Single-cell transcriptomics and a boundary wound model reveal temporal decoupling and microenvironmental heterogeneity in radiation-induced fibrosis.

BACKGROUND: Radiotherapy is a key treatment method for cancer, but high-dose treatment can lead to damage to normal tissues and chronic fibrosis. There is a lack of systematic temporal analysis of the dynamic progression from early damage signals to late extracellular matrix (ECM) deposition. METHODS AND MATERIALS: We reanalyzed publicly available scRNA-seq count matrices from GSE211713 to explore microenvironmental remodeling during fibrosis. Then, we established an in vivo local irradiation (12 Gy) mouse model with fully irradiated wounds and boundary-spanning wounds. For tissue repair and molecular phenotyping evaluations, we used tissue staining, IHC, qPCR, and WB. RESULTS: scRNA-seq revealed temporal decoupling between ECM-related transcription and myofibroblast function. Our mouse model showed spatial heterogeneity: although the fully irradiated wound exhibited typical inhibition of healing, the wound across the irradiation boundary showed a rapid, matrix-rich response within 14 days. TGF-β/SMAD-related signals, myofibroblast markers, and collagen deposition in the animal model were not strictly synchronous. Additionally, CellChat predicted SPP1-centered communication at the late fibrosis stage. CONCLUSIONS: Fibrosis after radiotherapy may be influenced not only by local damage but also by the boundary microenvironment. These findings support a shift toward ECM-dominant remodeling and provide a hypothesis-generating basis for targeted intervention.