Melatonin attenuates Drp1-mediated excessive mitochondrial fission through upregulation of SIRT1 in granulosa cells in PCOS

Mitochondrial dysfunction in GCs has been implicated in PCOS pathogenesis. Dynamin-related protein 1 (Drp1)-mediated mitochondrial fission is critical for maintaining intact mitochondrial function. This study aims to investigate whether mitochondrial fission contributes to mitochondrial dysfunction in the GCs of individuals with PCOS and the molecular effects of melatonin on mitochondrial fission. Transmission electron microscopy (TEM) of human GCs showed that the mitochondria exhibited a condensed and small spherical morphology with a tendency toward fragmentation in PCOS patients. At the molecular level, GCs from patients with PCOS presented significant increases in both the p-Drp1(Ser616)/(Ser637) ratio and mitochondrial fission factor (MFF). In a dihydrotestosterone (DHT)-induced PCOS-like mouse model, excessive mitochondrial fission in GCs was similarly observed, characterized by fragmented mitochondrial morphology via TEM and increased expression of the mitochondrion-localized Drp1 (mito-Drp1) protein. Similarly, in vitro experiments demonstrated that DHT treatment increased the expression of mito-Drp1, the p-Drp1(Ser616)/(Ser637) ratio, and MFF in KGN cells. Melatonin treatment effectively reversed these abnormalities, restoring mitochondrial morphology, reducing fission markers (mito-Drp1, the p-Drp1 ratio, and MFF), decreasing mitochondrial reactive oxygen species, and enhancing mitochondrial membrane potential. Mechanistically, melatonin upregulated SIRT1, which restored the imbalance of Drp1 phosphorylation and blocked its MFF-dependent mitochondrial recruitment, thereby attenuating Drp1-mediated excessive mitochondrial fission. Our findings reveal a novel protective mechanism of melatonin in PCOS via the SIRT1-Drp1 signaling axis, offering a potential therapeutic target for PCOS management.