Interactions between the endometrial epithelium and stroma are fundamental to development, physiology, and diseases of the uterus. Current endometrial organoid methods frequently rely on basement membrane extracts (BME), which support epithelial organoid growth in the absence of physiological mesenchyme/stroma. These systems typically generate cystic epithelial structures with apical-in polarity, limiting access to the luminal surface and preventing direct contact with the stroma. To overcome these limitations, we developed a protocol for generating BME-free assembloids composed of endometrial epithelial and stromal cells. This protocol details enzymatic isolation of primary cells from neonatal and adult mouse uteri and human endometrial biopsies, expansion of primary cells, and assembloid formation using a rotational culture system. Quality control benchmarks, including formation efficiency and cell type composition, were incorporated to ensure reproducibility. The resulting assembloids self-organize into structures consisting of a stroma core surrounded by a polarized epithelial monolayer and provide a tractable platform for investigating stromal-epithelial crosstalk during uterine development and endometrial disease.

Post-ovulatory supplementation of progesterone (P4) to cows has been associated with enhanced conceptus elongation. The aim of the present study was to investigate the impact of P4 supplementation on peri-implantation conceptus signaling including the secretion of interferon tau (IFNT), the maternal response to IFNT (mRNA abundance of interferon-stimulated genes [ISG; ISG15 and MX2]), and circulating pregnancy-associated glycoproteins (PAG). Two cohorts (A and B) of primiparous Angus beef cows were synchronized to estrus, artificially inseminated (d0), and assigned randomly to receive intravaginal P4 device (CIDR, 1.38 mg) from d3 to d12 or remain as untreated controls. Blood and cervical cytobrush were collected from d3 to d24 and a uterine cytobrush was collected on d24 from Cohort A to assess circulating concentrations of P4, PAG, mRNA abundance of ISG, and concentrations of IFNT, respectively. Blood and cytobrush (cervical and uterine) were collected from Cohort B on d18 to assess circulating concentrations of P4, mRNA abundance of ISG, and concentrations of IFNT, respectively. The mRNA abundance of ISG was assessed in luteal biopsies on d18 and d22. There was no effect of P4 supplementation on concentrations of IFNT in the uterus on d18. There was no effect of supplemental P4 on mRNA abundance or timing of ISG in endometrial, cervical, or luteal tissue. The earlier detection of PAG in circulation and lower uterine fluid concentrations of IFNT on d24 in P4-supplemented cows suggests advanced trophoblast differentiation and enhanced conceptus development beyond previously reported advances in conceptus elongation alone.

Environmental arsenic (As) exposure impairs spermatogenesis and testicular function, but the underlying mechanism remains unclear. In this study, 6-week-old male mice and rat primary Sertoli cells were utilized to explore the mechanism of arsenic-induced spermatogenic impairment. Serum testosterone levels were measured, testicular histology and ultrastructure were assessed, and the expression of tight junction (TJ) proteins, actin-related proteins, cytokines, and key components of the MAPK pathway were further investigated. Our results indicated that arsenic exposure reduced sperm quality, altered histological structure of the testis, and impaired TJs and actin cytoskeleton in the spermatogenic epithelia. Further analysis demonstrated that arsenic suppressed the expression of TJ proteins (CLAUDIN-11, ZO-1) in both testis tissues and primary Sertoli cells. The mRNA level and localization of the actin-related protein (ARP3) were also altered following arsenic exposure. Additionally, arsenic treatment markedly elevated testicular levels of the cytokines IFN-γ and TGF-β3. The gene expression and phosphorylation level of p38 Mapk were significantly increased in arsenic-exposed mice. These findings demonstrate that interference with Sertoli cell junctions is an important factor in arsenic-induced spermatogenic dysfunction. TJ barrier permeability and F-actin realignment, which might be mediated by TGF-β3/p38 MAPK pathway, collectively contribute to the disruption of cell junction structure.

Frankliniella occidentalis (western flower thrips, WFT) is a globally destructive pest causing major crop losses via oviposition, feeding, and plant virus transmission. Previous work has focused on its reproductive physiology and behavior, but comprehensive 3D structural knowledge remains limited. We used volume electron microscopy (vEM) to achieve nanoscale 3D visualization of the reproductive systems in both sexes, providing the first high-resolution, in situ reconstruction and establishing a new anatomical framework for insect reproductive physiology. Males exhibit extreme miniaturization, with reproductive organs occupying just 1.45% of body volume yet retaining full function within a compact abdomen. Females possess eight tightly interwoven ovarioles that maximize coelomic space for oocyte maturation, maintaining a slender form while housing large eggs-reflecting an evolutionary trade-off between reproductive efficiency and body size. Our reconstructions resolve a key ambiguity: the female accessory glands are a pair of fused, S-shaped structures joined by an internal duct opening near the ovipositor base. Quantitative analysis reveals the first complete germ cell census, with asymmetric ovary cell numbers suggesting an adaptive strategy for sustained fecundity. By integrating detailed morphology with systematic quantification, this study creates a foundation for research in functional morphology and for developing reproduction-targeted pest management. The structural insights enable precise strategies to disrupt thrips reproduction and reduce virus spread. Beyond advancing pest control innovation, the findings serve as a valuable educational and reference resource in insect morphology.

KIT signaling is a fundamental regulatory pathway that preserves cellular homeostasis and controls cell development and fate across a wide range of organs and cell types. Consistent with this pleiotropic role, mutations in c-KIT/Kit have been associated with a wide range of phenotypes, including sterility, piebaldism, nevus formation, mastocytosis, and multiple malignancies. The contribution of c-KIT/Kit to reproductive function has attracted sustained attention for several decades, underscoring its essential role in fertility and gonadal biology. KIT expression is observed in oocytes - localized to the oocyte membrane and the cytoplasm - as well as in theca cells and interstitial cells, suggesting a multifaceted role in follicular development. Notably, all Kit mutant models develop primary ovarian insufficiency (POI) with variable onset, characterized by endocrine dysfunction, impaired folliculogenesis, and eventual female infertility. These findings collectively establish KIT signaling as a critical regulator of ovarian integrity, as both gain- or loss-of-function mutations in Kit consistently recapitulate POI-associated phenotypes. However, despite substantial progress, the precise molecular mechanisms by which KIT signaling integrates these pathways to preserve primordial follicle survival and prevent POI remain incompletely understood. Here, we summarize current knowledge of KIT expression and the functional consequences of Kit mutations, with particular emphasis on oocytes across ovarian cell populations and in comparison to other organ systems in humans and mice. We further evaluate the physiological and pathological significance of ovarian KIT signaling in female fertility and highlight crucial knowledge gaps that must be addressed to fully elucidate its role in maintaining ovarian function.

Chemotherapy-induced premature ovarian failure (POF) represents a major challenge to female reproductive health, yet the potential regulatory role of vaginal microbiota in this process remains largely unexplored. Using a well-established model of chemotherapy-induced ovarian aging, we observed significant disruptions in vaginal microbial ecology characterized by depletion of Lactobacillus species and concomitant enrichment of pathogenic bacteria. Microbiota transplantation effectively reversed these dysbiosis patterns and restored ovarian function. Single-cell transcriptomic analysis revealed that microbial intervention promoted the recovery of granulosa and luteal cell populations while simultaneously suppressing inflammatory activation in ovarian stromal cells, demonstrating the vaginal microbiota's capacity to maintain follicular integrity. Further mechanistic insights showed that microbiota transplantation upregulated key antioxidant defense systems and ribosomal protein networks within ovarian cells, suggesting coordinated actions to mitigate oxidative stress and enhance cellular repair capacity, although the specific microbial metabolites mediating these effects require further elucidation. Our findings establish for the first time the existence of a functional vaginal microbiota-ovary axis and delineate its critical role in protecting against chemotherapy-induced ovarian damage. This work not only advances our fundamental understanding of microbial-endocrine crosstalk but also identifies concrete microbial targets for developing innovative strategies to preserve fertility in cancer patients.

Uterine adenomyosis is a widespread condition that impairs female fertility, with elevated serum levels of cancer antigen 125 (CA125) and prolactin (PRL) often observed in affected women. Mifepristone has been investigated as a potential therapeutic agent to improve pregnancy outcomes in uterine adenomyosis patients. This study aimed to examine the effects of mifepristone on serum CA125 and PRL levels in uterine adenomyosis mouse models. The Institute of Cancer Research (ICR) mice were treated with varying doses of mifepristone after uterine adenomyosis was induced. Immunohistochemistry was used to analyze estrogen receptor and CA125 expression in uterine tissues, while serum CA125 and PRL levels were measured by enzyme-linked immunosorbent assay (ELISA). The study found that mifepristone treatment significantly reduced serum CA125 and PRL levels, and the 20mg/kg/day dose was most effective in improving pregnancy outcomes. These findings suggest that mifepristone can reduce key biomarkers associated with uterine adenomyosis and improve fertility outcomes, offering a promising treatment option for women suffering from this condition.