Time‐Controlled Refrigerated Stem Cell Therapy Mitigates Scleroderma Fibrosis via Modulation of Mitochondrial Autophagy and Gut Metabolism

Scleroderma is a chronic autoimmune disease characterized by progressive fibrosis, associated with high morbidity and mortality, limited therapeutic efficacy, and significant systemic side effects. Therefore, there is an urgent need to develop novel treatment strategies with improved tissue targeting and safety profiles. In this study, a refrigerated-treated mesenchymal stem cell (RT-MSCs) system was established by culturing human umbilical cord-derived MSCs under controlled low-temperature conditions. The antifibrotic effects of RT-MSCs were evaluated through both in vivo and in vitro experiments, together with an assessment of their regulatory role in mitochondrial autophagy. Their lesion-targeting capacity was also investigated. Furthermore, the effects of RT-MSCs on gut microbiota composition and metabolic pathways in model mice were comprehensively analyzed using 16S rRNA sequencing and intestinal content metabolomics, and the safety of RT-MSCs was systematically evaluated. The results demonstrated that RT-MSCs effectively attenuated fibrosis progression by modulating mitochondrial autophagy. Within 30 min after administration, RT-MSCs accumulated at lesion sites and persisted for up to 7 days. RT-MSCs significantly improved intestinal microbiota dysbiosis in scleroderma mice and regulated the expression of associated intestinal metabolites. In summary, as an optimized stem cell-based therapeutic strategy, RT-MSCs offer new insights and potential avenues for treating scleroderma.