Fibroblast Mitochondrial Ca 2+ Overload Drives Skin Fibrosis via Mitochondrial DNA Leakage and Cyclic GMP ‐ AMP Synthase–Stimulator of Interferon Genes Activation in Systemic Sclerosis

Objective To explore the clinical relevance of mitochondrial DNA (mtDNA) in systemic sclerosis (SSc) and elucidate the mechanism by which mtDNA leakage drives fibroblast activation. Methods Plasma mtDNA was quantified by quantitative polymerase chain reaction in 50 patients with SSc and 20 healthy controls (HCs). Primary dermal fibroblasts from five patients with diffuse cutaneous SSc and five HCs (n = 5 per group) were analyzed for mitochondrial structure, function, and RNA sequencing profiling. The opening of the mitochondrial permeability transition pore (mPTP) and oligomerization of VDAC1 were examined with Ca 2+ modulators and inhibitors. Stimulator of interferon genes (STING) knockdown using small interfering RNA was performed for validation. The STING inhibitor H‐151 was evaluated in vitro and in a bleomycin‐induced skin fibrosis model (n = 6 mice per group). Results Plasma mtDNA levels were significantly higher in patients with SSc than in HCs, negatively correlating with forced vital capacity (r = −0.436, P < 0.01) and positively correlating with the modified Rodnan skin score (r = 0.807, P < 0.001), interleukin‐6 (r = 0.667, P < 0.001), and transforming growth factor β (r = 0.678, P < 0.001). SSc fibroblasts exhibited abnormal mitochondrial morphology and dysfunction. Increased MCU and VDAC1 levels promoted mitochondrial Ca 2+ overload, mPTP opening, VDAC1 oligomerization, and cytosolic mtDNA accumulation, which activated the cyclic GMP‐AMP synthase (cGAS)–STING pathway and triggered profibrotic responses. Pharmacologic blockade of mPTP or VDAC1 reduced cytosolic mtDNA, whereas H‐151 suppressed profibrotic markers in SSc fibroblasts and attenuated dermal thickening and collagen deposition in bleomycin‐treated mice. Conclusion The Ca 2+ –mtDNA–cGAS–STING axis drives fibroblast activation and skin fibrosis in SSc, representing a promising therapeutic target.