Reproduction最新文献

Profitability in the pork industry relies heavily on breeding herd productivity. However, traditional phenotypic and genetic methods are limited in their selection efficiency for some reproductive potential related traits. This leads to high culling rates due to poor reproductive performance and detriments to productivity and profits. This study investigated whether metabolomic profiling of urine, saliva, and serum applied to machine learning (ML) algorithms could predict female pig reproductive potential status. Urine, saliva, and serum samples from high reproductive potential (HRP; lifetime number of piglets born alive ≥ 13) or infertile (INF; nonpregnant after two consecutive rounds of artificial insemination) female pigs were analyzed using targeted liquid chromatography and mass spectrometry. Metabolomic profiles were applied to an ML pipeline including partial least squares discriminant analysis and recursive feature elimination complementary feature selection and six supervised ML algorithms. The urine-based logistic regression (F1 = 0.90 ± 0.03, Matthews correlation coefficient = 0.81 ± 0.11) and saliva-based logistic regression (F1 = 0.94 ± 0.04, Matthews correlation coefficient = 0.86 ± 0.10) classifiers achieved impressive predictive performance, demonstrating the potential to accurately measure female pig reproductive potential. Feature importance and univariate analysis highlighted key biomarkers, including specific carnitines, amino acids, and phospholipids, that distinguished the female pig reproductive metabolic phenotype. Overall, integrating urine, saliva, and serum metabolomic data with ML offers a promising approach to improve female pig reproductive potential prediction. This could assist breeding herd sow retention and culling protocols, improving production and economic outcomes. Future work should validate these classifiers for practical use by including additional samples from broader farming environments and assessing the feasibility of integrating this approach in commercial operations.

We report on the telomere length (TL) changes in metaphase chromosomes throughout preimplantation development of human embryos-a period of genome-wide epigenetic reprogramming. Using semiquantitative fluorescence in situ hybridization, we measured relative TLs in the metaphase chromosomes of 69 preimplantation embryos from the zygote up to and including the blastocyst stage. Relative TLs increased significantly from zygote to the 2-5-cell stage, remained almost unchanged at the stages of 2-5 and 6-12-cells and decreased by the blastocyst stage. Concurrently with relative TL decrease at the blastocyst stage, an increase in interindividual TL variability occurred. The zygote-inherited, but not newly synthesized chromatids maintained parent-specific telomeres (longer in paternal compared to maternal chromosomes) up to and including the 2-5-cell stage, with a follow-up TL equalization in 6-12-cell embryos. The extent of interchromatid TL asymmetry-a phenomenon potentially linked to telomere lengthening through recombination-was assessed by TL ratios between sister chromatids and showed similar patterns across all stages of preimplantation development. The longer telomere is presumably located in the highly hydroxymethylated sister chromatid of hemihydroxymethylated chromosomes, i.e., those having higher 5-hydroxymethylcytosine content in one sister chromatid than in the other due to global epigenetic reprogramming in early embryogenesis. To conclude, our study suggests that in human preimplantation development telomeres are reprogrammed in conjunction with genome-wide epigenetic reprogramming of an embryo. By the blastocyst stage, when epigenetic reprogramming comes to an end, parent-specific TLs are also completely reprogrammed and every embryo develops its own unique TL pattern.

Declining reproductive health and fertility are global public health issues affecting an estimated 15% of reproductive-aged couples worldwide. The reasons for declining fertility are complex. However, a male contribution is thought to occur in ∼50% of infertile couples. Deficits in sperm number and/or function are undeniably a major cause of infertility, but compelling evidence suggests that additional factors in the male ejaculate also play an influential and underappreciated role. In this review, we focus specifically on extracellular vesicles within human seminal plasma and explore their emerging roles in reproduction and fertility. These seminal extracellular vesicles (SEVs) are nano-sized membrane structures secreted by various cell lineages in virtually all regions of the male reproductive tract and exert key roles in intercellular communication. Consideration is given to the well-characterized effects of SEVs in supporting sperm as they transit through the female reproductive tract and their ability to modulate the immune environment within the female reproductive tract. Building on these important roles, we also detail the emerging links between dysregulated SEV production and male fertility status, and highlight the potential utility of leveraging these vesicles to improve fertility and reproductive outcomes in infertile couples. Altogether, this review highlights how expanding knowledge of SEVs provides a new perspective on the complexity of seminal fluid physiology and the underlying aetiology of male infertility.

Elements such as iron, copper, and zinc play essential roles in the mammalian oocyte, egg, and embryo; however, among these metals, zinc plays unique regulatory roles. Temporal fluctuations in zinc concentrations drive reproductive milestones such as meiotic resumption, egg activation, and initiation of the mitotic cell cycle. Roles for zinc in late preimplantation embryo development have not been well characterized. Using a quantitative element approach we report the inorganic profiles of mouse embryos progressing through the late blastocyst stage. We find that blastocysts, like oocytes and eggs, and distinct to somatic cells, maintain higher levels of zinc than copper and iron. All three of these essential metals are more abundant in the inner cell mass, which contains the population of pluripotent stem cells that gives rise to the fetus, relative to the trophectoderm which gives rise to the placenta and extraembryonic tissues. To test whether zinc abundance was associated with mitotic progress and cell-fate lineage, we perturbed zinc homeostasis during blastocyst formation by artificially raising intracellular zinc concentrations with zinc pyrithione. This treatment during the morula-to-blastocyst transition, when cell-fate lineages emerge, resulted in an elevation of zinc in the inner cell mass. This treatment did not impact cell number but did increase expression of the pluripotency and epiblast marker, Nanog. These results demonstrate that the inorganic profiles of the late preimplantation embryo retain elemental hallmarks of earlier developmental stages, and perturbation of zinc levels alters pluripotency gene expression in the blastocyst.

Embryo implantation requires transient modulation of epithelial integrity at the embryo- endometrium interface, yet the molecular mechanisms that govern epithelial remodelling remain incompletely understood. In our previous work, we demonstrated that the luminal epithelial cells undergo a partial epithelial-to-mesenchymal transition (pEMT) at the site of embryo implantation, regulated by the homeobox transcription factor HomeoboxA10 (HOXA10). Here, we identify NOD-like receptor family pyrin domain-containing 3 (NLRP3), a key inflammasome component, as a direct downstream effector of HOXA10 that is essential for luminal epithelial cell remodelling. CUT&RUN profiling revealed that HOXA10 binds to regulatory regions of several NLRP family members, including NLRP3, and loss of HOXA10 in vivo results in increased NLRP3 expression in the luminal epithelial cells. Single-cell RNA-seq and immunostaining confirmed that NLRP3 is specifically upregulated in luminal epithelial cells undergoing pEMT at the time of implantation. This was accompanied by co-localization of NLRP3 with apoptosis-associated speck-like protein containing a CARD (caspase activation and recruitment domain) and CASP1 in luminal epithelial cells, suggesting inflammasome activation at the time of implantation. Crucially, treatment with NLRP3 inhibitor (MCC950) on the day of implantation, impeded pEMT and blocked pyroptosis mediated by gasdermin D in the luminal epithelial cells, leading to the retention of luminal epithelium at the site of implantation and ultimately impairing embryo invasion. Our findings collectively show that HOXA10 preserves epithelial identity by inhibiting NLRP3, and its downregulation promotes NLRP3 mediated inflammasome production and pyroptosis, allowing for epithelial clearing for a successful implantation. This study demonstrates the dual function of NLRP3 in inducing epithelial plasticity and cell death at the embryo implantation site, highlighting a precisely regulated inflammatory mechanism.

Previous reports have indicated that placental trophoblast extracellular vesicles (EVs) possess unique properties that enable them significantly to inhibit the proliferation of ovarian cancer cells in vitro and slow ovarian tumour growth in an in vivo model, while EVs derived from monocytes did not. However, the mechanisms underlying the inhibitory effects of trophoblast EVs remain unclear. In this study, we characterized the microRNAs (miRNAs) uniquely present in placental trophoblast EVs but absent from THP-1 monocyte-derived EVs. Through bioinformatic analysis, we elucidated the potential involvement of these unique miRNAs in the negative regulation of proliferation pathways implicated in ovarian cancer. Functional assays demonstrated that placental trophoblast EVs inhibited ovarian cancer cell proliferation, and this effect was reversed upon blocking EV uptake, indicating the transfer of the contents of the EVs as a crucial mechanism modulating cancer cell viability. Using miRNA mimics, we also demonstrated that specific miRNAs from placental trophoblast EVs exhibited inhibitory effects on ovarian cancer cell proliferation, highlighting the potential of placental trophoblast EVs as therapeutic agents. These findings not only shed light on the molecular mechanisms underlying the therapeutic efficacy of placental trophoblast EVs but also provide valuable insights into the potential development of miRNA-based therapies for ovarian cancer, including the use of trophoblast EVs as a therapeutic for ovarian cancer.

Seminal plasma contains bioactive components capable of modulating the uterine environment. Although the boar ejaculate is fractionated, with each portion differing in composition, the cumulative effect of these fractions on the female reproductive tract remains poorly understood. This study evaluated how stepwise inclusion of ejaculate fractions affects uterine gene expression, cell proliferation, and immune response after artificial insemination. Semen doses were prepared using three combinations: FR1 (sperm-rich fraction), FR2 (FR1 + intermediate fraction), and FR3 (FR2 + post-sperm fraction). Each dose was used for intrauterine artificial insemination in sows (n = 5 per group), with a non-inseminated control group (n = 5). Uterine tissues (U) were collected 6 days post-estrus onset for transcriptomic and histological analysis. RNA-sequencing analysis identified the highest number of differentially expressed genes (DEGs; false discovery rate < 0.05 and |log2 FC|≥1) in U-FR2 group (n = 602), followed by U-FR3 (n = 465) and U-FR1 (n = 414), relative to the control. Most DEGs were down-regulated in inseminated sows, including immune-related genes such as VNN2. Gene ontology analysis revealed biological processes conserved across groups, including metabolism, immune modulation, and epithelial remodeling. Histological assessment showed reduced Ki-67 positivity and decreased CD3+ T-cell infiltration in all the inseminated groups (p < 0.05) compared to the control. Notably, uteri from U-FR2- and U-FR3-treated sows showed a faster transition toward a secretory phenotype (percentage of glandular tissue; p < 0.05). These findings demonstrate that the inclusion of different ejaculate fractions triggers distinct uterine transcriptomic and histological responses, highlighting the role of seminal plasma (SP) composition on the uterine environment and suggesting that fraction-specific SP components may influence early pregnancy outcomes.

Asthenozoospermia (AS), a prevalent contributor to male infertility, remains incompletely characterized at the metabolic level. This study aimed to define the energy metabolic profile of AS spermatozoa and identify key proteins associated with observed dysregulations. Sperm samples from 19 AS patients and 21 normozoospermic (NS) controls were analyzed. Aliquots from each sample were used to quantify energy-metabolic parameters [oxidative phosphorylation (OXPHOS)/glycolytic ATP production rates, electron-transport chain complex activities, reactive oxygen species (ROS) levels, mitochondrial membrane potential (MMP)] and perform proteomic sequencing. Differentially expressed proteins (DEPs) underwent functional enrichment analysis. Spearman correlation linked DEPs to energy-metabolic parameters to identify pathogenesis-associated candidates. AS spermatozoa demonstrated significantly reduced OXPHOS-derived ATP production, impaired electron-transport chain complex activities, decreased MMP, and elevated ROS levels compared to NS controls. Proteomic analysis identified 205 DEPs, with aerobic respiration as the top enriched pathway. Correlation analysis revealed significant associations between OXPHOS ATP/MMP parameters and six DEPs [Dickkopf (DKK)3, KAD9, TMCO1, DRC10, NDUAD, DCAF8]. DKK3 exhibited pronounced expression in human reproductive tissues and localized to the sperm mitochondrial sheath. Dkk3 knockout (KO) mice displayed phenotypes mirroring clinical AS, including reduced motility and mitochondrial dysfunction. Collectively, our integrated human and murine data demonstrate significant OXPHOS impairment with elevated ROS levels in asthenozoospermic sperm, phenotypes recapitulated in Dkk3-KO models. These findings support DKK3 deficiency as a contributor to AS pathogenesis through mitochondrial bioenergetic dysregulation. Further interventional studies remain essential to define mechanistic links between DKK3 loss and compromised sperm mitochondrial function and motility.

The regulation of endometrial-conceptus communication during early pregnancy and its association with pregnancy loss was investigated in beef cattle. Estrous cycles of heifers were synchronized, followed by either a sham procedure (n = 15) or embryo transfer (n = 71) on day 7 (day 1 = ovulation). Endometrial luminal epithelial cells were collected, and luteal function was assessed via Doppler ultrasonography on days 6, 15, and 20. Animals were categorized into five groups (n = 6-8/group): cycling (simulated embryo transfer), EL7-20, EL20-25, EL25-30 (embryo losses detected on days 20, 25, or 30, respectively), and pregnant (confirmed by day 30). Endometrial ISG15 and RSAD2 expression was upregulated in EL20-25, EL25-30, and pregnant groups on days 15 and 20, indicating conceptus-induced immune activation in heifers with maintained corpora lutea. IRF2 was downregulated on day 15 in the EL25-30 and pregnant group. On day 20, IFNγ, IL6, and IL10 were upregulated in EL20-25, EL25-30, and pregnant groups, reflecting immune modulation supportive of pregnancy, whereas IL1β and IL8 were increased only in EL20-25 and EL25-30 groups, indicating a pro-inflammatory state associated with pregnancy loss. TGFβ1 expression increased on day 20 in cycling and EL7-20 groups. TLR2 expression increased between days 6 and 20 in pregnant, EL20-25, and EL25-30 groups, while TLR4 was dysregulated across all groups except pregnant. These findings indicate that the conceptus stimulates early immune activation-characterized by the induction of interferon-stimulated genes, IL6, IL10, IFNγ, and TLR2-by day 15 in recipients with sustained luteal activity, independent of pregnancy outcomes. In contrast, dysregulated IL1β, IL8, and TLR4 expression is associated with early pregnancy failure.

The spindle apparatus is essential for both mitotic and meiotic cell division. In contrast to its mitotic counterpart, the meiotic spindle of mammalian oocytes lacks centrosomes and thus relies on acentrosomal microtubule organizing centers to assemble the spindle. In addition, the positioning of spindle in oocytes is mediated by the F-actin network, not the astral microtubules (MTs). Overall, the role of astral MTs in meiotic spindle positioning has been overlooked due to the lack of centrosomes. In this study, we present an optimized method for staining and super-resolution imaging of astral MTs during meiosis. Though lacking centrosomes, both interpolar and astral MTs were present in the spindle through meiosis. The growth of astral MTs was inhibited by CDK1 and actin-related protein 2/3. Premature extension of astral MTs at metaphase I impaired the positioning of meiotic spindle, resulting in symmetric division of oocytes after meiosis I. Collectively, these results provide novel insights into the regulation of astral MT growth to facilitate the positioning of meiotic spindle in oocytes.