Inactivation of Yes-Associated Protein Mediates Trophoblast Dysfunction: A New Mechanism of Pregnancy Loss Associated with Anti-Phospholipid Antibodies?

Pregnancy morbidity induced by anti-phospholipid antibodies (aPL+/PM+) is mainly thought to arise from placental abnormalities. We attempted to investigate the effect of aPL on the activity of Yes-associated protein (YAP) in the trophoblast and how YAP regulated human trophoblasts function. Thus, HTR-8 cells were treated with IgG purified from aPL+/PM+ women or normal controls. We found that aPL+/PM+ IgG impacted YAP activity via abrogating YAP expression. Further investigation of the anti-ß2GPI-IgG/ß2GPI complex showed an inhibition of nuclear YAP level and translocation in a dose-dependent manner, which might be rescued by progesterone in HTR-8 cells. YAP overexpression or knockdown HTR-8 cells were established for the evaluation of cell function and related gene expression in vitro. Loss of YAP arrested cell cycles in the G2/M phase, accelerated cell apoptosis by increasing the ratio of Bax/Bcl2, and disrupted MMP2/9-mediated cell migration and angiogenesis tube formation by VEGF. These findings support a new mechanism of PM associated with aPL through which YAP inactivation induced by aPL perturbs the trophoblast cell cycle, apoptosis, migration, and angiogenesis, finally developing into pregnancy failure.

Lower Fiber Consumption in Women with Polycystic Ovary Syndrome: A Meta-Analysis of Observational Studies.

Polycystic ovary syndrome is a common endocrine disorder associated with metabolic abnormalities and gut microbiota dysbiosis. The deficiency of dietary fiber, a crucial nutrient in the daily diet, is also associated with a wide range of metabolic and reproductive abnormalities, as well as an altered gut microbial ecosystem. This study is a meta-analysis to summarize the available evidence on the dietary fiber intake level in PCOS patients. Databases of PubMed, Embase, Cochrane Library, Web of Science, and ClinicalTrials.gov were searched for observational studies, and 13 studies were finally included. The pooled standardized mean difference (SMD) with the 95% confidence interval (CI) of daily dietary fiber intake and total energy intake were calculated using the random-effects model. The pooled result (12 studies) on absolute dietary fiber intake showed that while there was no significant difference in the total energy intake [−0.17 (−0.44, 0.09), p = 0.208], the dietary fiber intake was significantly lower in PCOS women than those of controls [−0.32 (−0.50, −0.14), p < 0.001]. However, significant heterogeneity was detected across the studies (I2 = 65.6%, p = 0.001). Meta-regression suggested that geographic region and dietary assessment method may confer borderline significance of influence on the heterogeneity. The pooled result (two studies) on dietary fiber intake which adjusted for total energy intake, however, showed no significant difference [−2.11 (−4.77, 0.56), p = 0.122]. In subgroup analyses based on absolute dietary fiber intake, a lower dietary fiber intake in PCOS was observed in studies conducted in Asia, adopted food diary or records or food recall as the dietary assessment method, had a case−control study design, or used Rotterdam criteria for PCOS diagnosis. The difference in SMD was still significant in the adult subgroup or in studies matched or unmatched for age.

Rapamycin maintains the primordial follicle pool and protects ovarian reserve against cyclophosphamide-induced damage.

Any abnormal activation of primordial follicles and subsequent depletion can irreversibly diminish the ovarian reserve, which is one of the major chemotherapy-induced adverse effects in young patients with cancer. Herein, we investigated the effects of rapamycin on the activation and development of ovarian follicles to evaluate its fertility-sparing therapeutic value in a cyclophosphamide (CTX)-treated mouse model. Based on ovarian histomorphological changes and follicle counting in 50 SPF female C57BL/6 mice, daily administration of 5 mg/kg rapamycin for 30 days was deemed an ideal dosage and duration for administration in subsequent experiments. Compared with the control group, rapamycin treatment inhibited the activation of quiescent primordial follicles, with no obvious side effects observed. Finally, 48 mice were randomly divided into four groups: control, rapamycin-treated, cyclophosphamide-treated, and rapamycin intervention. Body weight, ovarian histomorphological changes, number of primordial follicles, DDX4/MVH expression, apoptosis of follicular cells, and expression of apoptosis protease-activating factor (APAF)-1, cleaved caspase 3, and caspase 3 were monitored. Co-administration of rapamycin reduced primordial follicle loss and the development of follicular cell apoptosis, thereby rescuing the ovarian reserve after CTX treatment. On analyzing the mTOR signaling pathway, we observed that rapamycin significantly decreased CTX-mediated overactivation of mTOR and its downstream molecules. These findings suggest that rapamycin exhibits potential as an ovarian-protective agent that could maintain the ovarian primordial follicle pool and preserve fertility in young female patients with cancer undergoing chemotherapy.

CD55 is upregulated by cAMP/PKA/AKT and modulates human decidualization via Src and ERK pathway and decidualization-related genes.

Decidualization is an essential process for embryo implantation and maintenance of pregnancy, and abnormal decidualization contributed to several pregnancy disorders like a miscarriage. The objective of this study was to explore the regulation and function of CD55 in human decidualization. By immunohistochemical staining, it was found that CD55 expression was higher in first-trimester decidua than in the endometrium. In both primary endometrial stromal cells and immortalized cell line T-hESCs, CD55 was upregulated by induction of in vitro decidualization with medroxyprogesterone acetate (MPA) and 8-Br-cAMP. During decidualization in vitro, CD55 was stimulated by 8-Br-cAMP in a time- and concentration-dependent manner, which was reversed by a PKA inhibitor H89 and partially by an AKT activator SC79. Knocking down CD55 expression diminished the expression of decidualization markers prolactin (PRL) and insulin-like growth factor-binding protein 1 (IGFBP1), accompanied by inhibition of Src, aberrant activation of ERK and decreased expression of several decidualization-related genes, including FOXO1, EGFR, and STAT3. Furthermore, the decidua of unexplained miscarriage women and the endometrium of unexplained infertile women both exhibited decreased CD55 expression. Collectively, these findings revealed that 8-Br-cAMP promotes CD55 expression via PKA activation and AKT dephosphorylation, and decreased CD55 impairs decidualization by inactivation of Src, aberrant activation of ERK pathway, and compromised expression of decidualization-related genes, indicating that CD55 deficiency may contribute to the pathogenesis of spontaneous miscarriage and infertility.

Identification of predictors of the ovarian response to clomiphene citrate in infertile women with polycystic ovary syndrome: a prospective cohort study.

OBJECTIVE: To identify predictors of the ovarian response to clomiphene citrate (CC) in infertile patients with polycystic ovary syndrome (PCOS). METHODS: We performed a prospective cohort study of infertile patients with PCOS. The participants underwent assessments of their physical, endocrine, and metabolic characteristics, and treatment with CC at an initial dose of 50 mg/day and a maximum of 100 mg/day between days 3 and 7 of their menstrual cycles. Participants who ovulated were identified as responders and those who did not as non-responders. RESULTS: Of the 72 participants, 48 (66.7%) were identified as responders and 24 as non-responders. Sex hormone-binding globulin (SHBG) (odds ratio 1.022, 95% confidence interval: 1.000-1.045) was found to be associated with the ovarian response to CC using logistic multivariate regression analysis. Receiver operating characteristic analysis also showed that SHBG was a significant predictor of the response to CC (area under the curve 0.799). CONCLUSION: We have shown that SHBG is the best prognostic indicator of an ovulatory response to CC. However, larger prospective studies, in which more variables are assessed, are required to confirm this finding and to identify appropriate cut-off values.

Toll-Like Receptor 4 Inhibits Estradiol Secretion via NF-κB Signaling in Human Granulosa Cells.

Toll-like receptor 4 (TLR4) may play a critical role in regulating follicular development. Data are scarce on the role of TLR4 in the follicle. This study investigated the effects of TLR4 on steroidogenesis in human granulosa cells. Immunohistochemical analysis revealed stage-specific expression of TLR4 in the mouse ovarian cycle, and immunofluorescence showed TLR4 expression in the human granulosa-like tumor cell line (KGN). TLR4 agonist lipopolysaccharides (LPS) significantly inhibited follicular development and synthesis of estradiol (E2) in mice. In KGN cells, TLR4 activation significantly inhibited CYP19A1, FSHR and StAR, and TLR4 inhibition reversed these effects. TLR4 activation also inhibited forskolin-induced secretion of E2 by inhibiting CYP19A1, with no effect on progesterone. Further studies showed activation of p38, JNK and NF-κB signaling after TLR4 activation. Subsequent analyses showed that an NF-κB antagonist reversed the inhibitory effects on CYP19A1 expression and E2 secretion. Together, our results suggest that TLR4 activation may suppress CYP19A1 expression and E2 secretion via NF-κB signaling in human granulosa cells, with important implications for the regulation of ovarian pathophysiology.

Dydrogesterone treatment for menstrual-cycle regularization in abnormal uterine bleeding - ovulation dysfunction patients.

BACKGROUND: Dydrogesterone has shown significant efficacy in treatment of irregular menstrual cycle due to abnormal uterine bleeding - ovulation dysfunction (AUB-O), but there were few relevant studies. This observational study was designed to evaluate the effectiveness of dydrogesterone for the treatment of Chinese patients with AUB-O. AIM: To evaluate the effects of dydrogesterone on menstrual-cycle (MC) regularization and metabolism in the patients with AUB-O. METHODS: A prospective, non-interventional, single-arm, post-marketing observational study was conducted. Chinese women aged 16 years or above with AUB-O who had been prescribed dydrogesterone were enrolled. The patients were treated with dydrogesterone 10 mg from day 16 to day 25 of each cycle, consecutively for at least 3 cycles. The main outcome was defined as the percentage of patients whose MCs returned to normal (defined as 21 d < menstrual cycle ≤ 35 d) after three cycles of dydrogesterone treatment. RESULTS: One hundred and fourteen women with AUB-O were enrolled in the present study. Of 89 patients who completed treatment, 72 (80.9%) achieved a regular MC at the end of the 3rd circle. The level of androgen, including testosterone and dehydroepiandrosterone sulfate, declined significantly (P = 0.01 and 0.031, respectively), whereas other hormone levels remained steady. During the treatment, 44/80 (55.0%) subjects in the per-protocol set had reported biphasic basal body temperature. CONCLUSION: Dydrogesterone therapy was effective in achieving MC regularization for Chinese patients with AUB-O.

Follicle loss and PTEN/PI3K/mTOR signaling pathway activated in LepR-mutated mice.

Female mice (Y123F) with substitution mutations introduced through homologous gene targeting, replacing the three tyrosine residues of LepR, Tyr985, Tyr1077, and Tyr1138 with phenylalanine, could induce infertility. This study aimed to describe the reproductive alteration and to explore its mechanism. We compared the reproductive characteristics in the female homozygous (HOM) Y123F mice and wild-type (WT) littermates, analyzing the expression of downstream molecules of LepR, like protein kinase B (Akt)/mammalian target of rapamycin (mTOR), phosphatase and tensin homolog deleted on chromosome 10 (PTEN) and insulin receptor substrate (IRS) in the ovaries. The results showed that 10-week old female Y123F HOM exhibited no reproductive periods, declined anti-mullerian hormone (AMH) levels in the serum and ovaries, reduced primordial follicles, primary follicles, secondary follicles, antral follicles and hardly no corpus lutea (all p < .05). The phosphorylation of downsream Akt, mTOR, S6K1 and eIF4B of LepR were all elevated in the ovaries of the mutated female mice. They also presented a decreased phosphorylation of IRS-1, IRS-2, and PTEN, and a strengthened phosphorylation of FOXO-3A in the ovaries. In conclusions, LepR mutation could result in follicle loss and activation of PTEN/PI3K/Akt/mTOR pathway in adult female mice, independent of insulin signaling pathway.

Decay-accelerating factor promotes endometrial cells proliferation and motility under ovarian hormone stimulation.

The intent of the study was to explore the elevating expression of decay-accelerating factor(DAF) exerts influence on biological behaviors of endometrial stromal cells except in classical immunology on the basis of bioinformatic statistics and clinical miscarriages findings suggesting its potential role in the establishment of endometrial receptivity. We confirmed that DAF locates on the cellular surface of endometrial epithelium and stroma. By using plasmid transfection to down-regulate DAF expression in primary endometrial stromal cells(ESCs), we discovered that DAF expression in ESCs increases in response to estradiol and progesterone stimulation in dose- and time-dependent manners; moreover, tamoxifen and RU486 stimulations to block estrogen receptors(ERs) and progesterone receptors(PRs) respectively result in reduced DAF mRNA and protein, and it is more obvious to block PRs. Meanwhile, knocked-down DAF in ESCs weakens the proliferation, migration and invasion of endometrial cells. Cell cycle analysis showed knocked-down DAF accumulates cells in S phase and diminishes cells in G0/G1 phase, which substantiates DAF mediates endometrial cells proliferation. In conclusion, DAF is a potential molecule involving in endometrial cellular proliferation and motility to verify up-expressed DAF during the WOI may facilitate endometrial physiobiological behavior changes, which shed light on DAF function and potential role in the endometrial receptivity establishment.

Follicle Loss and Apoptosis in Cyclophosphamide-Treated Mice: What's the Matter?

With increasing numbers of young female cancer survivors following chemotherapy, chemotherapy-induced fertility loss must be considered. Menstrual disorder and infertility are of particular concern in female cancer patients. We showed that treatment with the alkylating agent cyclophosphamide (CTX) could cause severe primordial follicle loss and growing follicle apoptosis, resulting in loss of ovarian reserve. SPF C57BL/6 female mice were treated with a single dose of 120 mg/kg of CTX or saline as a control, and both sides of ovaries were collected three or seven days after injection. Following CTX treatment, the ovaries were mostly composed of collapsed oocytes and presented marked cortical fibrosis and a reduced number of follicles, especially primordial follicles. The loss of primordial follicles was confirmed by primordial follicle counting, immunohistochemistry and Western blot detection of DDx4/MVH. Follicle apoptosis was tested by a TUNEL assay and the number of TUNEL-positive follicle cells increased, as expected, in CTX-treated mice. Furthermore, expression of APAF-1 and cleaved caspase-3 was also increased after CTX treatment. Analysis of the PI3K/Akt/mTOR signaling pathway showed that CTX increased phosphorylation of Akt, mTOR and downstream proteins without affecting total levels. These results demonstrated that the CTX treatment led to the hyperactivation of the PI3K/Akt/mTOR signaling pathway in ovaries which may be related to primordial follicle loss and growing follicle apoptosis.