Environmental factors accelerate epigenetic aging of sperm via mechanistic target of rapamycin/blood-testis barrier mechanism

OBJECTIVE: To identify a common molecular mechanism of sperm epigenome reprograming by environmental factors by testing whether mechanistic target of rapamycin (mTOR)-dependent and mTOR-independent disruption of the blood-testis barrier (BTB) by environmental factors (heat stress [HS] and cadmium [Cd] exposure, respectively) accelerate epigenetic aging of sperm in a mouse model. DESIGN: Using Infinium methylation array, we developed a murine sperm epigenetic clock model and used it to assess epigenetic age shifts in adult mice exposed to environmental factors for the duration of two spermatogenesis cycles. SUBJECTS: C57BL/6 mice. EXPOSURE: Short-term acute intermittent whole-body HS protocol was designed to mimic human HS during heat waves. The Cd exposure group was treated with 1 μL/g body weight of water solution of CdCl₂. MAIN OUTCOME MEASURES: The activation of mTOR complexes by both stressors was analyzed using enzyme-linked immunosorbent assay. Effects of HS, Cd, and aging on sperm deoxyribonucleic acid methylation were compared using bioinformatic tools. RESULTS: We demonstrate that both mTOR-dependent BTB disruption by HS and mTOR-independent BTB disruption by Cd exposure accelerate sperm epigenetic aging, resulting in similar changes to sperm deoxyribonucleic acid methylation patterns, including changes in methylation of genes involved in embryonic development and neurodevelopment. CONCLUSION: The mTOR/BTB mechanism is a novel pathway through which environmental and other stressors influence sperm epigenetic aging. This pathway is a potential therapeutic target for the mitigation of environmental effects on epigenetic programs in sperm.