A thin endometrium can lead to low clinical pregnancy rates, low live birth rates, high spontaneous abortion rates, and low birth weight. However, current methods of treating thin endometria do not achieve ideal results. This study explored the effect of Indian Hedgehog (IHH) on thin endometrium and its mechanism of action. A thin endometrial rat model was established by infusion of 95% ethanol. IHH was overexpressed in model rats using adeno-associated viruses. The endometrial thickness and number of glands and vessels were determined using H&E staining. Endometrial fibrosis was detected using Masson’s trichrome staining. Immunohistochemistry was performed to detect α-SMA, MUC-1, and CK19. After modeling, the rats were mated, and the number of gestational sacs was counted for fertility assessment. Western blotting was used to detect the angiogenesis markers vWF, PCNA, and vim and Hedgehog signaling-related proteins SMO, GLI1, and GLI3. IHH overexpression reduced ethanol-induced edema and bruising, repaired the appearance of damaged tissue, increased endometrial thickness, promoted glandular and vascular regeneration, and alleviated endometrial fibrosis. IHH overexpression inhibited the expression of fibroblast marker α-SMA while promoting the expression of vWF, PCNA, vim, CK19, and MUC-1. It also increased the number of gestational sacs and promoted the expression of SMO, GLI1, and GLI3. In conclusion, IHH ameliorates ethanol-induced thin endometrium and improves fertility by activating the Hedgehog signaling pathway.

The cytoplasmic poly(A)-binding protein (PABPC) plays a central role in the life of poly(A) mRNAs, including their stability, translation, and decay. In addition to the nearly ubiquitous PABPC1, two testis-specific PABPCs, PABPC2 and PABPC6, are present in rodents, while one specific PABPC, PABPC3, is found in primate testes. These three PABPC proteins are each encoded by intronless genes that may have diverged independently due to the retroposition of prototypical Pabpc1 or PABPC1. PABPC2 and PABPC6 are distinguished from PABPC1 in that they barely associate with translationally active polysomal mRNAs and are enriched in male germ cell-specific nuage, termed chromatoid bodies. Despite these unique characteristics, spermatogenesis and male fertility were not compromised in mutant mice lacking either PABPC2 or PABPC6, suggesting functional redundancy between the two proteins. Here, we produced double-mutant mice lacking both PABPC2 and PABPC6 and found that the simultaneous absence of these two proteins failed to affect testicular protein synthesis, spermatogenesis, or male fertility in vivo. These results suggest that the functions of PABPC2 and PABPC6 are redundant with those of other coexisting PABPC proteins, including PABPC1. We propose that testis-specific PABPC proteins emerged because of transcriptional promiscuity in the testis.

During the periovulatory period, local production of cortisol surges in the bovine cumulus-oocyte complex (COC), although its physiological significance is not well understood. As a potent anti-inflammatory agent, cortisol may protect the COC from inflammation caused by lipopolysaccharide (LPS), an endotoxin known to cause infertility in postpartum cows. This study examined the effect of cortisol, together with progesterone (P4), on LPS-challenged bovine oocyte maturation. COCs were aspirated from follicles 2–5 mm in diameter and subjected to in vitro maturation for 21 h with various combinations of LPS, cortisol, cortisone (a substrate for cortisol production), trilostane (a P4 synthesis inhibitor), and nomegestrol acetate (NA; a synthetic progestogen). LPS (0.001, 0.01, 0.1, 1 μg/ml) suppressed oocyte maturation in a dose-dependent manner, and this effect was reversed by concomitant treatment with cortisol (0.1 μM). COCs converted cortisone to cortisol, and the locally produced cortisol (approximately 0.01 μM) was capable of negating the suppressive effect of LPS (1 μg/ml) on oocyte maturation. Trilostane suppressed oocyte maturation by eliminating P4 production, indicating the crucial role of P4 in this process. LPS equally suppressed oocyte maturation, regardless of the presence or absence of P4 or the various doses of NA (0.001–1 μM). This suggests that P4 alone does not inhibit the action of LPS. However, in the absence of P4, cortisol could not suppress the LPS effect on oocyte maturation. Collectively, these findings suggest that the bovine COC can protect itself from the suppressive effects of LPS by producing cortisol, with P4 being essential for this function.

The enzymatic isolation of preantral follicles (PAFs) is considered the most efficient method for retrieving a large number of intact follicles, offering significant advantages in terms of yield and processing time. However, the low success rate of enzymatically isolated follicles in long-term culture raises concerns regarding their impact on oocyte quality and developmental potential. This study addresses a critical gap in understanding how enzymatic retrieval of PAFs affects the oocyte-granulosa cell connection and its relationship with high mortality and culture failure observed during in vitro growth (IVG). By systematically comparing crude collagenases (IA and IV) and purified collagenases (Liberase TM and DH) with a mechanical isolation protocol, we identified the optimal enzyme concentrations that maximize follicle yield while minimizing cellular damage. Our results reveal that the enzymatic retrieval of PAFs corresponds to the loss of transzonal projections (TZPs) post-isolation, as well as premature oocyte extrusion and follicle deformities during IVG. Our findings also highlight the differential apoptotic responses in oocytes and granulosa cells. Although these enzymes sustain follicle cell integrity, they compromise oocyte viability during isolation. Notably, crude collagenases impair oocyte growth during prolonged culture, whereas purified collagenases preserve the developmental potential of oocytes. This study also provides the first evidence that enzymatic isolation of PAFs adversely affects TZPs. Overall, our study highlights the importance of selecting an appropriate method, enzyme type, and concentration for preserving the integrity of oocytes, follicles, and their connections, thereby supporting successful in vitro culture. Additionally, our results suggest that mechanical protocols and high-purity enzymes are preferred for maintaining oocyte competence.

Male infertility contributes substantially to overall infertility, with temperature changes adversely affecting the sperm quality. During infertility treatments, the exposure of home-collected semen to extreme temperatures during transport deteriorates the semen parameters. This study investigates the effectiveness of thermal containers and insulation materials for preserving semen quality at low temperatures. Semen samples from 35 healthy male partners undergoing fertility treatments were analyzed. The samples were segregated in three groups and assessed: standard collection containers (Group A), thermal containers (Group B), and thermal containers with warming materials (Group C). Samples exposed to 4°C exhibited a notable decline in motility and forward motility over time, whereas groups B and C maintained these parameters better. Group C maintained the internal temperature at approximately 20°C for up to 90 min, reducing cold-induced deterioration. These findings demonstrate that cost-effective thermal retention methods can preserve semen quality during transport, and potentially improve the outcomes of intrauterine insemination and in vitro fertilization.

The Rec8 gene is specifically expressed in fetal and adult gonads. Although the importance of REC8 in gametogenesis is widely acknowledged, the mechanisms underlying its germ cell-specific expression remain unclear. In this study, we utilized the mouse Rec8 gene sequence to construct a 2577 bp sequence, which included intron 1 (180 bp), exon 1 (118 bp), and an upstream 2279 bp region. The dual-luciferase assay results showed significant differences in promoter activity between –650 bp and –385 bp and between –89 bp and –35 bp. This indicated that the core promoter region of the Rec8 gene may exist within these regions. Bisulfite sequencing PCR results showed that CpGs 10–19 were largely unmethylated in the testes but hypermethylated in other tissues. Interestingly, correlation analysis between CpG methylation status and Rec8 mRNA expression levels showed that methylation of CpGs 10 to 19 was negatively correlated with Rec8 mRNA expression levels (Pearson’s r = −0.991, P = 0.009). Furthermore, RNA-Seq data and bioinformatic analyses suggested that the specific expression of Rec8 may be linked to the presence of TATA-like sequences within its core promoter region. Overall, these findings indicate that Rec8 expression is regulated by the low methylation of CpG sites and the presence of TATA-like sequences in its core promoter.

An association has been reported between a lower pH in the uterus and an increased rate of implantation. How low pH regulates endometrial function is unclear. This study investigated the effect of low pH on the expression of leukemia inhibitory factor (LIF), which is crucial for implantation, in a human endometrial carcinoma cell line, rat endometrial stromal cells, and porcine endometrial cells. LIF mRNA expression was quantified by real-time PCR and protein expression was assessed using western blot analysis. LIF mRNA and protein expression increased at low pH in human endometrial carcinoma cells. Increased LIF mRNA expression was also detected at low pH in rat endometrial stromal and porcine endometrial cells, suggesting that low intrauterine pH may create favorable conditions for implantation and endometrial receptivity across species. The increase in LIF mRNA expression in the three cell types was attenuated by the addition of amiloride, indicating that low pH promotes the expression of LIF via amiloride-sensitive molecules in the endometrium.