PFOA concentration in the PCOS group was negatively correlated with high-quality embryos (P < 0.05). To gain more insight into the impact of PFOA on the metabolic composition of follicular fluid, we classified the DOR group based on the PFOA concentration, for which metabolomic analysis was performed. In the high-concentration PFOA group, there was an increase and a decrease in three and nine metabolites, respectively, compared to that in the low-concentration group. These results suggest that PFOA may alter the metabolic composition of follicular fluid, thus, affecting ovarian reserve function.

Granulocyte colony-stimulating factor priming improves embryos and pregnancy rate in patients with poor ovarian reserve: a randomized controlled trial.

BACKGROUND: Granulocyte colony-stimulating factor (G-CSF) administration increased ovarian preantral follicles and anti-Müllerian hormone (AMH) in animal models with diminished ovarian reserve. We investigated whether G-CSF priming before treatment with assisted reproductive technology (ART) improved embryo development and pregnancy rate while increasing serum AMH in patients with poor ovarian reserve. METHODS: In this prospective randomized open-label controlled trial, 100 patients 20 to 42 years old with AMH below 2 ng/mL were randomized to priming or control groups (50 patients each). None had over 1 ART failure, day-3 follicle-stimulating hormone (FSH) above 30 IU/L, uterine anomalies, or a partner with azoospermia. All patients initially underwent conventional infertility treatment for 2 consecutive cycles in which the priming group but not controls received a subcutaneous G-CSF priming injection during the early luteal phase. Each group then underwent 1 cycle of in vitro fertilization/intracytoplasmic sperm injection and fresh embryo transfer (IVF/ICSI-fresh ET), followed by cryopreserved ET if needed until live birth or embryo depletion. AMH was measured before and after priming. RESULTS: Fertilization rate, embryonic development, and implantation rate by fresh ET were significantly improved by priming. Clinical and ongoing pregnancy rates by IVF/ICSI-fresh ET were significantly higher with priming (30% and 26% in 47 ART patients; 3 delivered with conventional treatment) than in controls (12% and 10% in 49 ART patients; 1 dropped out). With priming, significantly more patients achieved cryopreservation of redundant blastocysts. The cumulative live birth rate was 32% in 50 patients with priming, significantly higher than 14% in 49 controls (relative risk, 2.8; 95% confidence interval, 1.04-7.7). Infants derived from priming had no congenital anomalies, while infant weights, birth weeks, and Apgar scores were similar between groups. Among 4 variables (age, day-3 FSH, AMH, and priming), logistic regression significantly associated age and priming with cumulative live birth. Priming significantly increased serum AMH. No adverse effects of priming were observed. CONCLUSION: G-CSF priming improved embryonic development and pregnancy rate during ART treatment and increased AMH in patients with poor ovarian reserve. Enhanced preantral follicle growth likely was responsible. TRIAL REGISTRATION: UMIN registration in Japan (UMIN000013956) on May 14, 2014.  https://www.umin.ac.jp/ctr/index.htm .