Transcriptomics and proteomics association analysis demystify the molecular mechanisms underlying epididymal sperm maturation disorders in yaks with cryptorchidism.

Cryptorchidism is a prevalent reproductive disorder in male mammals. Yaks, being a unique species inhabiting high altitudes, exhibit a higher prevalence of cryptorchidism compared to cattle in lowland areas, leading to a significant constraint on their reproductive efficiency. This study conducted a comprehensive analysis of the pathological characteristics and molecular regulatory mechanisms of epididymal tissue in cryptorchidism yaks through the integration of histological, transcriptomic, and proteomic approaches. The pathological analysis showed that the epididymal wall of cryptorchid yak was significantly atrophic, the lumen was narrow and the sperm was less, accompanied by abnormal deposition of elastic fibers and collagen fibers, reticular fiber degradation, and other structural remodeling. Furthermore, electron microscopy observations indicated folded nuclear membranes in epididymal cells of cryptorchidism yaks, accompanied by chromatin condensation, marginalization, and mitochondrial swelling. Transcriptionomics screened out 287 differentially expressed genes (|log2FC|>1, FDR-adjusted P < 0.05). These DEGs were mainly enriched in GO entries such as extracellular matrix reconstruction, signal receptor binding, and steroid synthesis. In addition, KEGG analysis showed that Wnt, PI3K-Akt, tight junction and calcium ion signaling pathways were significantly downregulated in cryptordidymis epididymis. Proteomics identified 114 differential proteins, among which the expression of key Wnt pathway proteins (such as SYNPO and LMCD1) and tight junction proteins (TJP2 and ZO-1) were down-regulated, and lysosomal function-related proteins were significantly activated. Multiomic association analysis revealed that only a few genes were expressed consistently at mRNA and protein levels, indicating that post-translational modification plays an important role in cryptorchidism pathology. The qRT-PCR and Western blot (WB) experiments further confirmed that the differential genes were highly consistent with the expression trend and subcellular localization of the proteins. This study not only provides a potential target for the treatment of cryptorchidism in high altitude areas but also accumulates important data for understanding the regulation of epididymal function and the mechanism of male infertility.