Intrauterine exposure to glyphosate-based herbicide during pregnancy induces testicular impairment via activating ferroptosis in offspring

Glyphosate-based herbicide (GBH) is a broad-spectrum herbicide extensively used in agriculture, which poses increased residual risks in grains and food products, thereby endangering human health. However, the impact of maternal GBH exposure on fetal testicular development, as well as the protective effects of dietary betaine supplementation, remains inadequately understood. Hereby, pregnant sow models were established to elucidate the effects of GBH exposure at concentrations of 20 and 100 mg/kg and to investigate the protective mechanisms of betaine. The results showed that GBH and betaine had no significant differences on blood biochemical, colostrum composition and immunoglobulin. However, 100 mg/kg GBH treatment damaged the testicular morphology and barrier function, and disturbed testosterone synthesis in fetal testis. Subsequently, we found that high concentration GBH exposure triggered oxidative stress and inflammatory responses of testis, and inhibited the nuclear factor erythroid 2-related factor 2 (Nrf2)-mediated antioxidant signaling pathway, which further restrained mitochondrial fusion. Moreover, the accumulation of lipid peroxides and Fe2+ indicated that at a 100 mg/kg GBH treatment caused testicular ferroptosis. Conversely, dietary supplementation with betaine appears to inhibit ferroptosis by modulating the expression of ferritin light chain (FTL), acyl-coenzyme A (CoA) synthetase long-chain family member 4 (ACSL4), and ferritin, thereby promoting testosterone synthesis and enhancing testicular development to some extent. In conclusion, these findings enhance our understanding of the risks associated with fetal testicular exposure to GBH in pregnant women and elucidate the protective mechanisms of betaine against GBH poisoning.