Single-cell analysis reveals distinct functional heterogeneity of CD34⁺ cells in anagen wound and diabetic wound

Wound healing is a complex biological process involving multiple cell types with their critical functions. The diabetic wounds show delayed wound healing, while the anagen wounds display accelerated wound closure. However, the mechanisms underlying the effect of cellular heterogeneity on wound healing are still unclear. CD34⁺ cells exhibit high heterogeneity in wound skins and improve wound healing. Herein, we investigated the phenotypic and functional heterogeneity of CD34⁺ cells in normal, anagen, and diabetic wounds. We obtained CD34 lineage tracing mice, constructed distinct wound models, collected CD34⁺ cells from wound edges, and performed single-cell RNA sequencing. We identified 10 cell clusters and 6 cell types of CD34⁺ cells, including endothelial cells, fibroblasts, keratinocytes, neutrophils, macrophages, and T cells. 5 subclusters were defined as fibroblasts. The CD34⁺ fibroblasts C2 highly expressed papillary fibroblastic markers took up the largest proportion in anagen wounds and were associated with inflammation and extracellular matrix. Increased CD34⁺ endothelial cells, fibroblasts C4, and neutrophils as well as decreased fibroblasts C1 were discovered in diabetic wounds. We also filtered out differentially expressed genes (DEGs) of each cell cluster in anagen wounds and diabetic wounds. Functional enrichment analysis was performed on these DEGs to figure out the enriched pathways and items for each cell cluster. Pseudotime analysis of CD34⁺ fibroblasts was next carried out indicating fibroblast C4 mainly with low differentiation. Our results have important implications for understanding CD34⁺ cell type-specific roles in anagen and diabetic wounds, provide the possible mechanisms of wound healing from a new perspective, and uncover potential therapeutic approaches to treating wounds.