Molecular mechanisms of DBDPE-induced spermatogenic disorder: A network toxicology, machine learning, and in vivo experimental validation

• Integrated network toxicology and ML to investigate DBDPE toxicity. • Identified five core genes linked to spermatogenic disorder. • DBDPE disrupted HIF-1 signaling and gap junction pathways. • Strong binding observed between DBDPE and key target proteins. • In vivo experiments confirmed DBDPE-induced downregulation of core genes. Decabromodiphenyl ethane (DBDPE), a widely used novel brominated flame retardant, has been increasingly detected across environmental matrices and human samples. Owing to its marked persistence and bioaccumulation potential, DBDPE was designated as a Substance of Very High Concern (SVHC) by the European Chemicals Agency in 2025. However, evidence regarding its potential reproductive hazards remains limited. Here, we systematically investigated the molecular mechanisms underlying DBDPE-induced spermatogenic disorder using an integrated approach that combines network toxicology, machine learning, and in vivo validation. Intersecting 564 DBDPE-associated targets with 1,743 NOA-related differentially expressed genes identified 33 overlapping candidates that may mediate DBDPE-induced spermatogenic disorder. Enrichment analyses implicated the HIF-1 signaling and gap junction pathways in DBDPE-induced spermatogenic disorder. Machine learning prioritized five core genes, including PRKACG, WDR5, AURKC, PFKP, and ASGR1, all of which were downregulated in NOA patients. Molecular docking demonstrated stable binding between DBDPE and these core targets. In vivo , DBDPE exposure disrupted testicular structure, reduced sperm concentration and motility, and decreased expression of the five core proteins, confirming the predicted mechanisms. Additionally, we proposed an adverse outcome pathway (AOP) framework describing key events leading to DBDPE-induced spermatogenic impairment. Collectively, our findings revealed the reproductive toxicity of DBDPE and provided a mechanistic foundation for developing strategies to safeguard male fertility.