Mitochondria-targeted antioxidant Mito-Q induces granulosa cell apoptosis and impairs human pre-antral follicle growth in vitro

Abstract

The development of in vitro systems capable of supporting early human folliculogenesis holds significant promise for fertility preservation, particularly for patients who cannot undergo ovarian tissue transplantation. Mitochondria-targeted antioxidants have shown beneficial effects in later stages of oocyte maturation, but their impact during early folliculogenesis remains unclear. This study aimed to investigate the effects of the mitochondrial-targeted antioxidant Mito-Q on early-stage human follicles. Using human ovarian tissue and isolated pre-antral follicles obtained from women undergoing fertility preservation, two complementary in vitro models were established. Ovarian cortex fragments were cultured for up to 14 days, and isolated pre-antral follicles were maintained in three-dimensional culture for 8 days. Following an initial comparison of the oxidised and reduced forms, the reduced form was selected for further experiments. Outcomes included follicle density and morphology, apoptosis, cytotoxicity, growth, survival, and gene expression. Mito-Q did not improve follicle survival or overall follicle density. Instead, increasing concentrations were associated with a significant increase in apoptosis in granulosa cells within ovarian tissue and reduced growth of isolated pre-antral follicles. These effects were accompanied by changes in the expression of mitochondrial stress-related genes, suggesting activation of cellular stress responses without functional benefit for follicle development. The findings indicate that mitochondrial antioxidant supplementation does not universally benefit early folliculogenesis and may be detrimental at higher concentrations. These results highlight the importance of stage-specific approaches to redox modulation and suggest that careful optimisation of antioxidant strategies is required to improve in vitro follicle development and fertility preservation outcomes.