Background: In plastic surgery, tissue expansion is widely used for repairing skin defects. However, low expansion efficiency and skin rupture caused by thin, expanded skin remain significant challenges in promoting skin regeneration during expansion. S100 calcium-binding protein A9 (S100A9) is essential in promoting wound healing; however, its effects on skin regeneration during tissue expansion remain unclear. The aim of the present study was to explore the role of S100A9 in skin regeneration, particularly collagen production to investigate its importance in skin regeneration during tissue expansion. Methods: The expression and distribution of S100A9 and its receptors—toll-like receptor 4 (TLR-4) and receptor for advanced glycation end products were studied in expanded skin. These characteristics were investigated in skin samples of rats and patients. Moreover, the expression of S100A9 was investigated in stretched keratinocytes in vitro. The effects of S100A9 on the proliferation and migration of skin fibroblasts were also observed. TAK-242 was used to inhibit the binding of S100A9 to TLR-4; the levels of collagen I (COL I), transforming growth factor beta (TGF-β), TLR-4 and phospho-extracellular signal-related kinase 1/2 (p-ERK1/2) in fibroblasts were determined. Furthermore, fibroblasts were co-cultured with stretched S100A9-knockout keratinocytes by siRNA transfection and the levels of COL I, TGF-β, TLR-4 and p-ERK1/2 in fibroblasts were investigated. Additionally, the area of expanded skin, thickness of the dermis, and synthesis of COL I, TGF-β, TLR-4 and p-ERK1/2 were analysed to determine the effects of S100A9 on expanded skin. Results: Increased expression of S100A9 and TLR-4 was associated with decreased extracellular matrix (ECM) in the expanded dermis. Furthermore, S100A9 facilitated the proliferation and migration of human skin fibroblasts as well as the expression of COL I and TGF-β in fibroblasts via the TLR-4/ERK1/2 pathway. We found that mechanical stretch-induced S100A9 expression and secretion of keratinocytes stimulated COL I, TGF-β, TLR-4 and p-ERK1/2 expression in skin fibroblasts. Recombined S100A9 protein aided expanded skin regeneration and rescued dermal thinning in rats in vivo as well as increasing ECM deposition during expansion. Conclusions: These findings demonstrate that mechanical stretch promoted expanded skin regeneration by upregulating S100A9 expression. Our study laid the foundation for clinically improving tissue expansion using S100A9.

Background: Tissue expansion, a technique in which skin regeneration is induced by mechanical stretch stimuli, is commonly used for tissue repair and reconstruction. In this study, we aimed to monitor the autophagy levels of expanded skin after the application of expansion stimuli and explore the effect of autophagy modulation on skin regeneration.Methods: A rat scalp expansion model was established to provide a stable expanded skin response to mechanical stretch. Autophagy levels at different time points (6, 12, 24, 48 and 72 h after the last expansion) were detected via western blotting. The effect of autophagy regulation on skin regeneration during tissue expansion was evaluated via skin expansion efficiency assessment, western blotting, immunofluorescence staining, TUNEL staining and laser Doppler blood flow imaging.Results: The autophagic flux reached its highest level 48 h after tissue expansion. Activating autophagy by rapamycin increased the area of expanded skin as well as the thicknesses of epidermis and dermis. Furthermore, activating autophagy accelerated skin regeneration during tissue expansion by enhancing the proliferation of cells and the number of epidermal basal and hair follicle stem cells, reducing apoptosis, improving angiogenesis, and promoting collagen synthesis and growth factor secretion. Conversely, the regenerative effects were reversed when autophagy was blocked.Conclusions: Autophagy modulation may be a promising therapeutic strategy for improving the efficiency of tissue expansion and preventing the incidence of the complication of skin necrosis.

Mechanical stretch is utilized in the process of tissue expansion to promote skin regeneration, which is crucial for wound healing and organ reconstruction purposes. Enlarged dermal area is one of the significant histological characteristics of the expanded skin. However, the underlying biological processes and molecular pathways associated with dermal regeneration triggered by mechanical stretch are still not well understood. Twelve male Sprague-Dawley (SD) rats were divided into the expansion group and sham group randomly. Upon creating a rat scalp expansion model, the dermis was isolated from the full-thickness skin in both experimental groups for RNA sequencing. This process led to the identification of differentially expressed genes (DEGs). Subsequently, we conducted Gene Ontology (GO) analysis, Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis, and Gene Set Enrichment Analysis (GSEA) to identify the essential biological processes associated with dermal regeneration induced by mechanical stretch, leveraging data from the DEGs. A network of protein-protein interactions (PPI) was built to detect the critical modules and central genes. The expression levels of these hub genes were evaluated using quantitative real-time polymerase chain reaction (qPCR). Increased expanded skin area and dermal thinning which represent the typical changes of expanded skin were observed in the expansion group. A total of 782 DEGs were identified in the expansion group relative to the sham group. The DEGs were associated with several biological processes, including the organization of the extracellular matrix, the enhancement of macrophage activation, and the promotion of angiogenesis, among others. Cell components encompassing Toll-like receptor 2-Toll-like receptor 6 protein complex, interstitial matrix, extracellular matrix (ECM), and collagen trimer were discovered. Molecular function categories including integrin binding, insulin-like growth factor binding, and fatty acid elongase activity were involved. The KEGG pathway analysis demonstrated the significant enrichment of pathways including the PI3K-Akt signaling pathway, fatty acid metabolism, and extracellular matrix-receptor interactions. GSEA results displayed that mechanical stretch correlated with the regulation of cell activation processes, cytokine-mediated signaling pathways, and immune system processes. PPI network resulted in the identification of 598 nodes along with a total of 5,304 interaction pairs between proteins. And ten hub genes containing Ccl2, Cxcl10, Fasn, Itgad, Cd163, Mmp9, Cd36, Tlr2, Igf1, and Wnt2 were identified by bioinformatics analysis and validated by qPCR. This in vivo study for the first time revealed the DEGs related to mechanical stretch stimulated dermal regeneration and identified the involved pathways and hub genes correlated with macrophage recruitment and polarization, fibroblast proliferation and ECM production and angiogenesis, which may benefit further studies aimed at developing therapeutic strategies for facilitating expanded skin regeneration.