Objective:To investigate the effects and mechanisms of capsaicin on full-thickness skin defects in diabetic mice.Methods:This study was an experimental study. Thirty-six male db/db mice aged 6-8 weeks were taken. Circular full-thickness skin defect wounds (6 mm in diameter) were created on their backs. According to the random number table method (grouping method same below), the mice were divided into control group, low-concentration capsaicin group, and high-concentration capsaicin group injected with normal saline, 10 μmol/L capsaicin solution, and 20 μmol/L capsaicin solution, respectively ( n=12). Immediately after modeling and on day 2, 30 μL of the corresponding solution was injected locally into the wounds. At 4, 8, and 12 days after injury, wound healing status was observed grossly and the percentage of residual wound area was calculated. At 12 days after injury, the proportions of inflammatory cell, collagen fiber, and CD31-positive expression areas in the wound of mice were observed and detected respectively using hematoxylin and eosin staining, Masson staining, and immunohistochemical staining, and the protein expression of transient receptor potential vanilloid type 1 (TRPV1) in the wound tissue of mice was detected using Western blotting. Human primary fibroblasts were prepared from normal skin tissue obtained from 5 patients (2 male and 3 female patients, aged 20-45 years) who were admitted to the Department of Plastic and Reconstructive Surgery of Shanghai Ninth People's Hospital of Shanghai Jiao Tong University School of Medicine in October 2024. Cells in the logarithmic growth phase (passages 2-5) were used for subsequent experiments. Cells were divided into control group and high-concentration capsaicin group, cultured in complete media without or with 20 μmol/L capsaicin, respectively. After 24 hours of culture, differentially expressed genes (DEGs) between two groups were identified using the DESeq2 R package, followed by gene ontology (GO) and Kyoto encyclopedia of genes and genomes (KEGG) pathway enrichment analysis. The cells were divided into control group, low-concentration capsaicin group, and high-concentration capsaicin group, which were cultured in complete media without capsaicin, and with 10 μmol/L capsaicin, and with 20 μmol/L capsaicin, respectively. After 48 hours of culture, cell apoptosis status was assessed using flow cytometer. The protein expression levels of protein kinase B (Akt), phosphorylated Akt (p-Akt), mammalian target of rapamycin (mTOR), and phosphorylated mTOR (p-mTOR) in cells were detected by Western blotting, and the p-Akt/Akt and p-mTOR/mTOR ratios were calculated. At 12 days after injury, the protein expression levels of Akt, p-Akt, mTOR, and p-mTOR in the wounds of diabetic mice in both control group and high-concentration capsaicin group were detected by Western blotting, and the p-Akt/Akt and p-mTOR/mTOR ratios were calculated. All animal experiments used a sample size of 6, and all cellular experiments used 3. Results:At 4 days after injury, the wounds of three groups of diabetic mice began to heal gradually, and the percentages of residual wound area of mice in both low-concentration capsaicin group and high-concentration capsaicin group were significantly lower than that in control group (with t values of 2.31 and 2.87, respectively, P<0.05). At 8 days after injury, the percentages of residual wound area of mice in both low-concentration capsaicin group and high-concentration capsaicin group were significantly lower than that in control group (with t values of 2.55 and 5.38, respectively, P<0.05). At 12 days after injury, the percentages of residual wound area of mice in both low-concentration capsaicin group and high-concentration capsaicin group remained significantly lower than that in control group (with t values of 3.31 and 6.24, respectively, P<0.05), with the high-concentration capsaicin group showing a significantly greater reduction compared to that in low-concentration capsaicin group ( t=3.42, P<0.05). At 12 days after injury, the proportion of inflammatory cell area in the wound of mice in high-concentration capsaicin group was (6.2±1.8)%, significantly lower than (15.5±3.0)% in control group ( t=6.45, P<0.05). The proportion of collagen fiber area, proportion of CD31-positive expression area, and protein expression of TRPV1 in the wound of mice in high-concentration capsaicin group were significantly higher compared with those in control group (with t values of 5.48, 7.11, and 15.41, respectively, P<0.05). After 24 hours of culture, 51 DEGs with significantly differential expression were detected in high-concentration capsaicin group of cells compared with those in control group ( P<0.05), with 31 upregulated and 20 downregulated genes. GO analysis showed that the significantly upregulated and significantly downregulated DEGs mainly participated in biological processes such as extracellular matrix (ECM) polymerization, extracellular structure organization, collagen metabolic process regulation, and ECM component secretion regulation. KEGG analysis showed that the significantly upregulated and significantly downregulated DEGs mainly participated in cell apoptosis-related pathways such as the phosphatidylinositol 3-kinase/Akt pathway and tumor necrosis factor signaling pathway. After 48 hours of culture, the cell apoptosis rates in both low-concentration capsaicin group and high-concentration capsaicin group were significantly lower than that in control group (with t values of 6.38 and 9.09, respectively, P<0.05). The p-mTOR/mTOR ratio in cells in low-concentration capsaicin group was significantly higher than that in control group ( t=2.74, P<0.05). The p-Akt/Akt and p-mTOR/mTOR ratios in cells in high-concentration capsaicin group were significantly higher than those in control group (with t values of 4.43 and 3.33, respectively, P<0.05). At 12 days after injury, the p-Akt/Akt and p-mTOR/mTOR ratios in wounds of diabetic mice in high-concentration capsaicin group were 0.470±0.044 and 0.549±0.106, respectively, which were significantly higher than 0.189±0.058 and 0.241±0.120 in control group (with t values of 6.67 and 3.36, respectively, P<0.05). Conclusions:Capsaicin can promote the healing of full-thickness skin defect wounds in diabetic mice by activating the Akt/mTOR signaling pathway in fibroblasts, thereby inhibiting apoptosis.

Objective:To investigate the effects and mechanisms of capsaicin on full-thickness skin defects in diabetic mice.Methods:This study was an experimental study. Thirty-six male db/db mice aged 6-8 weeks were taken. Circular full-thickness skin defect wounds (6 mm in diameter) were created on their backs. According to the random number table method (grouping method same below), the mice were divided into control group, low-concentration capsaicin group, and high-concentration capsaicin group injected with normal saline, 10 μmol/L capsaicin solution, and 20 μmol/L capsaicin solution, respectively ( n=12). Immediately after modeling and on day 2, 30 μL of the corresponding solution was injected locally into the wounds. At 4, 8, and 12 days after injury, wound healing status was observed grossly and the percentage of residual wound area was calculated. At 12 days after injury, the proportions of inflammatory cell, collagen fiber, and CD31-positive expression areas in the wound of mice were observed and detected respectively using hematoxylin and eosin staining, Masson staining, and immunohistochemical staining, and the protein expression of transient receptor potential vanilloid type 1 (TRPV1) in the wound tissue of mice was detected using Western blotting. Human primary fibroblasts were prepared from normal skin tissue obtained from 5 patients (2 male and 3 female patients, aged 20-45 years) who were admitted to the Department of Plastic and Reconstructive Surgery of Shanghai Ninth People's Hospital of Shanghai Jiao Tong University School of Medicine in October 2024. Cells in the logarithmic growth phase (passages 2-5) were used for subsequent experiments. Cells were divided into control group and high-concentration capsaicin group, cultured in complete media without or with 20 μmol/L capsaicin, respectively. After 24 hours of culture, differentially expressed genes (DEGs) between two groups were identified using the DESeq2 R package, followed by gene ontology (GO) and Kyoto encyclopedia of genes and genomes (KEGG) pathway enrichment analysis. The cells were divided into control group, low-concentration capsaicin group, and high-concentration capsaicin group, which were cultured in complete media without capsaicin, and with 10 μmol/L capsaicin, and with 20 μmol/L capsaicin, respectively. After 48 hours of culture, cell apoptosis status was assessed using flow cytometer. The protein expression levels of protein kinase B (Akt), phosphorylated Akt (p-Akt), mammalian target of rapamycin (mTOR), and phosphorylated mTOR (p-mTOR) in cells were detected by Western blotting, and the p-Akt/Akt and p-mTOR/mTOR ratios were calculated. At 12 days after injury, the protein expression levels of Akt, p-Akt, mTOR, and p-mTOR in the wounds of diabetic mice in both control group and high-concentration capsaicin group were detected by Western blotting, and the p-Akt/Akt and p-mTOR/mTOR ratios were calculated. All animal experiments used a sample size of 6, and all cellular experiments used 3. Results:At 4 days after injury, the wounds of three groups of diabetic mice began to heal gradually, and the percentages of residual wound area of mice in both low-concentration capsaicin group and high-concentration capsaicin group were significantly lower than that in control group (with t values of 2.31 and 2.87, respectively, P<0.05). At 8 days after injury, the percentages of residual wound area of mice in both low-concentration capsaicin group and high-concentration capsaicin group were significantly lower than that in control group (with t values of 2.55 and 5.38, respectively, P<0.05). At 12 days after injury, the percentages of residual wound area of mice in both low-concentration capsaicin group and high-concentration capsaicin group remained significantly lower than that in control group (with t values of 3.31 and 6.24, respectively, P<0.05), with the high-concentration capsaicin group showing a significantly greater reduction compared to that in low-concentration capsaicin group ( t=3.42, P<0.05). At 12 days after injury, the proportion of inflammatory cell area in the wound of mice in high-concentration capsaicin group was (6.2±1.8)%, significantly lower than (15.5±3.0)% in control group ( t=6.45, P<0.05). The proportion of collagen fiber area, proportion of CD31-positive expression area, and protein expression of TRPV1 in the wound of mice in high-concentration capsaicin group were significantly higher compared with those in control group (with t values of 5.48, 7.11, and 15.41, respectively, P<0.05). After 24 hours of culture, 51 DEGs with significantly differential expression were detected in high-concentration capsaicin group of cells compared with those in control group ( P<0.05), with 31 upregulated and 20 downregulated genes. GO analysis showed that the significantly upregulated and significantly downregulated DEGs mainly participated in biological processes such as extracellular matrix (ECM) polymerization, extracellular structure organization, collagen metabolic process regulation, and ECM component secretion regulation. KEGG analysis showed that the significantly upregulated and significantly downregulated DEGs mainly participated in cell apoptosis-related pathways such as the phosphatidylinositol 3-kinase/Akt pathway and tumor necrosis factor signaling pathway. After 48 hours of culture, the cell apoptosis rates in both low-concentration capsaicin group and high-concentration capsaicin group were significantly lower than that in control group (with t values of 6.38 and 9.09, respectively, P<0.05). The p-mTOR/mTOR ratio in cells in low-concentration capsaicin group was significantly higher than that in control group ( t=2.74, P<0.05). The p-Akt/Akt and p-mTOR/mTOR ratios in cells in high-concentration capsaicin group were significantly higher than those in control group (with t values of 4.43 and 3.33, respectively, P<0.05). At 12 days after injury, the p-Akt/Akt and p-mTOR/mTOR ratios in wounds of diabetic mice in high-concentration capsaicin group were 0.470±0.044 and 0.549±0.106, respectively, which were significantly higher than 0.189±0.058 and 0.241±0.120 in control group (with t values of 6.67 and 3.36, respectively, P<0.05). Conclusions:Capsaicin can promote the healing of full-thickness skin defect wounds in diabetic mice by activating the Akt/mTOR signaling pathway in fibroblasts, thereby inhibiting apoptosis.