Involvement of IL-6/STAT3 Signaling in the Protective Effects of Melatonin Against Skin Fibrosis, Inflammation, and Oxidative Stress in Systemic Sclerosis

Systemic sclerosis (SSc) is a complex autoimmune disease characterized by immune and inflammatory dysregulation, microvascular injury, and progressive fibrosis. Emerging evidence suggests that oxidative stress contributes to disease progression. Melatonin (MT) possesses anti-inflammatory, antioxidant, and anti-fibrotic properties; however, its therapeutic effects and molecular mechanisms in SSc remain unclear. Primary dermal fibroblasts from patients with SSc and healthy controls were used to evaluate the effects of MT on fibrotic, inflammatory, and oxidative stress-related phenotypes. Network pharmacology and public single-cell RNA sequencing analyses were integrated to explore the molecular mechanisms underlying the protective effects of MT. IL-6 trans-signaling was reconstituted using IL-6 plus soluble IL-6 receptor (sIL-6R), and STAT3 was pharmacologically inhibited using Stattic to evaluate the involvement of IL-6/STAT3 signaling in the protective effects of MT. The therapeutic efficacy of MT was further validated in a bleomycin (BLM)-induced mouse model of skin fibrosis. MT concentration-dependently attenuated fibrotic phenotypes, inflammatory responses, and oxidative stress in primary dermal fibroblasts derived from patients with SSc. Network pharmacology identified 185 overlapping targets between MT and SSc, with IL-6 and STAT3 emerging as key nodes, while single-cell RNA sequencing revealed alterations in fibroblast subpopulation composition, functional states, and intercellular communication in SSc skin. Mechanistically, MT reduced IL-6 expression and inhibited STAT3 activation. Reconstitution of IL-6 trans-signaling with IL-6 plus sIL-6R partially restored STAT3 activation and TGF-β1/Smad3 signaling and partially reversed the protective effects of MT on fibrotic phenotypes, inflammatory responses, and oxidative stress, whereas pharmacological inhibition of STAT3 with Stattic partially attenuated these rescue effects. In vivo, MT reduced BLM-induced skin fibrosis, inflammatory responses, and oxidative stress, accompanied by decreased IL-6 expression and STAT3 activation. MT attenuates fibrotic, inflammatory, and oxidative stress-related changes in SSc patient-derived dermal fibroblasts and alleviates skin fibrosis in a BLM-induced mouse model. The effects of IL-6 plus sIL-6R reconstitution and Stattic-mediated inhibition support the involvement of IL-6/STAT3 signaling in the protective effects of MT. These findings provide experimental evidence supporting further investigation of MT as a potential therapeutic strategy for SSc-associated skin fibrosis.