Analysis of miRNA expression profiles during spermatogenesis of plateau zokor (Eospalax baileyi) under androgen antagonism treatment

Abstract

Seasonal reproduction in animals is characterized by programmed activation, dormancy of reproductive activities, and precise regulation of hormone levels. Androgens secreted by Leydig cells are essential for testicular development and spermatogenesis. To explore the mechanism of androgen in spermatogenesis in seasonal breeding animals, this study used the plateau zokor (Eospalax baileyi) as a model and, during the breeding season, treated it with the androgen receptor antagonist, flutamide, to analyze its effect on the microRNA (miRNA) expression profile in testicular tissue. Flutamide treatment resulted in marked changes in the expression of 91 miRNAs in the testes (28 upregulated and 63 downregulated), including 44 known and 47 newly predicted miRNAs. Functional enrichment analysis of the target genes of differentially expressed miRNAs demonstrated significantly enriched cell adhesion, hypoxia response, apoptosis process, inflammatory response， lipid metabolism (Gene Ontology analysis, P < 0.05) and ferroptosis, fatty acid metabolism and other signaling pathways (Kyoto Encyclopedia of Genes and Genomes analysis, P < 0.05). Based on this, an interaction network of miRNA–mRNA in the spermatogenesis and ferroptosis pathways was constructed. Dual-luciferase reporter gene assay confirmed that ACSL4 was a direct target gene of miR-486-y. The expression of miR-486-y was inhibited by an antagomir, which verified its involvement in the regulation of spermatogenesis in the plateau zokor during reproduction. Androgens regulate spermatogenesis in plateau zokors by inhibiting specific miRNAs (such as miR-486-y). This study provides molecular evidence and new perspectives on the regulatory network of seasonal reproduction in animals.