Upregulation of mitoNEET disrupts granulosa cell proliferation, apoptosis and mitochondrial homeostasis : a possible pathogenesis of follicular dysplasia in PCOS.

BACKGROUND

Polycystic ovary syndrome (PCOS), a leading cause of anovulatory infertility, is characterized by arrested follicular development due to imbalanced proliferation and apoptosis of granulosa cells (GCs). The molecular mechanisms of this dysfunction remain unclear. This study explores the role of mitochondrial NEET protein 1 (mitoNEET) in PCOS pathogenesis.

METHODS

The expression levels of mitoNEET in GCs from PCOS patients and testosterone-treated KGN cells were analyzed using PCR and Western blot. RNA sequencing was used to probe the transcriptional changes following mitoNEET knockdown. Western blot, flow cytometry analysis, immunofluorescence imaging, and transmission electron microscopy were used to detect mitochondrial function, cell proliferation, and apoptosis in KGN cells. MitoNEET inhibitor (NL-1) was administered to a dehydroepiandrosterone (DHEA)-induced PCOS mouse model to investigate its potential therapeutic effects.

RESULTS

We demonstrate that mitoNEET protein and mRNA are significantly upregulated in GCs from PCOS patients, correlating with hyperandrogenism. RNA sequencing highlights its regulatory effects on mitochondrial homeostasis, cell cycle, and apoptosis. MitoNEET knockdown reversed testosterone-induced mitochondrial dysfunction, including cristae disruption, ATP depletion, mitochondrial membrane potential injurion, Mito-SOX accumulation, and aberrant dynamics (OPA1 downregulation/DRP1 upregulation), while restoring mitochondrial unfolded protein response (UPRmt) markers. Mechanistically, mitoNEET silencing activated the PI3K/AKT pathway, enhancing KGN cell proliferation (via PCNA, Cyclin B1, and CDK1) and suppressing apoptosis (Bax/Bcl-2 ratio). Pharmacological inhibition of mitoNEET with NL-1 alleviated hyperandrogenemia, restored estrous cyclicity, improved ovarian morphology, and promote ovulation in PCOS mice.

CONCLUSIONS

These findings establish mitoNEET as a pivotal mediator of mitochondrial-PI3K/AKT crosstalk in PCOS-related follicular arrest, highlighting its therapeutic potential for restoring GC function and ovarian homeostasis.