OR14-05 Mechanistic Roles of Mitochondrial AKT in Granulosa Cells and Folliculogenesis - Implications on Ovarian Dysfunction in Metabolic Syndrome

Abstract Disclosure: H. Lo: None. S. Bhattacharya: None. K. Shaw: None. Y. Chen: None. P. Fueger: None. A. Ta: None. J. Wang: None. P.H. Wang: None. Insulin-resistant females of reproductive age face increased risks of ovarian dysfunction. Insulin induces the translocation of activated AKT into mitochondria. This study aims to investigate the role of mitochondrial AKT in ovarian granulosa cells (GCs) during folliculogenesis. Two novel transgenic mouse models were developed with Cre-LoxP system to selectively inhibit mitochondrial AKT signaling in GCs (ovdnAKT) or augment mitochondrial AKT (constitutively active mitochondrial AKT) in GCs (ovcaAKT). Results: Single-cell RNA sequencing (scRNA-seq) identified seven distinct GC clusters representing sequential stages of folliculogenesis, from progenitor cells through the preantral and antral phases, and ending in luteinization or atresia. In ovdnAKT mice, compared to controls, the number of antral GCs was significantly reduced, while luteinizing GC numbers increased. Specific gene markers and pathways were identified, which explain the transcriptional and development changes observed in ovdnAKT GCs. Histological analysis revealed a significant increase in the total number of follicles and preantral follicles in ovdnAKT ovary, indicating impaired maturation of preantral to antral phase, thus aligns with scRNA-seq results. Histology also found preantral follicles consisted of various GC types and were surrounded by luteinizing GCs. Anti-Müllerian hormone (AMH), a biomarker for preantral follicles, was markedly elevated in ovdnAKT mice compared to controls. The estrous cycles of ovdnAKT mice were irregular and prolonged (11.9 ± 3.7 days vs. 4.5 ± 0.5 days in controls; p < 0.001). To further explore the effect of GC mitochondrial AKT, mice were fed a high-fat/high-fructose diet (HFFD) or a normal chow diet (NCD) to study metabolic syndrome-associated ovarian dysfunction. In control mice, HFFD significantly reduced insulin-induced AKT translocation to mitochondria in GCs and prolonged the estrous cycle (HFFD: 7.8 ± 2.1 days; NCD: 4.7 ± 0.5 days; p = 0.0022). Histological analysis showed an increased number of preantral follicles in the control mice after HFFD. In ovcaAKT mice, which express active mitochondrial AKT in GCs, the duration of the estrous cycle was normalized under HFFD, and follicle distribution in the ovaries restored. Summary: Mitochondrial AKT in GCs plays a critical role in ovarian follicle recruitment and maturation by modulating GC transcription of key reproduction genes. Disruption of GC mitochondrial AKT signaling impeded the maturation of preantral to antral follicles and accelerated follicle luteinization. Activating mitochondrial AKT in GCs restored normal folliculogenesis and estrous cycling in the HFFD-induced metabolic syndrome. Impact: Impaired mitochondrial AKT signaling in GCs is a novel mechanistic link between insulin resistance and ovarian dysfunction. Presentation: Sunday, July 13, 2025