Immp2l gene knockout induces granulosa cell senescence by activation of cGAS-STING pathway via TFAM-mediated mtDNA leakage.

Granulosa cell-produced inflammatory factors may be key contributors to ovarian dysfunction, and Immp2l deficiency accelerates ovarian aging via granulosa cell senescence; however, the role of inflammation in granulosa cell senescence is largely unknown. Therefore, in this study, cGAS-STING-mediated inflammation was explored in Immp2l deficiency-induced granulosa cell senescence. Immp2l deficiency led to senescence-associated secretory phenotype (SASP) and granulosa cell senescence. Immp2l knockout caused mitochondrial dysfunction and mitochondrial DNA (mtDNA) leakage into the cytoplasm. The cytoplasmic mtDNA was recognized by the DNA-sensing molecule cGAS-STING, which activates cGAS-STING and key downstream interferon-stimulated genes (ISGs) and then promotes the secretion of proinflammatory factors, leading to SASP in senescent granulosa cells. Interestingly, the mitochondrial inner membrane pore protein (Cyclophilin D40) CyPD40 and the outer membrane pore protein voltage-dependent-anion channel 1 (VDAC1) were markedly increased in senescent granulosa cells, accompanied by significantly increased expression of the mtDNA stability protein mitochondrial transcription factor A (TFAM). Downregulation of TFAM with siRNA in senescent granulosa cells improved mitochondrial function, significantly decreased mtDNA in the cytoplasm, inhibited the cGAS-STING pathway and markedly decreased CyPD40 and VDAC1 protein levels in TFAM-treated senescent granulosa cells. The SASP phenotype was also alleviated. In addition, senescent granulosa cells were treated with procyanidin B2 (PCB2), which has anti-inflammatory effects, and the TFAM-mediated mtDNA-cGAS-STING pathway was inhibited, accompanied by a markedly reduced SASP phenotype and granulosa cell senescence. In conclusion, Immp2l gene knockout induced granulosa cell senescence by activation of the cGAS-STING pathway via TFAM-mediated mtDNA leakage into the cytoplasm through the CyPD40 and the VDAC1.