Ssc-miR-429 expression proliles and functions on inducing Leydig cells apoptosis.

Leydig cells (LCs) play an indispensable role in testosterone synthesis, and their dysfunction can result in male reproductive disorders. Previous transcriptome sequencing revealed differential expression of MicroRNA-429 (miR-429) in both Leydig stem cells (SLCs) and LCs, indicating its potential regulatory function in LCs. In this study, we examined the expression of miR-429 in seven pig tissues (heart, liver, spleen, lung, kidney, testis, epididymis, brain) and investigated its impact on the proliferation and apoptosis of testicular interstitial cells using various techniques such as CCK-8, EdU, TUNEL, Western blot, among others. The results demonstrated that miR-429 exhibited lower expression levels in the testis, particularly in the LCs of testicular tissue. Upon upregulation of miR-429, TM3 cell density significantly increased, while downregulation led to a slight elevation in cell density. Further research indicated that the observed phenotype was due to miR-429-induced cell apoptosis, independent of cell proliferation. Additionally, a dual-luciferase reporter system revealed no targeting relationship between miR-429 and the predicted target genes (BMI1 and SOX5). Previous reports confirm Bcl2 as a known target of miR-429, leading us to hypothesize that miR-429 diminishes LCs' anti-apoptotic capability by inhibiting Bcl2. In summary, our findings suggest that miR-429 may induce LC apoptosis, supporting its potential as a biomarker for male reproductive disorders linked to Leydig cell dysfunction.

Polymorphism of GTF2A1 gene is associated with litter size in sheep.

General transcription factor IIA subunit 1 (GTF2A1) is required for transcriptional activation from most promoters in eukaryotic transcription. Previous whole-genome association analyses articles have predicted the effect of this gene on lambing in sheep. In the study, a total of nine insertion/deletion (indel) variants named L1 to L9 in the gene were selected to detect in 550 Australian White sheep (AuW) adult ewes. The polymorphisms were detected in four loci (L1, L2, L3 and L8locus) and the polymorphism information content (PIC) values were 0.270, 0.375, 0.372 and 0.314. Further, our study revealed that L1, L2 and L3 loci of GTF2A1 gene were significantly related to the first parity litter size and the polymorphism of L8 was significantly correlated with litter size in the second parity. In details, for the first parity, individuals with the II genotype of the L1 locus had higher little size than that with the ID genotype, individuals with the ID and DD genotype of the L2 locus had higher little size than that with the II genotype, and individuals with the DD genotype of the L3 locus had higher little size than that with the II genotype. All four loci do not follow to Hardy-Weinberg equilibrium, and have no linkage between them. In conclusion, the polymorphisms of GTF2A1 were confirmed and analysis results demonstrated that there are some relevance between difference genotypes and litter size, and these findings may provide new insights for accelerating sheep molecular breeding through molecular marker-assisted selection (MAS).

Chlorpyrifos induces male infertility in pigs through ROS and PI3K-AKT pathway.

Chlorpyrifos (CPF) has been shown to have male reproductive toxicity in mice and rat. However, the association of CPF and male reproduction in pigs remains unknown. Therefore, this study attempts to investigate the damage of CPF on male reproduction in pigs and its potential molecular mechanisms. First, ST cells and porcine sperms were treated with CPF and then cell proliferation, apoptosis, motility of sperm, and oxidative stress levels were examined, respectively. Meanwhile, RNA sequencing was performed on ST cells before and after the treatment of CPF. The results of experiments in vitro showed that CPF had broad-spectrum toxic effects on ST cells and porcine sperms. The RNA-sequencing data and WB results indicated that CPF may regulate cell survival through the PI3K-AKT pathway. In conclusion, this study may lay the foundation for improving male fertility in pigs and provide theoretical information for human infertility.

Ssc-MiR-21-5p and Ssc-MiR-615 Regulates the Proliferation and Apoptosis of Leydig Cells by Targeting SOX5.

Leydig cells (LCs) are the predominant cells of androgen production, which plays key roles in spermatogenesis and maintaining male secondary sexual characteristics. Abnormal development of LCs affects androgen levels in vivo, affects fertility and may even lead to infertility. Little is known about the regulation mechanism on LCs' development and maturation in domestic animals, especially the regulation of non-coding RNAs. In this study, we continued to dig deeper in the previous RNA-seq data of porcine LCs from our group, combined with detecting the expression profiles in different tissues and different types of cells in the testis, to screen out candidate microRNAs (miRNAs) that may affect the regulation of LCs. A total of two miRNAs, ssc-miR-21-5p and ssc-miR-615 ("ssc" is omitted below), were finally determined. After overexpression and interference of miRNAs in vitro, the effects of candidate miRNAs on the proliferation and apoptosis of TM3 (mouse Leydig cell line) were explored. The results showed that miR-21-5p led to a decrease in TM3 cell density and p53 (apoptosis related protein) expression. Meanwhile, miR-21-5p decreased EdU positive cell numbers, but increased TUNEL positive cell numbers, suggesting miR-21-5p could inhibit proliferation and promote apoptosis. Conversely, miR-615 could increase TM3 cell density. Western blot and TUNEL assay indicated miR-615 inhibited apoptosis, but had no effect on proliferation. In addition, Sox5 was identified a potential target gene of these two miRNAs by Dual-Luciferase reporter system assay. Our findings about functions of miRNAs in TM3 and the mapping of miRNAs-target gene regulatory network would provide an important basis for the further elucidation of miRNAs in regulating pig LCs.

Role of IDO and TDO in Cancers and Related Diseases and the Therapeutic Implications.

Kynurenine (Kyn) pathway is a significant metabolic pathway of tryptophan (Trp). The metabolites of the Kyn pathway are closely correlated with numerous diseases. Two main enzymes, indoleamine-2,3-dioxygenase (IDO) and tryptophan-2,3-dioxygenase (TDO or TDO2), regulate the first and rate-limiting step of the Kyn pathway. These enzymes are directly or indirectly involved in various diseases, including inflammatory diseases, cancer, diabetes, and mental disorders. Presently, an increasing number of potential mechanisms have been revealed. In the present review, we depict the structure of IDO and TDO and explicate their functions in various diseases to facilitate a better understanding of them and to indicate new therapeutic plans to target them. Moreover, we summarize the inhibitors of IDO/TDO that are currently under development and their efficacy in the treatment of cancer and other diseases.