(WHS-L1.04) Transcriptional Analysis of Fibroblasts in Diabetic Wound Healing
Thursday, May 16, 2024
10:30 AM – 11:30 AM East Coast USA Time
Background Diabetic wound healing poses a significant public health concern, impacting approximately 30 million individuals in the United States alone, with projections indicating a doubling by 2050. The cellular mechanisms that prevent proper wound healing are still not fully understood. Fibroblasts are crucial contributors to all stages of wound healing. Given their significance, this study is dedicated to a comprehensive exploration of how heterogeneous populations of fibroblasts contribute to diabetic wound healing using transcriptomic analysis and various different mouse models over time. Methods We created full-thickness excisional wounds on the dorsum of C57/BL6 (WT) mice, WT mice subjected to a high-fat (HF) diet to induce pathophysiologic diabetes, and leptin-receptor deficient (DB) mice with genetically induced diabetes. Mice were sacrificed at post-operative days (POD) 0, 2, 7 and 30. Tissues were processed into single-cell suspensions for single-cell RNA sequencing. Bioinformatic analyses were used to identify and further analyze fibroblasts (Col1a1+, Col1a2+, Col5a1+, and Vim+). Results The focused analysis of fibroblasts revealed nine distinct subpopulations defined by a unique gene expression profile, indicative of its functional characteristics, including fibrotic (F), keloid-initiating (KI), altered epithelial-like (AEL), and angiogenic (A) clusters. Notably, on POD2, WT fibroblasts demonstrated considerable enrichment in A (12%) and F (14%) clusters, while HF exhibited a deficiency ( <1%) in both. Contrarily, DB only displayed a slight growth in A (3%), and F (4%). Across the different conditions at POD2, KI cluster displayed zero growth. At POD7, WT increased expression in A (15%), and declined in F (10%). These cells also demonstrated a sharp growth in KI (20%). DB rose to 8% in A and skyrocketed in F and KI (26% and 35% respectively). On the other hand, HF displayed a marginal increase of 3% in A and 1% in F with a substantial increase in KI (10.5%). Interestingly, HF displayed a significant increase in the AEL subpopulation with 40% vs <0.1% in DB and WT. By POD30, A, KI and AEL subpopulations dropped back to <0.7%. F remained marginally expressed in DB and HF at both 2% and WT regressed to 0%. Conclusion This study discovered distinct temporal patterns of A, F, AEL, and KI subpopulations of fibroblasts within WT, HF, and DB mice. Interestingly, A and F subpopulations were significantly weakened in the diabetic groups at POD2. At POD7, unique HF AEL clusters may critically affect impaired diabetic healing. Proportionally, the number of fibroblasts decreased by up to 100-fold by POD 30 compared to POD 7, demonstrating a resolution of the wound healing process. Creating therapies to increase A and fibrotic F subpopulations could improve diabetic wound healing.