(WHS-L1.01) WHAT MAKES A WOUND BECOME CHRONIC: CELLULAR AND MOLECULAR PROCESSES THAT LEAD TO CHRONIC WOUND INITIATION
Thursday, May 16, 2024
10:30 AM – 11:30 AM East Coast USA Time
Chronic wounds (CW) develop because of defective regulation of one or more processes needed for successful tissue repair, but how these alterations affect CW initiation, is still not known. The purpose of this study is to understand cellular and molecular processes that contribute to initiation of wound chronicity. We used a diabetic mouse model of CWs, scRNAseq, and bioinformatic approaches. Skin tissue was collected from the wound during the first 3 days of wounding from non-chronic (NCWs) and CWs, with four samples/group. After tissue processing, single cell isolation, and RNA sequencing, a total of 102,737 cells were profiled. After quality control and filtering (e.g. doublet removal), 75,511 cells were subjected to principal component analysis and cell clustering. We identified 17 clusters that were annotated using marker gene expression for different skin cell types: fibroblasts, keratinocytes, vascular and lymphatic endothelial cells, and leukocytes. Comparison of the clusters between NCWs and CWs, showed that there were significant differences between the same type of cluster under the two conditions. We identified 2 types of fibroblasts: one more prevalent in NCWs, healing fibroblasts (H-Fib), and the other more prevalent in CWs, non-healing fibroblasts (NH-Fib). Comparison of these 2 types of fibroblasts revealed several altered pathways. In NH-Fib, the Wnt signaling (upregulated SFRPs), integrin signaling (downregulation of Lamc3 and Megf6) and fibroblast growth factor signaling (downregulated Ppp2r2c), were all impaired. These are all processes needed for proper healing. We identified 5 subtypes of keratinocytes and of these, 2 showed significant differences. The cluster identified as channel keratinocytes is more abundant in CWs. However, the gene expression profile showed that the antiproliferative and apoptosis-inducing genes, IGFBP3 and Ptpn14, which inhibit IGF and YAP signaling, were overexpressed in these keratinocytes. The other type of keratinocyte is less abundant in CWs and shows upregulation of apoptotic pathways (e.g. Fas) and downregulation of migratory pathways (e.g. Mylk) suggesting impaired re-epithelialization in CW. Noteworthy, all 5 subtypes of keratinocytes show downregulation of electron transport chain complex 1 in CWs (e.g. mt-Nd1, mt-Nd4). The vascular and lymphatic endothelial cells in CWs had downregulation of mitochondrial complex 1 and antioxidant response genes (e.g. mt-Rnr2). The downregulation of mitochondrial genes indicates that the cells in CWs are not producing sufficient levels of ATP for proper function, which can contribute to the lack of: granulation tissue formation, re-epithelialization, blood flow, and excess of exudate produced by CWs. In conclusion, our findings show for the first time that the path for chronicity is established very early after wounding providing opportunities for the development of treatments that can reverse the path from non-healing to healing after debridement.