The effect of TiO2 nanoparticles on antral follicles is dependent on the nanoparticle internalization rate

Titanium dioxide nanoparticles (TiO2 NPs) are among the most widely produced metallic nanoparticles due to commercial and industrial applications in products including food, cosmetics, paints, and plastics. TiO2 NPs are released into the environment posing health risks for humans and wildlife. Widespread uses have raised concerns about the potential toxicity of TiO2 NPs in reproduction. The ovary is an important endocrine organ responsible for sex steroid hormone production and folliculogenesis. Nanoparticles can reach the ovary, but limited information is available regarding NP toxicity and its effects on ovarian antral follicles. Thus, we tested the hypothesis that exposure to TiO2 NP affects sex hormone synthesis, oxidative stress, and antioxidant response in ovarian antral follicles in vitro. In addition, we characterized the NP internalization in the antral follicles over time to determine any association between NP internalization and effects on the antral follicle. Antral follicles were exposed to vehicle control or TiO2 NPs (5, 25, and 50 µg/mL) for 96 h. The lowest NP concentration (5 µg/mL) showed no internalization and no effects in antral follicles. The 25 µg/mL concentration had the highest internalization rate, leading to increased mRNA ratio of Bax to Bcl2. Interestingly, the highest concentration (50 µg/mL) showed lower internalization compared to the 25 µg/mL, with altered levels of steroidogenic involved genes and increased levels of progesterone and testosterone compared to control. In conclusion, these data suggest that TiO2 NP is internalized in antral follicles as the first step process in impairing follicle functions.