Biology of Reproduction最新文献

Premature ovarian insufficiency (POI) impacts ~1-3.7% of women under the age of 40 globally and is characterised by an absence or complete loss of ovarian function. POI is clinically heterogenous in nature and researchers have identified >100 causative genes harbouring variants responsible for POI thus far. Genes identified to date include those associated with cell differentiation/development, mitochondrial maintenance, hormone receptors, transcription/translation factors, DNA repair/replication, and metabolic processes. Genes encoding cell components that facilitate these processes should therefore also be considered in POI gene candidature. The RNA exosome is a critical component in RNA processing, degradation, and biogenesis in eukaryotic cells. Catalytic activity of the RNA exosome is supplied by two subunits, DIS3 and EXOSC10. Dysregulation of RNA exosome function results in conditions known as exosomopathies that have a broad spectrum of phenotypic severity. RNA transcript regulation is essential in transcriptionally inactive maturing mammalian oocytes with its disruption negatively impacting meiosis and fertilization. Notably, oocyte depletion of Exosc10 significantly impacts the fertility of female mice. Herein we identified, via whole exome sequencing, the first instance of a human POI patient with an EXOSC10 homozygous missense variant. Using Drosophila melanogaster we modelled the impact of knockdown of the EXOSC10 ortholog, Rrp6, on both somatic and germline ovarian cells. We observed that Rrp6 is required in ovarian development in Drosophila. Due to the conserved role of EXOSC10 in fertility maintenance across species we contend that variants in EXOSC10 identified in POI patients may be causative.

Semen from mature, healthy bulls contains commensal microbes. Potential internal sources of seminal microbes have not been extensively evaluated. Objectives were to 1) assess whether the testes of the bull contain commensal microbes and to compare the composition of the putative testicular microbiome to that of semen and rumen fluid and 2) determine whether other organs of the bull reproductive system contain microbial populations. Here, we demonstrate that the testes of the bull contain a low biomass, yet diverse, microbiome. Interestingly, the microbial composition of rumen fluid, semen and testicular tissue were each dissimilar from one another, indicating that each source contains a unique microbiota. Only four core amplicon sequence variants, Acinetobacter, Enterobacteriaceae (E. coli), Jeotgalicoccus, and Kiritimatiellae WCHB1-41, were shared between semen and the testes. Along with the testes, microbial populations were also present within the penile and pelvic urethra, seminal vesicles and epididymis of mature bulls. Of these anatomic niches, the microbial populations within the penile and pelvic urethra were the most diverse and shared the greatest number of core taxa (n = 72). The microbiota of the seminal vesicles, epididymis and the testes were significantly dissimilar from each other. Only one core taxa, Mycoplasma, was shared between the testes and epididymis. Collectively, our results demonstrate that semen and organs of the bull reproductive tract contain unique microbial populations. Further research is necessary to determine whether the microbial composition of organs of the bull reproductive system, such as the testes and epididymis, influence sperm viability and bull fertility.

Stimulating follicle growth could have utility for fertility preservation before follicular culture or as part of ovarian stimulation protocol therapies for women with a limited ovarian reserve. Propranolol is a clinically approved beta-blocker. Beyond its beta-adrenergic receptors, recent investigations have suggested an alternative role in activating mTOR. Therefore, this study aimed to evaluate the efficacy of propranolol treatment on early follicle growth, both in vitro and in vivo. For in vitro studies, neonatal mouse ovaries were cultured for up to 3 days in the presence or absence of propranolol and then processed for histological analysis, quantitative PCR (qPCR), RNA in situ hybridization, and Western Blot (WB). For in vivo experiments, mice were treated with a low or a high dose of propranolol or no drug for 15 days. Then, analyses were performed, including body and ovarian weight measurements, histological analyses, WB, and qPCR. In vitro experiments demonstrated that treatment decreased primordial follicles and increased transitional, primary, and secondary follicles. Consistent with this, propranolol treatment resulted in the downregulation of Pten and an increased presence of Cpeb1, a novel biomarker of follicle activation, in primordial follicles. In our in vivo studies, propranolol promoted follicle activation, increased PI3K-p110 levels, and decreased Pten expression. In summary, propranolol increases the transition of primordial follicles to more advanced stages of development. These findings highlight propranolol as a potential drug to induce, in vivo and in vitro, early preantral follicle growth, potentially improving fertility preservation techniques and studies.

The common marmoset (Callithrix jacchus) is a promising animal model for preclinical biomedical research due to genetic, anatomic, and physiological similarity to humans. Assisted reproductive technologies can maximize the potential of captive research marmoset colonies, including preservation and propagation of genetically desirable or modified individuals. The literature on marmoset sperm cryopreservation is sparse, and limited assessments have been applied to the existing protocols that have yet to be fully optimized. In this study, we refined and standardized established cryopreservation components for common marmoset sperm, performed a comprehensive evaluation of frozen-thawed sperm quality, and identified accessible components for widespread adoption. While sperm were significantly less motile following cryopreservation compared to fresh sperm, no significant differences were detected across different cryostorage durations up to 6 months. Moreover, the average post-thaw recovery rates were 66% (total motility) and 36% (progressive motility), exceeding previously reported outcomes. Fresh and frozen-thawed sperm exhibited no significant differences in key structural parameters, including acrosome integrity and DNA fragmentation. The improved marmoset sperm cryopreservation protocol reported here will facilitate the sharing of genetically diverse and/or gene-edited sperm for research and colony management and provide a robust foundation for future studies aimed at enhancing the outcomes of frozen-thawed sperm in this and other NHP species.

Background and objective: Maternal obesity is known to adversely affect fetal development, with placental transcriptional dysregulation being one of the key mechanisms. However, the effect of a maternal high-fat diet (HFD) on dynamic placental gene expression, particularly in the context of obesity propensity, remains poorly understood. This study aimed to investigate the impact of a maternal HFD on time-dependent placental transcriptome alterations, with a focus on identifying key dysregulated pathways during mid-to-late gestation in rats.

Methods: Female Sprague-Dawley rats were fed either a high-fat diet (HFD) or control chow (CC) diet before and during pregnancy. HFD-treated rats were categorized into obese-prone (OP) and obese-resistant (OR) groups based on pre-pregnancy weight. Maternal and fetal characteristics, as well as fetal outcomes, were recorded at gestational days (GD) 14.5, 17.5, and 19.5. Plasma cytokine levels were also measured. RNA sequencing (RNA-seq) was used to compare the transcriptomes of the three groups at GD 14.5, GD 17.5, and GD 19.5.

Results: At GD 14.5, OP and OR groups showed significantly lower fetal body weight, placental weight, and efficiency compared to the CC group, with these measures increasing significantly by GD 17.5. Concurrently, RNA-seq time-series analysis revealed a significant dysregulation of the spliceosome pathway in the OR group and the parathyroid hormone synthesis pathway in the OP group during mid-to-late gestation. Specifically, 19 out of 229 annotated spliceosome genes were differentially expressed in the OR group. These transcriptomic findings were robustly validated by qPCR, which confirmed the upregulation of Sf3a1 and Sart1 in both the OR and OP groups at GD 14.5, while Sf3a2, Sf3b4, and Rbm22 were specifically elevated in the OP group.

Conclusion: Maternal high-fat diet disrupts placental transcriptome dynamics during mid-to-late gestation, particularly affecting spliceosome pathways, regardless of maternal obesity phenotype, contributing to placental dysfunction and adverse fetal outcomes. The observed time-dependent divergences highlight the need for phenotype- and gestational stage-specific interventions to mitigate developmental risks associated with maternal HFD.

The foundation for lifelong spermatogenesis depends on a highly coordinated program of prepubertal germline development, during which a precise balance between spermatogonial proliferation and differentiation is established to ensure sustained spermatogenesis. Any disruptions to this balance can impair germ cell maturation and overall fertility. However, factors critical in maintaining this balance remain incompletely understood. Our previous studies revealed that core-binding factor subunit-β (CBFβ) regulates both proliferation and differentiation during the onset of spermatogenesis. Canonically, CBFβ functions as a co-factor for the Runt-related transcription factor (RUNX) family by forming heterodimeric complexes that can act either as transcriptional activators or repressors. Here, we reveal interactions between CBFβ and RUNX proteins within the male germline and highlight distinct expression patterns of RUNX1 and RUNX3, particularly differential temporal expression during discrete cell cycle phases within spermatogonia. Moreover, Cleavage Under Targets & Release Using Nuclease (CUT&RUN) analyses revealed both overlapping and distinct genomic localization of RUNX1 and RUNX3. Surprisingly, knockdown studies determined that RUNX1 and RUNX3 act in opposition as either transcriptional activators or repressors within overlapping genomic targets. By contrast, genomic regions with differential RUNX1 or RUNX3 localization suggest distinct regulation of proliferation or differentiation, respectively. Furthermore, motif analysis revealed enrichment of disparate transcription factor motifs, including canonical regulators of the germline. Collectively, our findings suggest that CBFβ, RUNX1, and RUNX3 participate in a network to precisely coordinate proliferation and differentiation during prepubertal germline development, thus ensuring continuous spermatogenesis and male fertility.

Metabolism provides sperm with the energy needed to swim to and fertilize the oocyte. While mammalian sperm become motile during ejaculation and undergo maturation in the female genital tract, their energy demand increases. Investigations into the metabolism of sperm and the capacitation-induced increase in energy demand have been stymied by a lack of appropriate methodologies. Here, we present a detailed methodology to perform stable isotope labeling mass spectrometry in isolated mouse sperm, allowing to follow the fate of exogenous energy substrates through their metabolic pathways. As an example, mouse sperm are exposed to ubiquitously and positionally labelled 13C-glucose and the rate of accumulation of 13C in different metabolites is detected and analyzed. Using this assay in the presence of different exogenous energy substrates, with sperm from different species, genetically modified mouse lines, and/or pharmacological activators and/or inhibitors can provide important insight into the contribution of different metabolic enzymes and pathways to sperm energy homeostasis.

Pregnancy loss associated with excessive prostaglandin signaling and decidual injury remains a major clinical challenge, and few therapies directly stabilize the progesterone-dependent uterine environment. We tested whether allylestrenol (AL), an orally active progesterone receptor (PR) agonist, can protect against misoprostol (MSP)-induced pregnancy disruption in mice. In a GD7.5 MSP abortion model, AL co-treatment markedly reduced embryo resorptions and restored pregnancy success toward control levels. Histological examination of implantation-site decidua showed that MSP induced substantial epithelial and structural disruption, whereas AL largely preserved uterine architecture. Immunohistochemistry revealed that MSP downregulated PR and increased the inflammatory mediators COX-2 and IL-6, while shifting the BAX/BCL-2 balance toward a pro-apoptotic profile. AL reversed these molecular changes, restoring PR expression, attenuating COX-2 and IL-6 staining, and normalizing the BAX/BCL-2 ratio. Transcriptomic profiling supported these observations: RNA-seq demonstrated that MSP induced broad upregulation of inflammatory and apoptotic genes (including Il6, Il1b, Ccl4, and Bbc3) and downregulation of progesterone-responsive survival genes (Pgr, Stat6, Bcl2), whereas AL + MSP samples clustered closer to controls and showed partial restoration of these gene-expression patterns. Collectively, these data indicate that AL mitigates MSP-induced pregnancy loss by preserving PR signaling and limiting inflammatory and apoptotic responses in the decidua, supporting its potential as a progestin-based strategy for conditions in which pathological prostaglandin activity contributes to miscarriage.

Epigenetic dysregulation in the human endometrium contributes significantly to infertility, and especially to recurrent implantation failure (RIF). Impaired endometrial decidualization is a major cause of RIF, although the underlying epigenetic mechanisms remain unclear. We found that among the H3K4 methyltransferases, only the mRNA of SET Domain Containing 7 (SETD7) was upregulated in the human endometrium during decidualization and that SETD7 was indispensable for the transition of human endometrial stromal cells from proliferation to decidualization. SETD7 deficiency reduced H3K4me1 peaks at the transcription start site (TSS) center of decidual signature genes while increasing H3K4me1 enrichment downstream of the TSS in cell cycle genes. Integrated transcriptomic and epigenomic analyses revealed that Homeobox A10 (HOXA10) cooperated with SETD7 to facilitate H3K4me1 distribution at decidual signature genes. In silico prediction and molecular assays indicated that SETD7 directly interacted with HOXA10 with high confidence and methylated HOXA10, most likely at lysine (K)392 within the homeodomain, thereby enhancing HOXA10 binding transcriptional activity and protein stability. Consistently, HOXA10 knockdown alone markedly suppressed H3K4me1 enrichment at the TSS center of the decidual marker IGFBP1, resulting in reduced decidual protein levels within HESCs. In mid-secretory endometrium from a subset of RIF patients, SETD7 and HOXA10 expression was decreased, whereas the cell proliferation marker MKI67 was elevated. Collectively, these findings demonstrate that SETD7 actively directs histone methylation at specific genomic regions through HOXA10 methylation, and highlight the potential of targeted epigenetic therapy for RIF.

Over 25% of active NASA astronauts are women who will be exposed to low daily doses and dose rates of galactic cosmic rays (GCR) in space. We hypothesized that exposing mice to a preliminary simulated GCR mixed heavy ion beam composed of iron, silicon, and titanium ions induces follicle depletion and dose-dependent ovarian tumors. Female mice were exposed to 10, or 20 cGy each of Fe, Si, and Ti ions or sham-irradiation in quick succession within 15 minutes for total doses of 0, 30, or 60 cGy of the three beams. 16 months later, their ovaries were removed. Hyperplasia of the ovarian surface epithelium (OSE) was noted in 13%, 59%, and 22% of the 0, 30, and 60 cGy irradiated mice, respectively. The prevalence of mixed ovarian tumors was 0, 6, and 89%, respectively, in the 0, 30, and 60 cGy groups. Low numbers of Ki67 positive OSE and tumor cells supported a benign tumor phenotype. In a separate study, Si ion irradiation alone at 32 cGy did not induce ovarian tumors in mice; however, the mixed heavy ions at all doses and Si ion irradiation alone reduced the total number of healthy ovarian follicles. Mixed heavy ion exposure reduced lipid peroxidation, fibrosis, inflammation, and lipofuscin accumulation at 60 cGy compared to 0cGy, but elevated inflammation and lipofuscin accumulation at 30 cGy compared to 60 cGy. Preliminary simulated GCR exposure causes ovarian follicle death and tumorigenesis. This study provides insight into space-radiation induced ovarian damage and cancer risk in females.