Therapy to Teratology: Chronic Paternal Antioxidant Supplementation Alters Offspring Placental Architecture and Craniofacial Morphogenesis in a Mouse Model

Oxidative stress plays a significant role in regulating the mammalian epigenome, with emerging evidence suggesting imbalances in the cellular redox state trigger stress-responsive epigenetic modifications that drive various human diseases. However, it remains unclear whether, like worms, epigenetic changes caused by redox imbalance or mitochondrial stress can move through the mammalian germline, potentially affecting the health of future generations. Antioxidant therapies are commonly used to reduce oxidative damage and are widely employed in cases of male infertility, where high-dose supplementation is often recommended to enhance sperm quality and overall measures of male reproductive health. Interestingly, in non-stressed, ostensibly healthy males, recent research suggests that antioxidants may have a negative influence on sperm epigenetic markers, indicating a potential epigenetic liability. However, whether male antioxidant treatment can induce paternal effects on offspring growth and development remains unknown. Here, we employed micro-CT imaging and geometric morphometrics to determine whether chronic antioxidant supplementation in healthy male mice affects placental growth and craniofacial development in their offspring. Adult C57BL/6J male mice were given a six-week preconception regimen of N-acetyl-L-cysteine (NAC; 400 mg/kg/day) and selenium (0.04 mg/kg/day), which continued throughout breeding with treatment-naïve females. Although we observed modest alterations to the histological patterning of the female placenta, placental weights and efficiency remained unchanged. In contrast, we observed significant changes in facial shape and symmetry in both male and female offspring, with female offspring exhibiting significant reductions in eye spacing and head area. These changes occurred without any macro changes in paternal metabolic health, indicating that antioxidant-induced shifts in redox balance may disrupt developmental programming in the male germline independent of changes in overall health. Our findings emphasize the need for caution when using antioxidants as preconception interventions and broadly suggest that modulation of the paternal redox axis may result in altered developmental programming and teratogenic effects.