Biology of Reproduction最新文献

The corpus luteum (CL) produces progesterone to support pregnancy but undergoes luteolysis in response to prostaglandin F2A (PGF2A) when pregnancy is absent. Developing CL resist PGF2A-induced regression. This acquisition of luteolytic capacity (ALC) occurs around day 5 of the estrous cycle in cattle. To understand the mechanisms underlying ALC, we evaluated transcriptomic and proteomic changes in the CL on days 4 and 6 of the estrous cycle. 1190 transcripts were greater on day 4 and 119 were greater on day 6 (Padj <0.05, log2FC > |1|). In the proteomics dataset, 2 proteins were greater on day 4, with 12 greater on day 6 (Padj <0.05, log2FC > |1|). Data were integrated with datasets of luteal changes during luteolysis and during early pregnancy, and gene ontology analysis was used to identify candidate pathways regulating luteal PGF2A responsiveness, including apoptosis, cellular stress response, and cytokine signaling. Apoptosis regulators increased on day 6 and increased in response to PGF2A in vivo, indicating a likely role in luteolytic cell death. Changes to regulators of antigen presentation and T cell programming suggested increased T cell activation on day 6. A luteal cell culture experiment revealed that apoptosis pathways were regulated by interferon gamma and immune pathways were regulated by luteinizing hormone in vitro. In summary, integration across datasets from multiple luteal states allowed identification of proposed mediators governing luteal survival and PGF2A response. Apoptosis and immune-related pathways, changes indicating increased activated T cells, may poise the CL to regress in response to PGF2A.

Secretoneurin (SN) is a small, bioactive proteolytic product of secretogranin II (SCG2), a chromogranin protein involved in secretory vesicle packaging. SCG2 co-aggregates with luteinizing hormone (LH) in pituitary gonadotrope secretory granules and thus has long been associated with GnRH-regulated LH secretion. Previously, the SCG2-derived peptide SN has been identified as a regulator of LH production and secretion in fish and a murine gonadotrope-like cell line (LβT2). Though high levels of SCG2/SN are present in the mammalian pituitary, the role of pituitary SN has not been identified in mammals. It is further unknown if pituitary SN function varies with age and sex. Here, we identify a decrease in levels of precursor SCG2, and a trend towards increased levels of SN throughout postnatal developmental, particularly in females, following the onset of puberty. Additionally, proteomic and bioinformatic analyses of adult male and female pituitaries revealed protein-protein interactions between SCG2 and various peptides, many associated with prohormone processing and regulated secretion. Furthermore, we demonstrate for the first time that SN is capable of stimulating LH secretion from ex vivo mouse pituitaries, an effect previously documented only in non-mammalian vertebrate species and LβT2 cells. Exogenous SN stimulation significantly increased LH release from adult male (1.4x; p<0.01) and proestrous female (2.4x; p<0.0001) murine pituitaries. Notably, we found that SN treatment had no observed effect on LH secretion in prepubertal (PND14) male and female pituitaries, indicating that SN may function primarily in mature gonadotropes to promote fertility and estrous cycle progression.

The placenta is critical for fetal development, supporting nutrient transport, while also isolating the developing conceptus from adverse maternal signals. The porcine placenta creates an enzymatic barrier for thyroid hormones following activation of the fetal endocrine system in early gestation. Perturbation of the fetal thyroid system can have significant developmental consequences. However, research examining the relationship between fetal thyroid hormone availability, the placental epigenome, and global effects on placental gene expression is limited, particularly in swine. Existing work on select placental genes suggests a potential compensatory response to fetal endocrine disruption during late gestation. We hypothesize that global placental epigenetic and transcriptomic changes may arise in response to induced fetal hypothyroidism. To address this hypothesis, we analyzed placental tissues from gestation day 86 fetuses exposed to 21-day maternal methimazole (MMI) treatment and matched controls. CUT&RUN profiling revealed differential histone mark enrichment, with enrichment of both marks predominantly gained in MMI samples. The placental histone mark status from female fetuses was more substantially altered in response to maternal MMI treatment than placental tissue from male fetuses. In contrast, while differential gene expression was apparent between control and MMI conditions, transcriptional changes were more pronounced in placental tissue from male fetuses. Integration of these datasets revealed treatment- and sex-dependent epigenetic and transcriptional variation. This work expands our understanding of the intricate interplay between fetal thyroid hormone levels and placental gene expression regulation and provides novel insights into the existence of sex-specific differences in placental epigenetic and transcriptomic responses to fetal endocrine disruption.

Introduction: Intrauterine adhesion (IUA) is a leading cause of secondary infertility, characterized by fibrosis and impaired endometrial regeneration. Although inflammation and fibrotic pathways have been studied, the metabolic mechanisms remain unclear. This study aimed to identify metabolic alterations in intrauterine adhesion and to evaluate whether L-carnitine (LCA) supplementation could restore mitochondrial function and alleviate fibrosis.

Material and methods: Untargeted metabolomic profiling was performed on endometrial tissues from patients with IUA and controls using liquid chromatography-tandem mass spectrometry. An ethanol-induced mouse model of IUA was established and treated with LCA by intraperitoneal injection. Histological, immunohistochemical, and ultrastructural analyses assessed fibrosis, inflammation, and mitochondrial integrity. In vitro, lipopolysaccharide-stimulated endometrial epithelial cells were used to evaluate fatty acid oxidation (FAO), lipid droplet accumulation, mitochondrial function, reactive oxygen species (ROS) production, lipid peroxidation, and transforming growth factor beta secretion after L-carnitine treatment.

Result: Metabolomics revealed profound depletion of carnitine and derivatives in IUA, accompanied by enrichment of pro-inflammatory prostaglandins and fibrosis-associated metabolites. LCA treatment in mice restored uterine morphology, reduced apoptosis, fibrosis, and inflammation, and improved pregnancy outcomes. In vitro, LCA restored FAO activity, reduced lipid droplet accumulation, preserved mitochondrial membrane potential, suppressed ROS and lipid peroxidation, and decreased TGF-β secretion in LPS-stimulated cells.

Conclusion: This study establishes carnitine deficiency as a key driver of IUA, demonstrating that it disrupts mitochondrial FAO, promotes oxidative stress, and culminates in fibrosis. LCA supplementation not only restores metabolic homeostasis and reduces fibrotic progression but also improves reproductive outcomes, positioning it as a mechanistically grounded therapy for IUA.

Background: Premature rupture of membranes (PROM) is a major cause of preterm birth and neonatal mortality. Tight junctions are critical for maintaining fetal membrane barrier integrity. Ras-related protein Rab-3B (RAB3B), a key regulator of vesicle trafficking, may influence barrier homeostasis; however, its role in PROM remains unclear.

Methods: Whole-transcriptome RNA sequencing was performed on fetal membrane tissues from 31 PROM patients and 10 full-term controls to identify differentially expressed genes. A PROM mouse model was established to access RAB3B expression and related pathological changes. In vitro, RAB3B was overexpressed or slienced in human amniotic epithelial cells (hAECs) to evaluate its effect on cell migration, epithelial-mesenchymal transition (EMT), epithelial permeability, and tight junction integrity. Extracellular vesicles (EVs) were isolated to explore the role of RAB3B in EV internalization and EV-mediated regulation of tight junctions.

Results: RAB3B expression was significantly downregulated in fetal membranes from PROM patients and PROM mice. PROM mice exhibited increased membrane permeability, enhanced apoptosis, elevated pro-inflammatory cytokine levels, and reduced expression of tight junction proteins. In hAECs, RAB3B overexpression promoted cell migration, suppressed EMT, reduced epithelial permeability, and upregulated tight junction proteins, whereas RAB3B knockdown produced opposite effects. Furthermore, RAB3B enhanced EV internalization and potentiated the beneficial effects EVs on tight junction integrity.

Conclusion: RAB3B is markedly downregulated in PROM and plays a critical role in maintaining fetal membrane barrier integrity. By enhancing tight junctions and promoting EV-mediated material transport, RAB3B contributes to fetal membrane repair, highlighting its potential as a therapeutic target for PROM prevention and treatment.

The clinical management of unexplained recurrent pregnancy loss (URPL) poses significant challenges. Due to its unclear etiology, current treatment approaches exhibit limited therapeutic efficacy. Aberrant macrophage polarization towards the pro-inflammatory M1 phenotype at the maternal-fetal interface has been recognized as a pivotal factor contributing to the pathogenesis of URPL. However, therapies aimed at modulating macrophage polarization remain scarce. In this study, a high repetitive frequency pulsed magnetic field (HRFPMF) was employed as an alternative approach to optimize the biophysical parameters for inducing macrophage polarization, elucidate the underlying molecular mechanisms, and evaluate the therapeutic potential of HRFPMF-treated macrophages in abortion-prone mice. The findings demonstrated that HRFPMF effectively promoted the polarization of a diverse range of M2 macrophages in vitro. Mechanistic investigations revealed that HRFPMF-induced calcium influx coordinated cytoskeletal reorganization, thereby driving the phenotypic switch of macrophages. Functionally, HRFPMF-treated macrophages promoted the proliferation, migration, invasion and secretory capacity of trophoblasts. More importantly, adoptive transfer of HRFPMF-treated macrophages significantly improved pregnancy outcomes in abortion-prone mice. This was achieved through enhanced M2 polarization at the maternal-fetal interface and promotion of placental vascular remodeling. Collectively, this study highlights HRFPMF as a promising alternative, drug-free immunomodulatory strategy for the management of URPL, offering a novel approach to address this complex and challenging clinical condition.

Polycystic ovary syndrome is the most common endocrine condition in women and anovulatory cause of female infertility. While a pro-inflammatory cytokine and leukocyte bias in systemic circulation is well-documented in PCOS, it is not known how this inflammation extends to or affects the ovary. Additionally, the relationship between ovulation and inflammation in PCOS is not well-defined. We hypothesize that the ovarian follicular immune environment in PCOS is uniquely dysregulated, and that resolving anovulation through ovulation induction is not sufficient to alleviate this dysregulation. Using single-cell RNA and surface protein analysis of peripheral blood and follicular fluid from patients undergoing in vitro fertilization, we discovered that both control and PCOS follicles were immunologically distinct from circulation. At a systemic level, we find that ovulation induction in PCOS may alleviate systemic inflammation. In contrast, while healthy control ovaries experienced acute immune-directed ovulatory signaling, PCOS ovarian follicles were deficient in key pro-ovulatory cell to cell communication, and displayed instead a chronic low-grade inflammatory state with fibrotic features. Taken together, a picture emerges where acute ovulation demonstrates a well-ordered series of follicle-specific immune information flows, which are disrupted and replaced by low grade chronic inflammation in the PCOS follicle.

Programmable gene editing tools, particularly CRISPR/Cas9 and its advanced derivatives (base and prime editors), have revolutionized biomedical research and offer unprecedented potential for studying human embryogenesis and correcting monogenic diseases. This review systematically examines the evolution and challenges of these technologies in human embryos and mammalian models. We trace key methodological advancements, from initial studies hampered by low efficiency and mosaicism to refined strategies like RNP delivery and base editing that improved precision. A critical shift occurred with the discovery that CRISPR/Cas9 can cause severe on-target damage, such as large deletions and chromosomal loss, redirecting the field's focus toward safety. We present a comparative analysis of editing efficiencies across species (human, mouse, primate, pig, cow, rabbit) and tools (Cas9, BEs, PEs), consistently demonstrating the superiority of RNP for precise editing. Fundamental barriers to clinical translation are discussed, including the trade-off between efficiency and mosaicism, persistent off-target effects, and profound ethical concerns. The review concludes that while somatic gene therapy advances rapidly, heritable genome editing remains premature due to unresolved risks. Future progress depends on developing safer editors, understanding on-target consequences, and adhering to rigorous ethical standards.

Background: Advanced maternal age (AMA, ≥35 years) is increasingly common and is accompanied by rising threatened abortion (AMA-TA) rates, yet its molecular basis remains unclear.

Objective: To elucidate AMA-TA mechanisms by integrating metabolomics and transcriptomics, providing a foundation for biomarker and therapeutic discovery.

Methods: Untargeted serum metabolomics was performed in 9 AMA-TA patients and 7 age-matched healthy pregnant women. An AMA-TA mouse model was induced by mifepristone (4 mg/kg) to assess embryo resorption, placental morphology, and serum hormones (ELISA). Serum metabolomics and placental transcriptomic profiling (RNA-seq) were then conducted in AMA-TA mice to characterize metabolic and gene expression alterations. Cross-species and multi-omics integration was performed using HomoloGene and MetaboAnalyst 5.0. Key steroid biosynthesis-related genes were finally validated by RT-qPCR.

Results: Human serum metabolomics revealed the differential metabolites were mainly enriched in steroid hormone biosynthesis, lipid metabolism, and amino-acid metabolism. The AMA-TA model showed higher embryo resorption, abnormal placental architecture, and reduced progesterone and chorionic gonadotropin. RNA-seq revealed 111 up- and 1337 downregulated genes enriched in 68 pathways. Consistently, serum metabolomics in AMA-TA mice also showed significant metabolic disturbances, prominently involving steroid hormone biosynthesis. Integrated analysis converged on steroid hormone biosynthesis as a shared key dysregulated pathway. RT-qPCR further confirmed aberrant expression of steroid metabolism-related genes, including upregulation of Akr1d1 and Ugt family genes.

Conclusion: Disruption of steroid hormone biosynthesis represents central molecular feature to AMA-TA. Integrated multi-omics analysis offers mechanistic insight and supports the development of biomarkers and therapeutic targets for AMA-TA.

Persistence of unrepaired DNA damage in oocytes is detrimental and may cause genetic aberrations, miscarriage, and infertility. RPA, a single-stranded DNA (ssDNA)-binding complex, is essential for various DNA-related processes. Here we report that RPA plays a novel role in DNA damage repair during postnatal oocyte development after meiotic recombination. To investigate the role of RPA during oogenesis, we inactivated RPA1 (replication protein A1), the largest subunit of the heterotrimeric RPA complex, specifically in oocytes using two germline-specific Cre drivers (Ddx4-Cre and Zp3-Cre). We find that depletion of RPA1 leads to the disassembly of the RPA complex, as evidenced by the absence of RPA2 and RPA3 in RPA1-deficient oocytes. Strikingly, severe DNA damage occurs in RPA1-deficient GV-stage oocytes. Loss of RPA in oocytes triggered the canonical DNA damage response mechanisms and pathways, such as activation of ATM, ATR, DNA-PK, and p53. In addition, the RPA deficiency causes chromosome misalignment at metaphase I and metaphase II stages of oocytes, which is consistent with altered transcript levels of genes involved in cytoskeleton organization in RPA1-deficient oocytes. Absence of the RPA complex in oocytes severely impairs folliculogenesis and leads to a significant reduction in oocyte number and female infertility. Our results demonstrate that RPA plays a previously unrecognized role in DNA damage repair during mammalian folliculogenesis.