The role of autophagy in follicular development and atresia in rat granulosa cells.

OBJECTIVE

To investigate the involvement of autophagy in folliculogenesis and its correlation with apoptosis.

DESIGN

Animal model-based study.

SETTING

University medical center.

ANIMAL(S): Twenty-one day-old female Sprague-Dawley rats.

INTERVENTION(S): Ovaries obtained from established immature rat models primed with pregnant mare serum gonadotropin (PMSG) were used for the induction of follicular development and atresia. Granulosa cells isolated from developing follicles were cultured in serum-free condition with or without follicle-stimulating hormone.

MAIN OUTCOME MEASURE(S): Microtubule-associated light-chain protein 3 (LC3) and autophagic vacuoles were used as autophagic markers, and cleaved caspase-3 was used as an apoptotic marker in ovaries and/or granulosa cells.

RESULT(S): The LC3 protein was expressed mainly in granulosa cells during all developmental stages. In granulosa cells isolated from PMSG-primed immature rat ovaries, LC3-II expression showed a similar expression pattern to cleaved caspase-3. In addition, granulosa cells of atretic follicles that showed intense cleaved caspase-3 staining also showed intense LC3 immunoreactivity. An in vitro culture experiment revealed that the levels of LC3-II and cleaved caspase-3 proteins were gonadotropin-dependent. The induction and the gonadotropin dependency of granulosa cell autophagy were confirmed by the observation of autophagic vacuoles under transmission electron microscopy.

CONCLUSION(S): These preliminary results suggest that autophagy is induced mainly in granulosa cells during folliculogenesis and shows good correlation with apoptosis.