Abstract
BACKGROUND: Premature ovarian insufficiency (POI) profoundly compromises female reproductive health through accelerated follicle depletion and endocrine disruption. Emerging evidence highlights the therapeutic potential of mesenchymal stem cell-derived exosomes (MSC-Exs), particularly when their function is enhanced by hypoxic preconditioning. In this study, the ability of hypoxia-preconditioned MSC-Exs (H-Exs) to ameliorate oxidative damage to granulosa cells (GCs) and restore ovarian function, was systematically evaluated, and a POI rat model was used to investigate the underlying mechanism. METHODS: CircRNAs specifically expressed in H-Exs were identified and validated. The ability of H-Exs and their corresponding circPTP4A2 to repair oxidative damage and restore mitochondrial function were evaluated by antioxidant enzyme assays, reactive oxygen species (ROS) assays, JC-1 staining, ATP level assays, oxygen consumption rate (OCR) measurements and TEM. The interaction between circPTP4A2 and YBX1 was analysed by molecular dynamics simulations, RIP, CHX assays, and MG132 assays, and the restorative effect of the circPTP4A2/YBX1 axis on ovarian function was verified. RESULTS: Our findings revealed that compared with normoxic MSC-Exs (N-Exs), H-Exs exerted superior protective effects, significantly attenuating oxidative stress and restoring mitochondrial bioenergetics in KGN cells. Mechanistically, circPTP4A2 was identified as a hypoxia-responsive cargo selectively enriched in H-Exs. This circular RNA stabilized Y-box binding protein 1 (YBX1) through direct interaction, increasing its antioxidative capacity and mitochondrial regulatory functions. Hypoxia-inducible factor 1-alpha (HIF-1α) was further shown to transcriptionally upregulate circPTP4A2 via direct binding to the promoter region of its host gene PTP4A2. CONCLUSION: These results establish the circPTP4A2/YBX1 axis as a critical mediator of the therapeutic efficacy of H-Exs for POI, providing both mechanistic insights and a translational framework for exosome-based regenerative strategies.