MEHP exposure damages the basement membrane by suppressing the expression of its constituent proteins in peritubular myoid cells of the rat testis

Abstract The basement membrane (BM) is a crucial support structure of the seminiferous epithelium, mainly made up of collagens, fibronectin, laminins, and proteoglycans. Along with the collagen fiber network, the peritubular myoid cell (PTMC) forms the tunica propria, which acts as a protective barrier for the blood-testis barrier (BTB). Here, we examined how PTMCs respond to mono-2-ethylhexyl phthalate (MEHP) exposure by altering the transcriptomic and proteomic profiles of extracellular matrix proteins, which are essential structural components of the BM. Bulk RNA was used for transcriptomics, and protein lysates were prepared for proteomics from primary PTMC isolates. Interstitial fluid (IF) was utilized to assess the baseline microenvironment of the testis, and the in vivo relevance of the protein changes was confirmed through immunofluorescence of testis cross-sections. Gene Set Enrichment Analysis (GSEA) revealed a suppression of genes in the basement membrane-linked pathway after MEHP exposure. These findings are further supported by proteomics analysis of PTMC lysates. Notably, downregulation of key basement membrane components—such as collagen IV, fibronectin, laminin, and others—was observed alongside the classical complement activation pathway in the testis microenvironment, specifically in the interstitial fluid. The data indicate that MEHP exposure downregulated the expression of BM component proteins in PTMC, underscoring the interconnections among BM degradation, BTB disruption, spermatogenesis, and disorganization of seminiferous tubule architecture during MEHP-induced testicular injury.