Transplantation of human endometrial perivascular stem cells with hydroxy saffron yellow A promotes uterine repair in rats.

BACKGROUND: Intrauterine adhesions (IUAs) jeopardise uterine function in women, which is a great challenge in the clinic. Previous studies have shown that endometrial perivascular cells (En-PSCs) can improve the healing of scarred uteri and that hydroxysafflor yellow A (HSYA) promotes angiogenesis. The purpose of this study was to observe whether the combination of En-PSCs with HSYA could improve the blood supply and fertility in the rat uterus after full-thickness injury. METHODS: En-PSCs were sorted by flow cytometry, and the effect of HSYA on the proliferation and angiogenesis of the En-PSCs was detected using CCK-8 and tube formation assays. Based on a previously reported rat IUA model, the rat uteri were sham-operated, spontaneously regenerated, or treated with collagen-loaded PBS, collagen-loaded HSYA, collagen-loaded En-PSCs, or collagen-loaded En-PSCs with HSYA, and then collected at both 30 and 90 days postsurgery. HE staining and Masson staining were used to evaluate uterine structure and collagen fibre deposition, and immunohistochemical staining for α-SMA and vWF was used to evaluate myometrial regeneration and neovascularization in each group. A fertility assay was performed to detect the recovery of pregnancy function in each group. RNA-seq was performed to determine the potential mechanism underlying En-PSCs/HSYA treatment. Immunofluorescence, tube formation assays, and Western blot were used to validate the molecular mechanism involved. RESULTS: The transplantation of Collagen/En-PSCs/HSYA markedly promoted uterine repair in rats with full-thickness injury by reducing fibrosis, increasing endometrial thickness, regenerating myometrium, promoting angiogenesis, and facilitated live births. RNA sequencing results suggested that En-PSCs/HSYA activated the NRG1/ErbB4 signaling pathway. In vitro tube formation experiments revealed that the addition of an ErbB inhibitor diminished the tube formation ability of cocultured En-PSCs and HUVECs. Western blot results further showed that elevated levels of NRG1 and ErbB4 proteins were detected in the Collagen/En-PSCs/HSYA group compared to the Collagen/En-PSCs group. These collective results suggested that the beneficial effects of the transplantation of Collagen/En-PSCs/HSYA might be attributed to the modulation of the NRG1/ErbB4 signaling pathway. CONCLUSIONS: The combination of En-PSCs/HSYA facilitated morphological and functional repair in rats with full-thickness uterine injury and may promote endometrial angiogenesis by regulating the NRG1/ErbB4 signaling pathway.

High levels of fatty acid-binding protein 5 excessively enhances fatty acid synthesis and proliferation of granulosa cells in polycystic ovary syndrome.

BACKGROUND: Polycystic ovary syndrome (PCOS) is one of the most complex endocrine disorders in women of reproductive age. Abnormal proliferation of granulosa cells (GCs) is an important cause of PCOS. This study aimed to explore the role of fatty acid-binding protein 5 (FABP5) in granulosa cell (GC) proliferation in polycystic ovary syndrome (PCOS) patients. METHODS: The FABP5 gene, which is related to lipid metabolism, was identified through data analysis of the gene expression profiles of GSE138518 from the Gene Expression Omnibus (GEO) database. The expression levels of FABP5 were measured by quantitative real-time PCR (qRT‒PCR) and western blotting. Cell proliferation was evaluated with a cell counting kit-8 (CCK-8) assay. Western blotting was used to assess the expression of the proliferation marker PCNA, and immunofluorescence microscopy was used to detect Ki67 expression. Moreover, lipid droplet formation was detected with Nile red staining, and qRT‒PCR was used to analyze fatty acid storage-related gene expression. RESULTS: We found that FABP5 was upregulated in ovarian GCs obtained from PCOS patients and PCOS mice. FABP5 knockdown suppressed lipid droplet formation and proliferation in a human granulosa-like tumor cell line (KGN), whereas FABP5 overexpression significantly enhanced lipid droplet formation and KGN cell proliferation. Moreover, we determined that FABP5 knockdown inhibited PI3K-AKT signaling by suppressing AKT phosphorylation and that FABP5 overexpression activated PI3K-AKT signaling by facilitating AKT phosphorylation. Finally, we used the PI3K-AKT signaling pathway inhibitor LY294002 and found that the facilitation of KGN cell proliferation and lipid droplet formation induced by FABP5 overexpression was inhibited. In contrast, the PI3K-AKT signaling pathway agonist SC79 significantly rescued the suppression of KGN cell proliferation and lipid droplet formation caused by FABP5 knockdown. CONCLUSIONS: FABP5 promotes active fatty acid synthesis and excessive proliferation of GCs by activating PI3K-AKT signaling, suggesting that abnormally high expression of FABP5 in GCs may be a novel biomarker or a research target for PCOS treatment.

EHD1 impaired decidualization of endometrial stromal cells in recurrent implantation failure: role of SENP1 in modulating progesterone receptor signalling†.

Recurrent implantation failure (RIF) patients exhibit poor endometrial receptivity and abnormal decidualization with reduced effectiveness and exposure to progesterone, which is an intractable clinical problem. However, the associated molecular mechanisms remain elusive. We found that EH domain containing 1 (EHD1) expression was abnormally elevated in RIF and linked to aberrant endometrial decidualization. Here we show that EHD1 overexpressed in human endometrial stromal cells significantly inhibited progesterone receptor (PGR) transcriptional activity and the responsiveness to progesterone. No significant changes were observed in PGR mRNA levels, while a significant decrease in progesterone receptor B (PRB) protein level. Indeed, EHD1 binds to the PRB protein, with the K388 site crucial for this interaction. Overexpression of EHD1 promotes the SUMOylation and ubiquitination of PRB, leading to the degradation of the PRB protein. Supplementation with the de-SUMOylated protease SENP1 ameliorated EHD1-repressed PRB transcriptional activity. To establish a functional link between EHD1 and the PGR signalling pathway, sg-EHD1 were utilized to suppress EHD1 expression in HESCs from RIF patients. A significant increase in the expression of prolactin and insulin-like growth factor-binding protein 1 was detected by interfering with the EHD1. In conclusion, we demonstrated that abnormally high expression of EHD1 in endometrial stromal cells attenuated the activity of PRB associated with progesterone resistance in a subset of women with RIF.

Maternal and embryonic signals cause functional differentiation of luminal epithelial cells and receptivity establishment.

Embryo implantation requires temporospatial maternal-embryonic dialog. Using single-cell RNA sequencing for the uterus from 2.5 to 4.5 days post-coitum (DPC) and bulk sequencing for the corresponding embryos of 3.5 and 4.0 DPC pregnant mice, we found that estrogen-responsive luminal epithelial cells (EECs) functionally differentiated into adhesive epithelial cells (AECs) and supporting epithelial cells (SECs), promoted by progesterone. Along with maternal signals, embryonic Pdgfa and Efna3/4 signaling activated AECs and SECs, respectively, enhancing the attachment of embryos to the endometrium and furthering embryo development. This differentiation process was largely conserved between humans and mice. Notably, the developmental defects of SOX9-positive human endometrial epithelial cells (similar to mouse EEC) were related to thin endometrium, whereas functional defects of SEC-similar unciliated epithelial cells were related to recurrent implantation failure (RIF). Our findings provide insights into endometrial luminal epithelial cell development directed by maternal and embryonic signaling, which is crucial for endometrial receptivity.

Targeting CD301+ macrophages inhibits endometrial fibrosis and improves pregnancy outcome.

Macrophages are a key and heterogeneous cell population involved in endometrial repair and regeneration during the menstrual cycle, but their role in the development of intrauterine adhesion (IUA) and sequential endometrial fibrosis remains unclear. Here, we reported that CD301+ macrophages were significantly increased and showed their most active interaction with profibrotic cells in the endometria of IUA patients compared with the normal endometria by single-cell RNA sequencing, bulk RNA sequencing, and experimental verification. Increasing CD301+ macrophages promoted the differentiation of endometrial stromal cells into myofibroblasts and resulted in extracellular matrix accumulation, which destroyed the physiological architecture of endometrial tissue, drove endometrial fibrosis, and ultimately led to female infertility or adverse pregnancy outcomes. Mechanistically, CD301+ macrophages secreted GAS6 to activate the AXL/NF-κB pathway, upregulating the profibrotic protein synthesis. Targeted deletion of CD301+ macrophages or inhibition of AXL by Bemcentinib blunted the pathology and improved the outcomes of pregnancy in mice, supporting the therapeutic potential of targeting CD301+ macrophages for treating endometrial fibrosis.

Downregulated INHBB in endometrial tissue of recurrent implantation failure patients impeded decidualization through the ADCY1/cAMP signalling pathway.

PURPOSE: This study aims to identify the mechanism of Inhibin Subunit Beta B (INHBB), a member of the transforming growth factor-ß (TGF-ß) family involved in the regulation of human endometrial stromal cells (HESCs) decidualization in recurrent implantation failure (RIF). METHODS: RNA-seq was conducted to identify the differentially expressed genes in the endometria from control and RIF patients. RT-qPCR, WB, and immunohistochemistry were performed to analyse the expression levels of INHBB in endometrium and decidualised HESCs. RT-qPCR and immunofluorescence were used to detect changes in the decidual marker genes and cytoskeleton after knockdown INHBB. Then, RNA-seq was used to dig out the mechanism of INHBB regulating decidualization. The cAMP analogue (forskolin) and si-INHBB were used to investigate the involvement of INHBB in the cAMP signalling pathway. The correlation of INHBB and ADCY expression was analysed by Pearson's correlation analysis. RESULTS: Our results showed significantly reduced expression of INHBB in endometrial stromal cells of women with RIF. In addition, INHBB was increased in the endometrium of the secretory phase and significantly induced in in-vitro decidualization of HESCs. Notably, with RNA-seq and siRNA-mediated knockdown approaches, we demonstrated that the INHBB-ADCY1-mediated cAMP signalling pathway regulates the reduction of decidualization. We found a positive association between the expression of INHBB and ADCY1 in endometria with RIF (R2 = 0.3785, P = 0.0005). CONCLUSIONS: The decline of INHBB in HESCs suppressed ADCY1-induced cAMP production and cAMP-mediated signalling, which attenuated decidualization in RIF patients, indicating that INHBB is an essential component in the decidualization process.

Hormone replacement therapy alone or in combination with tamoxifen in women with thin endometrium undergoing frozen-thawed embryo transfer: A retrospective study.

Research question: To investigate the effects of two protocols (hormone replacement therapy (HRT) alone or in combination with tamoxifen) on the endometrium and pregnancy outcome of patients with thin endometrium in frozen-thawed embryo transfer (FET) cycles. Design: A total of 465 infertile patients with thin endometrium who underwent FET between January 2020 to June 2021 at the Drum Tower Hospital affiliated with Nanjing University Medical School were retrospectively analyzed. A total of 187 patients were given tamoxifen in addition to HRT (TMXF-HRT group), whereas 278 patients were given only HRT (HRT group). Clinical data were compared between the two groups, including general characteristics, endometrial thickness, and clinical pregnancy outcomes. Results: There were no significant differences in baseline characteristics of all enrolled patients between two groups. Serum progesterone (P) was higher in HRT group than in the TMXF-HRT group (0.28 ± 0.53 ng/mL vs. 0.15 ± 0.25 ng/mL, P = 0.002). There was a significant increase in endometrial thickness in the TMXF-HRT group compared with the HRT group (OR: 1.54, 95% CI: 1.32-1.75, P < 0.001). There were no significant differences in the clinical pregnancy rate, embryo implantation rate, early miscarriage rate, or live birth rate between these two groups. Conclusion: Although tamoxifen when used in combination with hormone replacement therapy can significantly increase endometrial thickness, it may not have a role in improving the pregnancy outcomes of patients with thin endometrium undergoing FET cycles.

Mst1-mediated phosphorylation of Nur77 improves the endometrial receptivity in human and mice.

BACKGROUND: Successful embryo implantation requires the attachment of a blastocyst to the receptive endometrial epithelium, which was disturbed in the women with recurrent implantation failure (RIF). Endometrial ß3-integrin was the most important adhesion molecule contributing to endometrial receptivity in both humans and mice. Nur77 has been proven indispensable for fertility in mice, here we explore the role of Nur77 on embryo-epithelial adhesion and potential treatment to embryo implantation failure. METHODS: The expression and location of Mst1 and Nur77 in endometrium from fertile women and RIF patients were examined by IHC, qRT-PCR and Western blotting. In vitro kinase assay following with LC-MS/MS were used to identify the phosphorylation site of Nur77 activated by Mst1. The phosphorylated Nur77 was detected by phos-tag SDS-PAGE assay and specific antibody against phospho-Nur77-Thr366. The effect of embryo-epithelium interaction was determined in the BeWo spheroid or mouse embryo adhesion assay, and delayed implantation mouse model. RNA-seq was used to explore the mechanism by which Nur77 derived peptide promotes endometrial receptivity. FINDINGS: Endometrial Mammalian sterile 20 (STE20)-like kinase 1 (Mst1) expression level was decreased in the women with RIF than that in the fertile control group, while Mst1 activation in the epithelial cells promoted trophoblast-uterine epithelium adhesion. The effect of Nur77 mediated trophoblast-uterine epithelium adhesion was facilitated by active Mst1. Mechanistically, mst1 promotes the transcription activity of Nur77 by phosphorylating Nur77 at threonine 366 (T366), and consequently increased downstream target ß3-integrin expression. Furthermore, a Nur77-derived peptide containing phosphorylated T366 markedly promoted mouse embryo attachment to Ishikawa cells ([4 (2-4)] vs [3 (2-4)]) and increased the embryo implantation rate (4 vs 1.4) in a delayed implantation mouse model by regulating integrin signalling. Finally, it is observed that the endometrial phospho-Nur77 (T366) level is decreased by 80% in the women with RIF. INTERPRETATION: In addition to uncovering a potential regulatory mechanism of Mst1/Nur77/ß3-integrin signal axis involved in the regulation of embryo-epithelium interaction, our finding provides a novel marker of endometrial receptivity and a potential therapeutic agent for embryo implantation failure. FUNDING: National Key Research and Development Program of China (2018YFC1004400), the National Natural Science Foundation of China (82171653, 82271698, 82030040, 81971387 and 30900727), and National Institutes of Health grants (R01HL103869).

Prolonging the time of progesterone supplementation to improve the pregnancy outcomes of single day 6 blastocyst transfer in frozen-thawed cycles: study protocol for a randomized controlled trial.

BACKGROUND: Infertility is one of the most important and underappreciated reproductive health problems in developing countries. Currently, in vitro fertilization and embryo transfer is the most effective treatment strategy for infertility. In a frozen-thawed cycle, single-blastocyst transfer can not only ensure relatively higher pregnancy and live birth rates but also effectively reduce the risk of maternal and neonatal complications. In frozen-thawed cycles, progesterone is initiated to promote the final phase of endometrial preparation prior to embryo transfer. However, the optimal duration of exposure to progesterone has remained inconclusive. Therefore, we designed a randomized controlled trial (RCT) to compare the effects of different prolonged progesterone transformation times (P+6 and P+7) on the pregnancy outcomes of D6 single blastocyst transfer in a frozen-thawed cycle. METHODS: This is a single-center, prospective, randomized controlled clinical trial involving 900 patients with single blastocyst transfer in the frozen-thawed cycle, aged from 20 to 38 years, with less than three transfers, and with HRT-cycle single D6 blastocyst transfer in the current cycle. Participants will be randomly assigned (1:1) into two parallel groups: the transfer of day 6 blastocysts on the 7th day of progesterone supplementation and the transfer of day 6 blastocysts on the 6th day of progesterone supplementation. The primary outcome measure is the clinical pregnancy rate. Secondary outcome measures include the miscarriage rate and live birth rate. DISCUSSION: This is the first randomized controlled trial to compare the transfer of day 6 blastocysts on the 6th and 7th day of progesterone supplementation. The results of this study will provide evidence for whether to prolong the duration of exposure to progesterone prior to embryo transfer. TRIAL REGISTRATION: ClinicalTrials.gov, ID: NCT04938011. Registered on 19 June 2021.

Primary Cilia Restrain PI3K-AKT Signaling to Orchestrate Human Decidualization.

Endometrial decidualization plays a pivotal role during early pregnancy. Compromised decidualization has been tightly associated with recurrent implantation failure (RIF). Primary cilium is an antenna-like sensory organelle and acts as a signaling nexus to mediate Hh, Wnt, TGFß, BMP, FGF, and Notch signaling. However, whether primary cilium is involved in human decidualization is still unknown. In this study, we found that primary cilia are present in human endometrial stromal cells. The ciliogenesis and cilia length are increased by progesterone during in vitro and in vivo decidualization. Primary cilia are abnormal in the endometrium of RIF patients. Based on data from both assembly and disassembly of primary cilia, it has been determined that primary cilium is essential to human decidualization. Trichoplein (TCHP)-Aurora A signaling mediates cilia disassembly during human in vitro decidualization. Mechanistically, primary cilium modulates human decidualization through PTEN-PI3K-AKT-FOXO1 signaling. Our study highlights primary cilium as a novel decidualization-related signaling pathway.

A novel lncRNA lncSAMD11-1: 1 interacts with PIP4K2A to promote endometrial decidualization by stabilizing FoxO1 nuclear localization.

Decidualization is essential for a successful pregnancy and determines embryo implantation and pregnancy maintenance. Abnormal decidualization is one of the main causes of recurrent implantation failure (RIF). Studies have shown that large amounts of long noncoding RNAs (lncRNAs) are abnormally expressed in endometrial samples from patients with RIF. However, the functional contributions of lncRNAs to decidualization in RIF have not been explored. In this study, we found that lncSAMD11-1:1 was significantly declined in the endometria of patients with RIF. The knockdown of lncSAMD11-1:1 in human endometrial stromal cells (hESCs) restrained decidualization and embryo implantation in vitro, while the overexpression of lncSAMD11-1:1 facilitated hESC decidualization and embryo implantation in vitro and ameliorated decidualization in RIF patients. Mechanistically, lncSAMD11-1:1 and phosphatidylinositol-5-phosphate 4-kinase type 2 alpha (PIP4K2A) translocated out of nucleus and bound to each other during decidualization, thereby inhibiting the phosphorylation of AKT and promoting FoxO1 nuclear localization. These data suggest that lncSAMD11-1:1 might be a critical novel lncRNA functionally required for human decidualization, and the dysregulation of lncSAMD11-1:1 in the endometrium may be a new predisposing factor of RIF.

Adverse impact of elevated progesterone levels on human chorionic gonadotropin trigger day on blastocyst transfer outcomes in gonadotropin-releasing hormone agonist cycles.

OBJECTIVE: Dose an elevated serum progesterone (P) level on the human chorionic gonadotropin (hCG) trigger day have a negative effect on clinical pregnancy outcomes for embryos transferred at different stages of development in long-acting gonadotropin-releasing hormone agonist (GnRHa) in vitro fertilization-embryo transfer (IVF-ET) cycles? STUDY DESIGN: This was a noninterventional, retrospective, observational, single-centre cohort study. A total of 1951 patients received long-acting GnRHa for pituitary downregulation in IVF-ET cycles at Nanjing Drum Tower Hospital from January 2018 to December 2020. The serum P levels on the day of hCG administration were measured, together with other cycle parameters, to explore the relationship between P levels and the clinical pregnancy rate (CPR) of different embryos transferred. RESULTS: When the serum P level on the hCG day was higher than 1.5 ng/mL, the CPR did not decrease significantly. There was no correlation between the CPR of cleavage-stage embryo transfer and the serum P level on the hCG day. In addition, the interaction analysis suggested that the CPR of patients undergoing blastocyst transfer decreased as serum P levels on the hCG day increased. Progesterone levels on the day of hCG administration were closely related to the CPR of blastocyst transfer rather than cleavage-stage embryo transfer. CONCLUSION: An increased serum P level on the day of hCG administration did not affect the CPR of cleavage-stage embryo transfer, but it reduced the CPR of blastocyst transfer cycles.

The relationship between serum oestrogen levels and clinical outcomes of hormone replacement therapy-frozen embryo transfer: a retrospective clinical study.

BACKGROUND: This study aimed to explore the relationship between serum oestrogen (E2) levels before endometrial transformation and pregnancy outcomes of hormone replacement therapy-frozen embryo transfer (HRT-FET) cycles, which has been investigated for years without any consensus. METHODS: A retrospective cohort study of 10,209 cycles HRT-FET cycles was conducted at the Reproductive Medicine Center of Nanjing Drum Tower Hospital from March 2017 to December 2020. A smooth fitting curve was constructed to identify the relationship between serum E2 levels before endometrial transformation and the clinical pregnancy rate. Then, threshold and saturation effect analysis was employed to explore the cut-off value of serum E2 levels. In addition, patients were divided into 2 groups based on their levels of serum E2 measured before progesterone-induced endometrial transformation: Group 1, < 300 pg/mL (n = 6251) and Group 2, ≥ 300 pg/mL (n = 3958). The clinical pregnancy and miscarriage rates of all groups were compared. Further smooth fitting curve analysis was employed by different subgroups segmented according to different endometrial thicknesses. RESULTS: When the serum E2 level was greater than 300 pg/mL, the clinical pregnancy rate decreased significantly (62.9% vs. 59.8%, p < 0.01), but the miscarriage rates were similar (13.5% vs. 15.6%, p = 0.14). While serum E2 level reached or exceeded 1400 pg/mL, there was no significant correlation between the clinical pregnancy rate and E2 level. The clinical pregnancy rate reached its higher level at lower E2 levels, regardless of the different endometrail thicknesses. CONCLUSIONS: Patients with a lower pretransformation serum E2 level (less than 300 pg/mL) have a higher clinical pregnancy rate and there was no correlation between the clinical pregnancy rate and a higher serum E2 level (greater than 1400 pg/mL) in HRT-FET cycles.

Calpain7 negatively regulates human endometrial stromal cell decidualization in EMs by promoting FoxO1 nuclear exclusion via hydrolyzing AKT1.

Endometrial receptivity damage caused by impaired decidualization may be one of the mechanisms of infertility in endometriosis (EMs). Our previous study demonstrated that Calpain-7 (CAPN7) is abnormally overexpressed in EMs. Whether CAPN7 affects the regulation of decidualization and by what mechanism CAPN7 regulates decidualization remains to be determined. In this study, we found CAPN7 expression decreased during human endometrial stromal cell (HESC) decidualization in vitro. CAPN7 negatively regulated decidualization in vitro and in vivo. We also identified one conserved potential PEST sequence in the AKT1 protein and found that CAPN7 was able to hydrolyse AKT1 and enhance AKT1's phosphorylation. Correspondingly, CAPN7 notably promoted the phosphorylation of Forkhead Box O1 (FoxO1), the downstream of AKT1 protein, at Ser319, leading to increased FoxO1 exclusion from nuclei and attenuated FoxO1 transcriptional activity in decidualized HESC. In addition, we detected endometrium CAPN7, p-AKT1, and p-FoxO1 expressions were increased in EMs. These data demonstrate that CAPN7 negatively regulates HESC decidualization in EMs probably by promoting FoxO1's phosphorylation and FoxO1 nuclear exclusion via hydrolyzing AKT1. The dysregulation of CAPN7 may be a novel cause of EMs.

Association between gestational trophoblastic disease (GTD) history and clinical outcomes in in vitro fertilization/intracytoplasmic sperm injection (IVF/ICSI) cycles.

BACKGROUND: Gestational trophoblastic disease (GTD) usually affects young women of childbearing age. After treatment for GTD, 86% of women wish to achieve pregnancy. On account of the impacts of GTD and treatments as well as patient anxiety, large numbers of couples turn to assisted reproductive technology (ART), especially in vitro fertilization/intracytoplasmic sperm injection (IVF/ICSI). But few studies have investigated whether a history of GTD affects the outcomes of IVF/ICSI in secondary infertile patients and how it occurs. We investigate whether a history of GTD affects the IVF/ICSI outcomes and the live birth rates in women with secondary infertility. METHODS: This retrospective cohort study enrolled 176 women with secondary infertility who underwent IVF/ICSI treatment at the reproductive medical center of Nanjing Drum Tower Hospital from January 1, 2016, to December 31, 2020. Participants were divided into the GTD group (44 women with GTD history) and control group (132 women without GTD history matched from 8318 secondary infertile women). The control group and the study group were matched at a ratio of 3:1 according to patient age, infertility duration, number of cycles and body mass index (BMI). We assessed retrieved oocytes and high-grade embryos, biochemical pregnancy, miscarriage, ectopic pregnancy, gestational age at delivery, delivery mode and live birth rates. RESULT(S): We found a significantly reduced live-birth rate (34.1% vs 66.7%) associated with IVF/ICSI cycles in patients with a GTD history compared to those without a GTD history. The biochemical pregnancy and miscarriage rates of the GTD group were slightly higher than those of the control group. In addition, there was a difference in gestational age at delivery between the GTD and control groups (p < 0.001) but no differences in the mode of delivery (p = 0.267). Furthermore, the number of abandoned embryos in the GTD group was greater than that in the control group (p = 0.018), and the number of good-quality embryos was less than that in the control group (p = 0.019). The endometrial thickness was thinner (p < 0.001) in the GTD group. Immunohistochemistry (IHC) showed abnormal endometrial receptivity in the GTD group. CONCLUSION(S): The GTD history of patients undergoing IVF/ICSI cycles had an impact on the live-birth rate and gestational age at delivery, which might result from the thinner endometrium and abnormal endometrial receptivity before embryo transfer.

Transcriptome analysis of eutopic endometrium in adenomyosis after GnRH agonist treatment.

BACKGROUND: Adenomyosis is a chronic gynecological disease characterized by invasion of the uterine endometrium into the muscle layer. In assisted reproductive technology (ART), gonadotropin-releasing hormone agonist (GnRHa) is often used to improve pregnancy rates in patients with adenomyosis, but the underlying mechanisms are poorly understood. METHODS: Eutopic endometrial specimens were collected from patients with adenomyosis before and after GnRHa treatment in the midsecretory phase. RNA sequencing (RNA-Seq) of these specimens was performed for transcriptome analysis. The differentially expressed genes (DEGs) of interest were confirmed by real-time PCR and immunohistochemistry. RESULTS: A total of 132 DEGs were identified in the endometrium of patients with adenomyosis after GnRHa treatment compared with the control group. Bioinformatics analysis predicted that immune system-associated signal transduction changed significantly after GnRHa treatment. Chemokine (C-C motif) ligand 21 (CCL21) was found to be highly expressed in the eutopic endometrium after GnRHa treatment, which may be involved in the improvement of endometrial receptivity in adenomyosis. CONCLUSION: This study suggests that molecular regulation related to immune system-associated signal transduction is an important mechanism of GnRHa treatment in adenomyosis. Immunoreactive CCL21 is thought to regulate inflammatory events and participate in endometrial receptivity in adenomyosis.

The mitochondrial protease LONP1 maintains oocyte development and survival by suppressing nuclear translocation of AIFM1 in mammals.

BACKGROUND: Oogenesis is a fundamental process of human reproduction, and mitochondria play crucial roles in oocyte competence. Mitochondrial ATP-dependent Lon protease 1 (LONP1) functions as a critical protein in maintaining mitochondrial and cellular homeostasis in somatic cells. However, the essential role of LONP1 in maintaining mammalian oogenesis is far from elucidated. METHODS: Using conditional oocyte Lonp1-knockout mice, RNA sequencing (RNA-seq) and coimmunoprecipitation/liquid chromatography-mass spectrometry (Co-IP/LC-MS) technology, we analysed the functions of LONP1 in mammalian oogenesis. FINDINGS: Conditional knockout of Lonp1 in mouse oocytes in both the primordial and growing follicle stages impairs follicular development and causes progressive oocyte death, ovarian reserve loss, and infertility. LONP1 directly interacts with apoptosis inducing factor mitochondria-associated 1 (AIFM1), and LONP1 ablation leads to the translocation of AIFM1 from the cytoplasm to the nucleus, causing apoptosis in mouse oocytes. In addition, women with pathogenic variants of LONP1 lack large antral follicles (>10 mm) in the ovaries, are infertile and present premature ovarian insufficiency. INTERPRETATION: We demonstrated the function of LONP1 in regulating oocyte development and survival, and in-depth analysis of LONP1 will be crucial for elucidating the mechanisms underlying premature ovarian insufficiency. FUNDING: This work was supported by grants from the National Key Research and Development Program of China (2018YFC1004701), the National Nature Science Foundation of China (82001629, 81871128, 81571391, 81401166, 82030040), the Jiangsu Province Social Development Project (BE2018602), the Jiangsu Provincial Medical Youth Talent (QNRC2016006), the Youth Program of the Natural Science Foundation of Jiangsu Province (BK20200116) and Jiangsu Province Postdoctoral Research Funding (2021K277B).

Attenuated monoamine oxidase a impairs endometrial receptivity in women with adenomyosis via downregulation of FOXO1†.

The establishment of endometrial receptivity is a prerequisite for successful pregnancy. Women with adenomyosis possess a lower chance of clinical pregnancy after assisted reproductive technology, which is partially due to impaired endometrial receptivity. The establishment of endometrial receptivity requires the participation of multiple processes, and proper endometrial epithelial cell (EEC) proliferation is indispensable. Monoamine oxidase A (MAOA) is a key molecule that regulates neurotransmitter metabolism in the nervous system. In the present study, we demonstrated a novel role for MAOA in the establishment of endometrial receptivity in women with adenomyosis and in an adenomyotic mouse model. Attenuated MAOA impairs endometrial receptivity by promoting inappropriate proliferation of EECs via the downregulation of FOXO1 during the window of implantation. These results revealed that MAOA plays a vital role in endometrial receptivity in female reproduction.

A high level of KLF12 causes folic acid-resistant neural tube defects by activating the Shh signaling pathway in mice†.

Although adequate periconceptional folic acid (FA) supplementation has reduced the occurrence of pregnancies affected by neural tube defects (NTDs), the mechanisms underlying FA-resistant NTDs are poorly understood, and thus NTDs still remain a global public health concern. A high level of Krüppel-like factor 12 (KLF12) exerts deleterious effects on heath in most cases, but evidence for its roles in development has not been published. We observed KLF12-overexpressing mice showed disturbed neural tube development. KLF12-overexpressing fetuses died in utero at approximately 10.5 days post-coitus, with 100% presenting cranial NTDs. Neither FA nor formate promoted normal neural tube closure in mutant fetuses. The RNA-seq results showed that a high level of KLF12 caused NTDs in mice via overactivating the sonic hedgehog (Shh) signaling pathway, leading to the upregulation of patched 1, GLI-Krüppel family member GLI1, hedgehog-interacting protein, etc., whereas FA metabolism-related enzymes did not express differently. PF-5274857, an antagonist of the Shh signaling pathway, significantly promoted dorsolateral hinge point formation and partially rescued the NTDs. The regulatory hierarchy between a high level of KLF12 and FA-resistant NTDs might provide new insights into the diagnosis and treatment of unexplained NTDs in the future.