The mechanisms of arsenic-induced ovotoxicity, ultrastructural alterations, and autophagic related paths: An enduring developmental study in folliculogenesis of mice

Arsenic (As) exerts a wide range of adverse effects on biological systems, including the reproductive organs in males and females. However, the mechanisms of As-induced reproductive toxicity are mostly obscure. Recently, we showed that autophagy is an essential route for As2O3-induced reprotoxicity through the hypothalamic-pituitary-gonadal-sperm (HPG-S) axis in pubertal and matured F1-male mice. However, the role of autophagy in As2O3- induced ovarian toxicity is mostly unknown. Hence, this study aimed to elucidate the role of oxidative stress, mitochondrial impairment, and autophagic processes in the ovary of As-exposed female mice. For this purpose, mature female mice were challenged with 0, low (0.2), medium (2), and high (20 ppm) As2O3 from 35-days before mating till weaning their pups, and the F1- females from weaning until maturity. Then, all the mice were sacrificed, and oxidative stress parameters, mitochondrial indices, electron microscopic evaluation of the ovaries, expression of autophagic-related genes and proteins, and autophagosome formation were assessed. It was shown that medium and high As2O3 doses were a potent inducer of oxidative stress, mitochondrial dysfunction, and autophagy in the ovary of F1-generation. A dose-dependent increment in the gene expression of PDK1, PI3K, TSC2, AMPK, ULK1, ATG13, Beclin1, ATG12, ATG5, LC3, P62, ATG3, ATG7, and p62, as well as protein expression of Beclin1, and LC3- I, II, was evident in the ovaries of the As-treated animals. Moreover, a dose-dependent decrease in the expression of mTOR and Bcl-2 genes, and mTOR protein was detected with increasing doses of As, suggesting that As treatment-induced autophagy. Along with a dose-dependent increase in the number of MDC-labeled autophagic vacuoles, transmission electron microscopy also confirmed more autophagosomes and injured mitochondria in medium and high As2O3 doses groups. As2O3 also negatively affected the mean body weight, litter size, organ coefficient, and stereological indices in female mice. Finally, in physiological conditions, arsenic trioxide (As2O3) leads to an increased level of autophagy in the oocyte when many oocytes were being lost. These findings indicated that an imbalance in the oxidant-antioxidant system, mitochondrial impairment, and the autophagic process, through inhibition of mTOR, dependent and independent pathways, and Bcl-2, as well as activation of AMPK/PI3K/Beclin1/LC3 routes, could play a pivotal role in As-induced reproductive toxicity through ovarian dysfunction in females.