Shandong Cancer Hospital and Institute Jinan, Shandong
H. Yu1, Y. Ma1, Y. Xu1, J. Yu2, D. Chen2, and Z. Zhang1; 1Shandong Cancer Hospital and Institute, Jinan, Shandong, China, 2Shandong Cancer Hospital and Institute, Shandong First Medical University and Shandong Academy of Medical Sciences, Jinan, Shandong, China
Purpose/Objective(s): To identify the critical pathogenic cellular subpopulations and their interactions within Radiation-induced skin injury (RISI), and screen key targets for alleviate the complications linked to RISI. Materials/
Methods: 18 mouse skin samples (C57/BL6, female, 6 weeks old) were included in this study. Initially, mice were subjected to a single fraction (20 Gy) of radiation therapy targeted at the right dorsal region, specifically above the right forelimb using Small Animal Radiation Research Platform (SARRP, from a medical technology company). Subsequently, integrated single-cell RNA sequencing (scRNA-seq) was conducted to analyze 71,412 cells, encompassing 10 major cell types and 32 subclusters from mouse skin samples collected at 7 and 30 days post-radiation exposure, as well as samples without radiation therapy. The combined gene expression matrix was converted into a Seurat object using the R package Seurat, followed by downstream analysis. Various techniques such as cell-cycle analysis, pseudotime analysis, RNA velocity analysis, Gene Ontology (GO) enrichment analysis, cellchat analysis, single sample gene set enrichment analysis (ssGSEA), and analysis of ligand-receptor interactions were utilized to characterize and uncover the distinct features of each subcluster. Additionally, reverse transcription polymerase chain reaction (RT-PCR), multiplex immunofluorescent staining, hematoxylin-eosin staining, KO (Knockout) mice and datasets from public databases were employed to validate our findings. Results: We discovered that cycling keratinocytes (KCs) with BMP signaling pathway enriched could activate the Wnt pathway, as well as the SMAD, driving the wound healing and fibrosis process in RISI. In addition, we identified terminally differentiated secretory-papillary fibroblasts (Fibs) capable of attracting immune cells in RISI. Lymphatic endothelial cells (ECs) with pro-inflammatory properties played an active role in the pathogenesis of RISI by facilitating leukocyte migration. Our analysis also highlighted enhanced ligand-receptor interactions, notably between chemokines such as Cxcl10, Ccl2, and Ackr1, across subclusters of inflammatory KCs, Fibs, ECs, and immune cells, underscoring their pivotal role in leukocyte recruitment in RISI. Conclusion: Cycling KCs, terminally differentiated secretory-papillary Fibs and lymphatic ECs represent critical roles in RISI progression. Targeting the interactions of these subclusters with immune cells might help improve the degree of RISI.
