Endothelial IFI16 drives vasculopathy and profibrotic remodeling in systemic sclerosis by activating JAK/STAT signaling.

OBJECTIVE: Systemic sclerosis (SSc) is characterized by a prominent type I interferon (IFN-I) signature, yet the upstream drivers that link this signature to fibrosis remain unclear. This study aims to identify Interferon gamma-inducible protein 16 (IFI16) as an endothelial mediator that contributes to vasculopathy and fibrotic remodeling in SSc. METHODS: Machine-learning screening of public bulk RNA-seq was applied to identify fibrosis-associated IFN-I regulators. IFI16 expression pattern was validated by scRNA-seq, spatial transcriptomics and immunofluorescence. Functional and pathway analyses were performed in IFI16-overexpressing endothelial cells. RESULTS: A machine-learning-based analysis of bulk RNA-seq datasets demonstrated that IFI16 is significantly upregulated in SSc skin and ranks among the IFN-I-associated genes positively correlated with more severe skin fibrosis, shorter non-Raynaud disease duration, and elevated extracellular matrix deposition. Single-cell and spatial transcriptomic analyses revealed the enrichment of IFI16 in endothelial cells, and the co-localization of IFI16 with vasculature was validated by immunofluorescence. IFI16-high endothelial cells exhibited higher IFN activity and enhanced extracellular matrix remodeling. Stimulation with profibrotic or proinflammatory cytokines (IL-6, IL-1β, TGF-β and IFN-⍺) upregulated IFI16 expression in endothelial cells. Gain-of-function experiments showed that IFI16 overexpression impaired endothelial proliferation, migration, and tube formation, while driving the transition of endothelial cells towards mesenchymal cells through JAK-STAT signaling. Inhibition of JAK-STAT signaling reversed IFI16-induced endothelial-mesenchymal transition. CONCLUSION: Collectively, endothelial IFI16 coupled IFN-I signaling to vasculopathy and fibrotic remodeling via JAK-STAT activation, highlighting IFI16 as a promising therapeutic target in SSc vasculopathy and the subsequent fibrosis.