Exploring the Therapeutic Potential of Adipose-Derived Stem Cells in Attenuating Skin Fibrosis: Insights from a Bleomycin-Induced Murine Model

BACKGROUND: Scleroderma, characterized by excessive collagen deposition and dysregulated immune responses, leads to debilitating skin fibrosis. Adipose-derived stem cells (ADSCs) exhibit antifibrotic and immunomodulatory properties, but their dynamic interactions within fibrotic microenvironments remain underexplored. This study investigates the temporal effects of ADSCs on skin fibrosis and elucidates their molecular mechanisms. METHODS: A bleomycin (BLM)-induced skin fibrosis model was established in C57BL/6J mice. GFP-labeled ADSCs were subcutaneously injected into fibrotic lesions, with follow-up analyses at days 1, 7, 14, and 21 post-injection. Histopathological evaluation (H&E, Masson’s trichrome, α-SMA staining), qRT-PCR, flow cytometry, and RNA sequencing were performed to assess fibrosis progression, inflammatory cytokine levels, and ADSC behavior. In vitro co-culture experiments with human scleroderma-derived fibroblasts validated key pathways. RESULTS: ADSCs localized preferentially to hair follicles and perivascular regions, with survival declining by day 21. BLM-induced dermal thickening, collagen disorganization, and α-SMA expression were significantly reduced by ADSC treatment (p<0.05). Proinflammatory cytokines (TNF-α, IL-6, IL-1β) surged post-BLM induction but decreased sharply after ADSC administration. RNA sequencing revealed time-dependent transcriptomic shifts: immunomodulatory pathways (e.g., IL-6 production, VEGF signaling) dominated at day 7, while antifibrotic pathways (collagen matrix regulation, TGF-β/Smad inhibition) emerged by day 14. TSG-6, a critical antifibrotic mediator, was upregulated in ADSCs within fibrotic environments. Silencing TSG-6 in ADSCs abolished their antifibrotic effects, restoring COL1 and α-SMA expression in fibroblasts (p<0.05). In vivo, TSG-6-deficient ADSCs failed to suppress TGF-β/Smad2/3 activation, confirming its pivotal role. CONCLUSION: ADSCs exert dual-phase therapeutic effects: early immunomodulation (days 1–7) followed by sustained antifibrotic action (days 14–21), mediated partly via TSG-6-dependent TGF-β/Smad pathway inhibition. These findings highlight ADSCs as a promising cell-based therapy for fibrotic skin disorders, offering insights into optimizing timing and mechanisms for clinical translation.