Ovarian aging‐associated downregulation of GPX4 expression regulates ovarian follicular development by affecting granulosa cell functions and oocyte quality

In the physiological state, female fertility declines with age, as evidenced by a steady decline in oocyte quantity and quality. Aging of the first organ, the ovary, is accompanied by increased oxidative stress levels in the ovary, causing a decline in the ovarian reserve and follicular atresia. Ferroptosis is a novel mode of programmed cell death discovered in recent years and is involved in the onset and development of various diseases. To investigate whether ferroptosis regulates ovarian aging, we first examined granulosa cells from patients with a normal ovarian reserve, decreased ovarian reserve (DOR), and advanced age (Aged). GPX4, a key gene involved in ferroptosis, was identified. The marker of its activity, glutathione (GSH), was significantly downregulated in granulosa cells from the DOR and Aged groups. Transmission electron microscopy confirmed abnormal changes in mitochondrial morphology in granulosa cells from the DOR and Aged groups. In vitro, granulosa cell culture results showed that ferroptosis inducers inhibited cell growth by downregulating GPX4 expression. In contrast, ferroptosis inhibitors reversed the inhibitory effects of ferroptosis on granulosa cell growth by upregulating GPX4 expression. The results of mice in vivo experiments showed that the expression level of GPX4 was significantly decreased in the oocytes of aged mice and that Fer-1, an inhibitor of ferroptosis, reversed the decrease in the number of oocytes retrieved and the quality of oocytes in aged mice. Cyclophosphamide (CTX) was used to generate a mouse model of premature ovarian failure. The results showed that Fer-1 treatment significantly restored the inhibitory effect of CTX on GPX4 expression in the cumulus cells and partially reversed the adverse effects of CTX on the follicular reserve in the ovaries, the number of oocytes retrieved, and the quality of the oocytes in mice. The study findings suggest that ferroptosis is involved in regulating ovarian aging and that GPX4 is a key gene in regulating ovarian follicle development and ferroptosis and a potential key target for treating ovarian aging.