Molecular Reproduction and Development最新文献

Inflammatory damage in ovarian granulosa cells (GCs) is a key mechanism in polycystic ovary syndrome (PCOS), cytoplasmic polyadenylation element binding protein-1 (CPEB1) is important in inflammatory regulation, however, its role in PCOS is unclear. We aim to research the mechanism of CPEB1 in ovarian GCs in PCOS using dehydroepiandrosterone (DHEA)-induced PCOS rat models and testosterone-incubated GC models. The pathophysiology in PCOS rats was analyzed. Quantitative-realtime-PCR, TUNEL, immunohistochemistry, and Western blot were applied for quantification. Additionally, cell counting kit-8, flow cytometry, immunofluorescence, Western blot, and Monodansylcadaverine staining were performed. We found that PCOS rat models exhibited a disrupted estrus cycle, elevated serum levels of testosterone, luteinizing hormone (LH), and follicle-stimulating hormone (FSH), increased LH/FSH ratio, and heightened ovarian index. Furthermore, reduced corpus luteum and increased follicular cysts were observed in ovarian tissue. In ovarian tissue, autophagy and apoptosis were activated and CPEB1 was overexpressed. In vitro, CPEB1 overexpression inhibited cell viability and sirtuin-1 (SIRT1), activated tumor necrosis factor-α, and interleukin-6 levels, as well as apoptosis and autophagy; however, CPEB1 knockdown had the opposite effect. In conclusion, overexpression of CPEB1 activated autophagy and apoptosis of ovarian GCs in PCOS.

During male fetal development, testosterone plays an essential role in the differentiation and maturation of the male reproductive system. Deficient fetal testosterone production can result in variations of sex differentiation that may cause infertility and even increased tumor incidence later in life. Fetal Leydig cells in the fetal testis are the major androgen source in mammals. Although fetal and adult Leydig cells are similar in their functions, they are two distinct cell types, and therefore, the knowledge of adult Leydig cells cannot be directly applied to understanding fetal Leydig cells. This review summarizes our current knowledge of fetal Leydig cells regarding their cell biology, developmental biology, and androgen production regulation in rodents and human. Fetal Leydig cells are present in basement membrane-enclosed clusters in between testis cords. They originate from the mesonephros mesenchyme and the coelomic epithelium and start to differentiate upon receiving a Desert Hedgehog signal from Sertoli cells or being released from a NOTCH signal from endothelial cells. Mature fetal Leydig cells produce androgens. Human fetal Leydig cell steroidogenesis is LHCGR (Luteinizing Hormone Chronic Gonadotropin Receptor) dependent, while rodents are not, although other Gα_s-protein coupled receptors might be involved in rodent steroidogenesis regulation. Fetal steroidogenesis ceases after sex differentiation is completed, and some fetal Leydig cells dedifferentiate to serve as stem cells for adult testicular cell types. Significant gaps are acknowledged: (1) Why are adult and fetal Leydig cells different? (2) What are bona fide progenitor and fetal Leydig cell markers? (3) Which signaling pathways and transcription factors regulate fetal Leydig cell steroidogenesis? It is critical to discover answers to these questions so that we can understand vulnerable targets in fetal Leydig cells and the mechanisms for androgen production that when disrupted, leads to variations in sex differentiation that range from subtle to complete sex reversal.

The parameters of sperm apoptosis and capacitation during liquid storage at 17°C can indicate the quality of pig sperm and the potential development of early embryos. However, the effect of kojic acid (KA) on semen preservation and its mechanism has not been fully understood. In this study, we discovered that adding KA to the diluent improved the antioxidant capacity of sperm mitochondria, maintained the normal structure of sperm mitochondria, and reduced sperm apoptosis. Western blot analysis revealed that KA prevented the release of Cytochrome c from mitochondria to the cytoplasm, reduced the expression of pro-apoptosis proteins cleaved Caspase-3 and cleaved Caspase-9, and increased the expression of the antiapoptosis protein Bcl-XL. Furthermore, KA also enhanced the motility parameters, oxidative phosphorylation level, adenosine triphosphate level, and protein tyrosine phosphorylation of capacitated sperm, while preserving the acrosome integrity and plasma membrane integrity of capacitated sperm. In conclusion, this study offers new insights into the molecular mechanism of how KA inhibits porcine sperm apoptosis and improves capacitated sperm parameters. Additionally, it suggests that KA can serve as an alternative to antibiotics.

Extracellular vesicles (EVs) play an important role in the development and function of mammalian ovarian follicles. However, the mechanisms by which they are taken up by the follicular granulosa cells remain unclear. In addition, while oocytes play a pivotal role in follicular development, the possible interactions between oocyte-derived paracrine factors (ODPFs) and EV signals are unknown. Therefore, this study aimed to elucidate the mechanism of EV uptake and the effects of ODPFs on EV uptake by follicular somatic mural granulosa cells in mice. Fluorescence-labeled transferrin (TRF) and cholera toxin B (CTB), substrates for clathrin- and caveolae-mediated endocytosis, respectively, were taken up by mural granulosa cells in vitro. Their uptake was inhibited by Pitstop 2 and genistein, inhibitors of clathrin and caveolae pathways, respectively. Mural granulosa cells took up EVs, and this uptake was suppressed by Pitstop 2 and genistein. Moreover, ODPFs promoted the uptake of EVs and CTB, but not TRF, by mural granulosa cells. These results suggest that mural granulosa cells take up EVs through both clathrin- and caveolae-mediated endocytosis and that oocytes may promote caveolae-mediated endocytosis to facilitate the uptake of EVs.

Premature ovarian insufficiency (POI) patients experience a decline in ovarian function and a reduction in serum reproductive hormones, leading to a significant impact on the outcomes of assisted reproductive technology. Despite the absence of an effective clinical treatment to restore fertility in POI patients, recent research has indicated that cord blood plasma (CBP) derived from human umbilical cord blood (hUCB) may offer therapeutic benefits for various degenerative diseases. The primary aim of this study is to explore approaches for enhancing ovarian function and serum reproductive hormones through the administration of CBP in a murine model. Initially, hUCB was utilized to obtain CBP (CBP), which was subsequently analyzed for cytokine and growth factor profiles in comparison to adult blood plasma (ABP) by use of flow cytometry. Subsequently, POI mouse models were established through the induction of 4-vinylcyclohexene diepoxide, followed by the injection of CBP into the tail. At 7, 14, and 21 days posttreatment, mouse ovaries and blood were collected, and their estrus cycle, body weight, and ovarian weights were evaluated using precise electronic balance. Finally, ovarian morphology and follicle number were assessed through HE staining, while serum levels of anti-Müllerian hormone (AMH), estradiol (E2) and follicle-stimulating hormone (FSH) were determined by ELISA. Our study revealed that individuals with CBP exhibited significantly lower concentrations of proinflammatory cytokines, including IL-β (p < 0.01) and IL-2 (p < 0.05), while displaying elevated levels of anti-inflammatory cytokines and chemokines, such as IL-2, IL-4, IL-6, IL-8, IL-12P70, IL-17A, IP-10, interferon-γ, and tumor necrosis factor-α (p < 0.01). Furthermore, CBP demonstrated remarkably higher levels of growth factors, including transforming growth factor-β1, vascular endothelial growth factor, and insulin-like growth factor-1 (p < 0.01) than ABP. Notably, our investigation also revealed that CBP restored the content of serum reproductive hormones, such as AMH, E2, and FSH (p < 0.05), and increased the number of primordial and primary follicles (p < 0.01) and decreased the number of luteal and atretic follicles (p < 0.01) in vivo. Our findings suggested that CBP-secreted cytokines and growth factors could be restored POI ovarian function, enhanced serum reproductive hormones and rescued follicular development in vivo. These findings further support the potential of CBP as a promising strategy in clinical applications for POI related infertility.

Conventional intracytoplasmic sperm injection (ICSI) is not recommended for nonmale infertile patients to avoid fertilization failure (FF) in view of controversies regarding safety issues. Among the strategies that may help to promote the use of in vitro fertilization (IVF) for women with a high risk of FF, rescue ICSI (R-ICSI) has attracted attention. This study aimed to evaluate the efficacy of short-term insemination combined with early R-ICSI in the treatment of unexplained infertility. A total of 952 controlled ovarian stimulation (COS) cycles for unexplained infertility were divided into short-term IVF (short-term insemination without R-ICSI, n = 500), R-ICSI (short-term insemination with R-ICSI, n = 141), and ICSI (conventional ICSI, n = 311) groups. Patients underwent consecutive transfer cycles until live birth, or until all embryos from the first COS cycle were used. Laboratory data and clinical outcomes from short-term IVF, R-ICSI, and ICSI groups were compared. Short-term IVF and ICSI groups were superior to R-ICSI group in polyspermy rate, available embryo rate, and top-quality embryos rate. Short-term IVF, R-ICSI, and ICSI groups underwent 705 (500 fresh and 205 frozen-thawed cycles), 190 (141 fresh and 49 frozen-thawed cycles), and 445 (311 fresh and 134 frozen-thawed cycles) transfer cycles, resulting in 294, 76, and 190 live birth cycles, respectively. Comparison of pregnancy outcomes among these three groups demonstrated similar clinical pregnancy rates and live birth rates in fresh and frozen-thawed embryo transfer cycles. There was no significant difference in the cumulative live birth rate among these three groups. R-ICSI group showed similar neonatal outcomes compared with short-term IVF and ICSI groups, including the rates of low birth weight, fetal macrosomia, small for gestational age and large for gestational age. Short-term insemination combined with early R-ICSI achieved satisfactory pregnancy and neonatal outcomes, albeit with a high polyspermy rate, which was an effective alternative to avoid excessive use of ICSI in unexplained infertility.

Ant queens can maintain a large number of sperm cells for over a decade after mating at the beginning of their adult lives until they die. This sperm storage ability is prominent; however, the cellular mechanisms involved remain unclear. Sperm cells are maintained in the female sperm storage organ—the spermatheca—which supplies a suitable environment for sperm cells. To reveal the molecular basis of the long-term sperm storage mechanisms in ant queens, protein profiles enriched in the spermathecal fluid relative to the hemolymph were identified in Lasius japonicus using data-independent acquisition-based quantitative proteomics technology. Proteins related to the extracellular matrix, antioxidants, metabolic pathways, proteases, chaperones, and with uncharacterized functions were especially abundant with higher log ratio values in the spermathecal fluid relative to the hemolymph. These enriched proteins were shared with highly expressed genes previously detected by transcriptome analyses of the spermatheca in queens of Crematogaster osakensis that belong to a different subfamily than L. japonicus. It is likely that the ability for long-term sperm storage evolved early in the ant lineage. Therefore, the common proteins identified in these two ant species are possibly crucial for this ability.

Boar seminal plasma (SP) proteins were associated with differences on sperm resistance to cooling at 17°C. However, information about seminal plasma proteins in boars classified by capacity of semen preservation and in vivo fertility remains lacking. Thus, the objective was to evaluate the SP proteome in boars classified by capacity of semen preservation and putative biomarkers for fertility. The ejaculates from high-preservation (HP) showed higher progressive motility during all 5 days than the low-preservation (LP) boars. There was no difference for farrowing rate between ejaculates from LP (89.7%) and HP boars (88.4%). The LP boars presented lower total piglets born (14.0 ± 0.2) than HP (14.8 ± 0.2; p < 0.01). A total of 257 proteins were identified, where 184 were present in both classes of boar, and 41 and 32 were identified only in LP and HP boars, respectively. Nine proteins were differently expressed: five were more abundant in HP (SPMI, ZPBP1, FN1, HPX, and C3) and four in LP boars (B2M, COL1A1, NKX3-2, and MPZL1). The HP boars had an increased abundance of SP proteins related to sperm resistance and fecundation process which explains the better TPB. LP boars had a higher abundance of SP proteins associated with impaired spermatogenesis.

Cystic ovary disease (COD) is a common cause of subfertility in dairy cattle. Therefore, the aim of this study was to provide novel concepts for cyst classification and to investigate the effects of COD on tubal microarchitecture, oviductal metabolic function, and the formation of the sperm reservoir. Bovine Fallopian tubes affected by follicular cysts, follicular cysts with luteinization and luteal cysts were investigated by a variety of microscopic and histological techniques and compared to control cows in metestrus and diestrus. We defined three types of cysts involved in COD, each of which had a characteristic wall thickness, inner wall appearance and cellular pattern within the cyst aspirate. Regarding the Fallopian tube, each cyst type was associated with a characteristic morphology, specifically the microarchitecture of the folds in ampulla, epithelial cell ratios, and ciliated/secretory cell size and form. Furthermore, each cyst type showed different patterns of tubal glycoprotein and acidic mucopolysaccharide synthesis, which was highly variable as compared to the controls. Our studies are the first to characterize the effects of COD on the Fallopian tube, which promotes the establishment of novel, cyst-specific therapeutic concepts in cattle and helps gain a holistic view of the causes of subfertility in cows with COD.

This is the first work using gonads from undifferentiated, genetically-sexed Siberian sturgeon describing expression changes in genes related to steroid synthesis and female and male sex differentiation. One factor identified as relevant for ovarian differentiation was the gene coding for the enzyme Hsd17b1, which converts estrone into estradiol-17β. hsd17b1 was highly activated in female gonads at 2.5 months of age, around the onset of sex differentiation, preceding activation of two other genes involved in estrogen production (cyp19a1 and foxl2). hsd17b1 was also strongly repressed in males. Two known foxl2 paralogs are found in Siberian sturgeon—foxl2 and foxl2l—but only foxl2 appeared to be associated with ovarian differentiation. With regard to the male pathway, neither 11-oxygenated androgens nor classic male genes (amh, dmrt1, sox9, and dhh) were found to be involved in male sex differentiation, leaving open the question of which genes participate in early male gonad development in this ancient fish. Taken together, these results indicate an estrogen-dependence of female sex differentiation and 11-oxygenated androgen-independence of male sex differentiation.