Assessment of Reproductive Risks of Bisphenol S in male BALB/c mice through functional histological and multi-omics investigations.

Bisphenol S (BPS) has increasingly replaced bisphenol A, yet the molecular mechanisms underlying its low-dose hazards on male fertility remain incompletely characterized. This study systematically investigates BPS-induced reproductive toxicity by exposing male BALB/c mice to environmentally relevant doses (0, 10, 100, and 1000 μg/kg/day) for 14 or 35 days, utilizing an integrated phenotypic-to-multi-omics workflow. Notably, phenotypic and functional impairments exhibited a distinct time-dependent pattern; while 14-day exposure induced no significant alterations, prolonged (35-day) BPS exposure at 100 μg/kg impaired overall sperm motility, reduced mean angular displacement (MAD), and induced testicular germ cell apoptosis without causing measurable systemic toxicity, revealing a selective gonadotoxic phenotype. Mechanistically, RNA sequencing of testicular tissue and untargeted metabolomic profiling of epididymal sperm uncovered a coordinated "gene-pathway-metabolite-phenotype" regulatory axis. BPS transcriptionally suppressed steroidogenic pathway components, an effect corroborated by a dose-dependent decline in intratesticular testosterone (T) across all treatment groups and a compensatory elevation of luteinizing hormone (LH) and follicle-stimulating hormone (FSH), with LH showing greater sensitivity at the lowest dose. Sperm metabolomics further identified a statistically specific upregulation of prostaglandin F2α (PGF2α) dimethyl amine within the arachidonic acid metabolism network, with all adjacent prostaglandin branches remaining unaltered. This perturbation was cross-validated by the congruent dysregulation of upstream phase-I and phase-II biotransformation transcripts, Cyp2e1 and Ugt1a1, identified as shared regulatory nodes across both omics platforms. Collectively, these findings provide mechanistic insight into how BPS impairs sperm motility via a targeted disruption of the steroidogenesis-prostaglandin signaling axis and offer cross-omics evidence supporting a re-evaluation of BPS safety margins as a bisphenol A substitute.