Integrated Analysis of Transcriptome and sRNA Sequencing Reveals Mmu-miR-503-5p Regulates the Aluminum Chloride Stress Response of GC-1spg Cells by Targeting Islr

Aluminum chloride (AlCl₃), a widely used inorganic polymeric coagulant in everyday products and industrial materials, has been associated with male reproductive toxicity, though its molecular mechanisms remain poorly understood. To investigate the complex molecular mechanisms underlying GC-1spg cells' responses to AlCl₃ exposure, transcriptome and small RNA (sRNA) sequencing analyses were performed. Transcriptome sequencing identified 1168 differentially expressed genes (DEGs), while sRNA sequencing detected 65 differentially expressed microRNAs (DEMs). An mRNA-miRNA regulatory network was established, and functional enrichment analysis showed that its target genes were significantly associated with multiple signaling pathways, particularly the p53 pathway. Further validation via Western blot and Hoechst 33342 staining assays confirmed that GC-1spg cells underwent apoptosis upon AlCl₃ exposure via the p53 signaling pathway. Among the identified DEMs, mmu-miR-503-5p was found to enhance GC-1spg cells' tolerance to AlCl₃-induced stress. Moreover, dual-luciferase reporter assays and RT-qPCR confirmed that mmu-miR-503-5p directly binds to the Islr gene, which plays a role in modulating GC-1spg cell tolerance to AlCl₃-induced stress. These findings provide critical insights into the molecular mechanisms governing GC-1spg cells' responses to AlCl₃ exposure.