Kisspeptin exacerbates androgen-induced follicular dysplasia by promoting Drp1 phosphorylation imbalance and mitochondrial excessive fission in granulosa cells of polycystic ovary syndrome.

BACKGROUND: An excess of androgens induces follicular dysplasia. Kisspeptin is a polypeptide hormone that is expressed in both the central nervous system and ovary, and it is correlated with follicle development. However, the mechanism underlying its role in androgen-induced follicular dysplasia of polycystic ovary syndrome (PCOS) is unclear. This study aimed to explore whether Kisspeptin can exacerbate androgen-induced follicular dysplasia by promoting Drp1 phosphorylation imbalance and excessive fission of mitochondria in granulosa cells (GCs) of PCOS. METHODS: Thirty PCOS patients and thirty control females who received assisted reproductive technology treatment were recruited. The specimens collected included discarded human follicular fluid (hFF) and GCs. In vivo and ex vivo studies were conducted using the PCOS rat models, two ovarian GC lines, and GCs collected from PCOS and control subjects. The PCOS rat models were established by subcutaneous injection of Dehydroandrosterone(DHEA) daily with a high-fat diet. We used enzyme-linked immunosorbent assays to detect Kisspeptin expression and light and transmission electron microscopy to assess ovarian tissue morphology and mitochondria. Additionally, we tested the cell activity, reactive oxygen species (ROS), membrane potential and the content of ATP in GCs. The expression of mitochondrial dynamics-related proteins and mRNA was analyzed using western blotting and RT-qPCR. The colocalization of the Drp1, p-Drp1 (Ser616), and p-Drp1 (Ser637) with the mitochondria was further detected by immunofluorescence staining. RESULTS: It was found that the expression of Kisspeptin in the hFF of patients with PCOS was significantly increased and negatively correlated with the number of high-quality follicles. The expression of mitochondrial fission proteins Fis1, MFF, and Drp1 increase was observed in ovarian GCs of PCOS patients. In vitro experiments showed that ROS level increased, mitochondrial membrane potential decreased, and intracellular ATP content decreased significantly in GCs after Kisspeptin stimulation. Compared with the control group, mitochondrial cristae of GCs were thickened and swollen and Fis1, MFF increased after Kisspeptin stimulation. The p-Drp1 (Ser616) expression increased, and p-Drp1 (Ser637) expression reduced significantly. Drp1, p-Drp1 (Ser616), and mitochondria colocalization were significant. Serum and ovarian levels of Kisspeptin in PCOS rats were significantly increased, which were positively correlated with the number of preantral follicles and negatively correlated with the thickness of the GCs layer. Additionally, the Kisspeptin group exhibited a significantly damaged mitochondrial structure, and Fis1, MFF expression was higher, accompanied by pDrp1 (Ser616) overexpression and a higher p-Drp1 (Ser616)/p-Drp1 (Ser637) ratio. CONCLUSION: Kisspeptin exacerbates androgen-induced Drp1 phosphorylation imbalance, driving mitochondrial fission in granulosa cells. Excessive fission impairs mitochondrial function, leading to apoptosis and disrupted follicular development. Thus, Drp1 may be considered a potential target for the treatment of PCOS.