Analysis of weighted gene co-expression networks and clinical validation identify hub genes and immune cell infiltration in the endometrial cells of patients with recurrent implantation failure.

Background: About 10% of individuals undergoing in vitro fertilization encounter recurrent implantation failure (RIF), which represents a worldwide social and economic concern. Nevertheless, the critical genes and genetic mechanisms underlying RIF are largely unknown. Methods: We first obtained three comprehensive microarray datasets "GSE58144, GSE103465 and GSE111974". The differentially expressed genes (DEGs) evaluation, enrichment analysis, as well as efficient weighted gene co-expression network analysis (WGCNA), were employed for distinguishing RIF-linked hub genes, which were tested by RT-qPCR in our 30 independent samples. Next, we studied the topography of infiltration of 22 immune cell subpopulations and the association between hub genes and immune cells in RIF using the CIBERSORT algorithm. Finally, a novel ridge plot was utilized to exhibit the potential function of core genes. Results: The enrichment of GO/KEGG pathways reveals that Herpes simplex virus 1 infection and Salmonella infection may have an important role in RIF. After WGCNA, the intersected genes with the previous DEGs were obtained using both variance and association. Notably, the subsequent nine hub genes were finally selected: ACTL6A, BECN1, SNRPD1, POLR1B, GSK3B, PPP2CA, RBBP7, PLK4, and RFC4, based on the PPI network and three different algorithms, whose expression patterns were also verified by RT-qPCR. With in-depth analysis, we speculated that key genes mentioned above might be involved in the RIF through disturbing endometrial microflora homeostasis, impairing autophagy, and inhibiting the proliferation of endometrium. Furthermore, the current study revealed the aberrant immune infiltration patterns and emphasized that uterine NK cells (uNK) and CD4+ T cells were substantially altered in RIF endometrium. Finally, the ridge plot displayed a clear and crucial association between hub genes and other genes and key pathways. Conclusion: We first utilized WGCNA to identify the most potential nine hub genes which might be associated with RIF. Meanwhile, this study offers insights into the landscape of immune infiltration status to reveal the underlying immune pathogenesis of RIF. This may be a direction for the next study of RIF etiology. Further studies would be required to investigate the involved mechanisms.

Clinical outcomes of frozen-thawed embryo transfer in natural cycles with luteinized unruptured follicles.

OBJECTIVE: To assess the effect of luteinized unruptured follicles (LUF) on frozen-thawed embryo transfer cycles performed in natural cycles (FET-NC). METHODS: Retrospective cohort study, held in a university hospital with 3415 cycles for frozen-thawed embryo transfer, performed between June 2019 and September 2022. Using propensity score matching, 242 patients with a diagnosis of LUF (LUF group) were matched with 484 ovulated patients (ovulation group). Stratified by the type of embryo transferred, the LUF group included 168 blastocyst transfer patients (blastocyst group) and 74 cleavage-stage embryo transfer patients (cleavage-embryo group). The ovulation group included 324 patients with blastocyst transfer (blastocyst group) and 160 patients with transferred cleavage-stage embryos. Clinical pregnancy rate was retrospectively analyzed between the LUF and ovulation groups, as well as between each subgroup. RESULTS: After using propensity score matching, the general characteristics of the LUF and ovulation groups were similar. The implantation and clinical pregnancy rates in the LUF group were not significantly different from those in the ovulation group (44.98 % vs. 45.29 %, p = 0.93; 53.72 % vs. 52.48 %, p = 0.75). The implantation and pregnancy rates of transferred cleavage-stage embryos in the LUF group were also not significantly different from those in the ovulation group (32.39 % vs. 36.40 %, p = 0.42; 47.30 % vs. 47.50 %, p = 0.98). The implantation and pregnancy rates of transferred blastocysts in the LUF group were also not significantly different from those in the ovulation group (53.11 % vs. 52.03 %, p = 0.82; 56.55 % vs. 54.94 %, p = 0.73). There was also no significant difference in the miscarriage rate between the groups. CONCLUSION: In the natural cycle, LUF does not affect the clinical pregnancy outcomes of FET. If adequate luteal support is given, the clinical pregnancy outcomes were similar between the LUF group and ovulation group.

Overexpression of hypoxia-inducible factor 1α and excessive vascularization in the peri-implantation endometrium of infertile women with chronic endometritis.

Objective: Chronic endometritis (CE) contributes to impaired endometrial receptivity and is closely associated with poor in vitro fertilization (IVF) outcomes. However, the mechanisms underlying CE are unclear. Here, we investigated the role of the hypoxic microenvironment and endometrial vascularization in the peri-implantation endometrium of infertile women with CE. Methods: This retrospective study involved 15 fertile women and 77 infertile patients diagnosed with CE based on CD138+ ≥1/10 high-power fields (HPFs). The CE patients were divided into Group 1 (CD138+ 1-4/10 HPFs, 53 cases) and Group 2 (CD138+ ≥5/10 HPFs, 24 cases). The expression levels of hypoxia-inducible factor 1α (HIF1α), vascular endothelial growth factor A (VEGFA), and vascular endothelial growth factor receptor 2 (VEGFR2) in peri-implantation endometrium were assessed by qRT-PCR and western blot analyses. Spatial levels of HIF1α, VEGFA, and VEGFR2 in various endometrial compartments was determined using immunohistochemistry and H-score analysis. Microvascular density (MVD) was determined using CD34 staining and scored using Image J. Finally, we used qRT-PCR to assess changes in the expression of HIF1α, VEGFA, and VEGFR2 in CE patients after treatment with first-line antibiotics. Results: Relative to Group 1 and control group, during the implantation window, protein and mRNA levels of HIF1α, VEGFA, and VEGFR2 were markedly high in Group 2 (P<0.05). H-score analysis showed that HIF1α, VEGFA, and VEGFR2 in the luminal, glandular epithelium, and stromal compartments were markedly elevated in Group 2, comparing to control group and Group 1 (P<0.05). Moreover, markedly elevated MVD levels were observed in Group 2. Notably, the above indexes did not differ significantly in the control group versus Group 1. Treatment with antibiotics significantly suppressed the endometrial HIF1α and VEGFA levels in CE-cured patients. Conclusions: Here, we for the first time report the upregulation of HIF1α, VEGFA, and VEGFR2, as well as excessive endometrial vascularization in the peri-implantation endometrium of CE patients. Our findings offer new insights into reduced endometrial receptivity in CE-associated infertility.

The Clinical Efficacy of Personalized Embryo Transfer Guided by the Endometrial Receptivity Array/Analysis on IVF/ICSI Outcomes: A Systematic Review and Meta-Analysis.

Objective: To assess the prevalence of displaced window of implantation (WOI) in infertile women, and the clinical utility of personalized embryo transfer (pET) guided by the endometrial receptivity array/analysis (ERA) on IVF/ICSI outcomes. Methods: The protocol was registered at Prospero: CRD42020204237. We systematically searched all published English literature related to the prevalence of WOI displacement and ongoing pregnancy rate/live birth rate in the overall good-prognosis infertile patients (GPP) and/or repeated implantation failure (RIF) patients undergoing IVF/ICSI-ET cycles after ERA test until August 2021. Result(s): 11 published studies were enrolled in the final analysis. The estimate of the incidence of WOI displacement based on ERA was 38% (95%CI 19-57%) in GPP and 34% (95%CI 24-43%) in RIF, respectively. There was no difference in OPR/LBR between patients undergoing routine ET without ERA test and those who following pET with ERA (39.5 vs. 53.7%, OR 1.28, p = 0.49, 95%CI 0.92-1.77, I 2 = 0%) in relative GPP. Notably, the meta-analysis revealed that OPR/LBR of patients with RIF undergoing pET who had non-receptive ERA increased to the level of to those undergoing sET with receptive ERA (40.7 vs.49.6%, OR 0.94, p = 0.85, 95%CI 0.70-1.26, I 2 = 0%). Conclusion: Considering the approximately one third of infertile women could suffered from displaced WOI, the ERA test emerged as a promising tool. Although the present meta-analysis demonstrates that patients with general good-prognosis may not benefit from ERA, pET guided by ERA significantly increases the chances of pregnancy for non-receptive patients with RIF of endometrial origin.

Dyslipidemia Is Negatively Associated With the Cumulative Live-Birth Rate in Patients Without PCOS Following IVF/ICSI.

Objective: To evaluate the effect of dyslipidemia on the cumulative live-birth rate (cLBR) in patients without polycystic ovary syndrome (PCOS) undergoing in vitro fertilization/intracytoplasmic sperm injection-embryo transfer (IVF/ICSI-ET) cycles. Methods: A total of 1,132 patients from the Yantai Yuhuangding Hospital Affiliated to Qingdao University from January 2016 to December 2017 were retrospectively included. The subjects were distributed into two groups based on their lipid profiles, namely, dyslipidemia group (n = 195) and control group (n = 937). The clinical and laboratory parameters of the two groups were analyzed, and a multivariate logistic regression analysis of the cLBR was conducted. In addition, subgroup analysis was carried out to avoid deviation according to the body mass index (BMI). Results: Patients with dyslipidemia had significantly greater BMI and longer duration of infertility, as well as lower antral follicle count and basal follicle-stimulating hormone level compared with patients without dyslipidemia. Stratified analysis showed that dyslipidemia was associated with a significantly higher total gonadotrophin dosage required for ovarian stimulation as well as lower number of oocytes retrieved, independent of obesity. The live-birth rate in fresh cycle and cLBR were higher in the control group, although the difference between the groups was not significant (54.9% vs. 47.3%, p = 0.116; 67.6% vs. 62.1%, p = 0.138). However, multivariate logistic regression analysis adjusting for potential confounders showed that dyslipidemia was negatively associated with cLBR (OR, 0.702, 95% CI, 0.533-0.881, p = 0.044). Conclusion: Our findings demonstrate for the first time that dyslipidemia has a deleterious impact on cLBR, independent of obesity, in non-PCOS population considered to have good prognosis. Assessment of serum lipid profiles as well as the provision of nutritional counseling is essential for increasing successful outcomes in assisted reproductive techniques.

Esculentoside A rescues granulosa cell apoptosis and folliculogenesis in mice with premature ovarian failure.

Follicular atresia is one of the main processes for the loss of granulosa cells and oocytes from the mammalian ovary and any impairment to premature ovarian failure. Large numbers of studies have demonstrated that granulosa cell apoptosis causes follicular atresia, yet the rescue of these cells remains elusive. We aimed to use Esculentoside A (3-O-b-D-glucopyranosyl-1, 4-b-D-xylopyranosyl) phytolaccagenin, a saponin extracted from Phytolacca esculenta roots, as a potential rescue agent for the apoptosis of granulosa cells. Our results revealed the rescue of normal body and ovary weights, normal ovarian histo-architecture of ovaries, and hormones levels with regular estrus cycle. Consistently, the expression of proliferating and anti-apoptotic markers, i.e. KI67 and BCL-2 in granulosa cells, was enhanced. Meanwhile, the expressions of pro-apoptotic markers, which were BAX and CASPASEs (CASPASE-9 and CASPASE-3), were prominently reduced in Esculentoside A-induced premature ovarian failure mice. Additionally, PPARγ, a potential therapeutic target, has also rescued its expression by treating the premature ovarian failure mice with Esculentoside A. Our results advocated that Esculentoside A could restore folliculogenesis in premature ovarian failure mice. Furthermore, it has the potential to be investigated as a therapeutic agent for premature ovarian failure.