An advanced ADSC therapy for keloid prevention using identification of functional subgroups by single cell transcriptomic analysis

Abstract

Abstract Background Keloids are an excessive growth of skin fibres that occur during the pathological wound healing process following skin injury. Although the use of adipose-derived mesenchymal stem cells (ADSCs) to treat keloids and their anti-fibrotic effects have been a subject of intense research, the precise mechanism and efficacy of this therapy remain unclear. This study aimed to identify an effective ADSC therapy for keloids. Methods We compared the transcriptome characteristics of keloid fibroblasts (KF) and normal fibroblasts, as well as keloid keratinocytes (KK) and normal keratinocytes. Using differentially expressed gene analysis, gene set enrichment analysis, protein-protein interaction network analysis, hub gene analysis, and transcription factor regulation analysis; we identified the molecular characteristics underlying keloid pathogenesis. Furthermore, we used single-cell RNA sequencing and bioinformatics analysis to identify functional ADSC subclusters with inhibitory effects on keloid development. Finally, we investigated the effectiveness of the identified ADSC subclusters in preventing hypertrophic scar formation using a miniature pig model. Results We found that genes such as NOG and IL6 were up-regulated in KF, and interacted with other molecules to promote immune regulation, angiogenesis, and tissue regeneration, leading to the formation of KF. In KK, genes such as APP, NOTCH1 regulated migration, invasion, proliferation and fate determination of cells, leading to the formation of KK. We successfully identified hADSC subclusters that inhibited the development of KF, KK, or both, and miniature pigs treated by pADSCs with higher ratio of these therapeutic clusters shown better hypertrophic scar prophylaxis. Conclusion Our study provides a comprehensive understanding of the molecular characteristics of the keloid in humans, and is the first to identify therapeutic ADSC clusters for keloids, offering new avenues for stem cell-based keloid treatments.