Yaks, a valuable livestock species endemic to China's Tibetan plateau, have a low reproductive rate. Cryptorchidism is believed to be one of the leading causes of infertility in male yaks. In this study, we compared the morphology of the normal testis of the yak with that of the cryptorchidism, and found dysplasia of the seminiferous tubules, impaired tightness of the Sertoli cells, and a disruption of the integrity of the blood-testis barrier (BTB) in the cryptorchidism. Previous studies have shown that CAV1 significantly contributes to the regulation of cell tight junctions and spermatogenesis. Therefore, we hypothesize that CAV1 may play a regulatory role in tight junctions and BTB in Yaks Sertoli cells, thereby influencing the development of cryptorchidism. Additional analysis using immunofluorescence, qRT-PCR, and Western blotting confirmed that CAV1 expression is up-regulated in yak cryptorchidism. CAV1 over-expression plasmids and small RNA interference sequences were then transfected in vitro into yak Sertoli cells. It was furthermore found that CAV1 has a positive regulatory effect on tight junctions and BTB integrity, and that this regulatory effect is achieved through the FAK/ERK signaling pathway. Taken together, our findings, the first application of CAV1 to yak cryptorchidism, provide new insights into the molecular mechanisms of cell tight junctions and BTB. This paper suggests that CAV1 could be used as a potential therapeutic target for yak cryptorchidism and may provide insight for future investigations into the occurrence of cryptorchidism, the maintenance of a normal physiological environment for spermatogenesis and male reproductive physiology in the yak.

Pig cryptorchidism is a congenital anomaly, which requires surgical intervention. Pain assessment in pig farming has some limitations and consumers are increasingly concerned about suffering linked to on-farm procedures. The PGS seems to be a promising tool for pain assessment in piglets, although it has not been investigated whether this tool is applicable to growing pigs. This study was designed to determine whether the PGS could be useful to assess pain in growing pigs undergoing on-farm cryptorchidectomy. Ten mixed-breed cryptorchid pigs were pre-medicated with azaperone and ketamine. Anesthesia was induced and maintained using IV sodium based. Pigs were filmed pre- and 6 h post-surgery to evaluate their behavior (scan sampling every minute). Besides, 36 pictures of the face expressions (18 pre- and 18 6 h post-surgery) were scored with the PGS by three treatment-blind observers. The pre-surgery pig's activity ratio was 59%, while it was 2% 6 h post-surgery. While the PGS inter-observer reliability was excellent (Interclass Correlation Coefficient value of 0.87), the PGS score increased significantly in pigs 6 h post-surgery (Paired sample t-test, p = 0.02). The PGS proved to be a potentially effective method to assess pain associated with cryptorchidectomy. However, further validation studies are required to validate this tool for other potentially painful procedures.

Testes produce sperm, and investigations into gene expression in the testes will enhance the understanding of the roles of testicular genes in male reproduction. Cryptorchidism, the failure of one or both testes to descend into the scrotal sac, is a common congenital malformation in horses. The major clinical consequence of this abnormality is impaired fertility. The aim of this study was to analyze the expression patterns of testicular genes and to identify the differentially expressed genes (DEGs) in testes between cryptorchid and normal horses. In this study, the gene expression patterns in equine testes and the DEGs between mature descended testes (DTs) and undescended testes (UDTs) were identified by RNA-seq and validated by real-time qPCR. Our results provide comprehensive transcriptomic data on equine testes. The transcriptomic analysis revealed 11 affected genes that were downregulated in UDTs, possibly as a result of the higher temperature in the abdomen than in the scrotal sac. These 11 genes have previously been associated with male reproduction, and their downregulation might explain the impaired fertility of cryptorchid horses. Two homozygous missense mutations detected in horses with cryptorchidism were absent in normal horses and were listed as potential pathogenic mutations; these mutations should be verified in the future.

In the biological process of testicular spermatogenesis, the expression and interaction of many genes are regulated by microRNAs (miRNAs). However, comparisons of miRNA expression between descended testes (DTs) and undescended testes (UDTs) are rarely done in horses. In this study, we selected two UDTs (CKY2b and GU4b) from Chakouyi (CKY) and Guanzhong (GU) horses and eight DTs (GU1-3, CKY1, CKY3, CKY2a, GU4a, and GU5). Three groups were compared to evaluate expression patterns of testicular miRNA in stallion testes. Group 1 compared normal CKY horses and GU horses (CKY1 and CKY3 vs. GU1-3). Group 2 (CKY2a and GU4a (DTs) vs. CKY2b and GU4b (UDTs)) and group 3 (GU1-3, CKY1, CKY3 (DTs) vs. CKY2b and GU4b (UDTs)) compared the expression levels in unilateral retained testes to normal testes. The results show that 42 miRNAs (7 upregulated and 35 downregulated) had significantly different expression levels in both comparisons. The expression levels of eca-miR-545, eca-miR-9084, eca-miR-449a, eca-miR-9024, eca-miR-9121, eca-miR-8908e, eca-miR-136, eca-miR-329b, eca-miR-370, and eca-miR-181b were further confirmed by quantitative real-time PCR assay. The target genes of differentially expressed miRNAs in three comparisons were predicted, and the functions were annotated. The putative target genes of the 42 co-differentially expressed miRNAs were annotated to 15 functional terms, including metal ion binding, GTPase activator activity, zinc ion binding, intracellular, cytoplasm, and cancer pathways, and osteoclast differentiation. Our data indicate that the differentially expressed miRNAs in undescended testis suggests a potential role in male fertility and a relationship with cryptorchidism in horses. The discovery of miRNAs in stallion testes might contribute to a new direction in the search for biomarkers of stallion fertility.