The Role Of MiR-155 In Wound Healing

Skin tissue defect is very common in clinic, which is generally caused by acute and chronic injury factors, such as mechanical damage, burns, body surface tumor, chronic ulcer causes. Wound healing is performed under complex pathophysiological enviroment by variety of cells and cytokines, mainly contains three classic stages: inflammation, wound epithelial and shrinkage and remodeling after collagen deposition. All the three stages are correlated with each other. It has always been the hot spot that how to promote wound healing and reduce the degree of fibrosis after burn and interdisciplinary. However, there is still no ideal result. Therefore, exploring new factor that could accelerate wound healing and reduce the degree of fibrosis is one of the important clinical problems that needed to be resolved.mi RNAs is a kind of short chains of non-coding RNA, through regulating gene expression level in the cell, play an important role in wound healing and fibrosis diseases. mi R-203 and mi R-210, respectively, by promoting keratinocytes migration and proliferation could promote wound healing, and mi R-99 and mi R-200 family form inhibiting keratinocytes migration could delaye wound healing. mi R-155 in the early research focus in the field of immune, and found that mi R-155 plays an important role in inflammatory cells and immune response regulation, a recent study also found that mi R-155 could promote the migration of epithelial cells by promoting the transformation of epithelial cell to mesenchymal cell. Therefore, we assume that mi R-155 plays an important role in wound healing and fibrosis after wound healing.Directly local injections of mi R-155 plasmid and antagomir in wound edge were used to increase or reduce mi R-155 levels in wound local, and in vitro cell model to study the role of mi R-155 in epithelium, wound contraction, wound inflammation, collagen deposition after wound healing, and its possible mechanism. Methods:1. After 1 x 1 cm full-thickness skin defects were made in the back of SD rats, direct local injections of mi R-155 plasmid to study the effect on wound healing and epithelium; Fibroblasts three dimensional gel contraction and α-SMA dyeing were used to observe the effect of mi R-155 plasmid on wound contraction in vivo and in vitro.2. Keratinocyte cells scratches and Transwell cell transmembrane experiments were used to detect the effect of mi R-155 on cell migration; Ki-67 dyeing, MTT and flow cytometry cycle measurement were used to detect the effect of mi R-155 plasmid on keratinocyte cell proliferation.3. Molecular mechanism of mi R-155 promoting keratinocyte cell migration were detected through F-actin, E-cadherin, β-catenin, MMPs and TIMPs as a protein involved in cell migration.4. Through the wounds HE and immune cells immunohistochemical staining to observe affect of mi R-155 plasmid in wound inflammation; to observe collagen deposition we detect the effect of mi R-155 on fibrosis after wound healing.5. Direct injection of mi R-155 antagomir in C57 mice back full-thickness skin defects model to detect effects on wound healing; local inflammatory cells HE and immunohistochemical staining, inflammatory factor levels were used to observe the affect of wound local injections of mi R-155 antagomir on wound local inflammatory response.6. By detecting partial collagen deposition quantity after the wound healing and Masson staining of collagen to observe mi R-155 antagomir local injections effects on fibrosis after wound healing. Results:1. In SD rats back wound model, wound local injections of mi R-155 plasmid could increase wound local mi R-155 level significantly compared with control group, and speeding up the wound healing, wound HE staining showed that mi R-155 can obviously promote wound epithelial change; three dimensional gel contraction of fibroblasts were not changed signficantly, a-SMA expression had no significant effect in vivo and in vitro. The result indicates that mi R- 155 can accelerate wound and promote wound healing, and had no obvious effect on wound contraction.2. Scratches and Transwell cell transmembrane cells experiments have established that mi R-155 can form promote cutin cell migration; mi R-155 has no obvious influence on keratinocyte Ki-67 expression, growth curve, cell cycle. The result indicates that mi R-155 could promote keratinocytes migration while had no obvious effect on the cell proliferation.3. Western blot and immunohistochemistry/fluorescence staining showed that mi R-155 had no obvious effect on E-cadherin, b-catenin, F-actin and MMP-9 protein expressions; mi R-155 can cause increased expression of MMP-2, and MMP-2 specific inhibitors ARP101 can partial inhibition mi R-155 induced increasing migration. The result indicates that MMP-2 play an important role in the mi R-155 induced cell migration faster and wound healing speeding up.4. Wound local injections of mi R-155 plasmid can increase the quantity wound local neutrophil infiltration, Col1 and Col3 deposits increase after wound healing. Mi R-155 can enhance wound local inflammatory reaction and lead to increased after the degree of fibrosis.5. C57 mice back wound local injections of mi R-155 antagomir can significantly reduce the wound mi R- 155 levels, the number of macrophage neutrophil infiltration to wound local decreased and proinflammatory factor TNF-a, IL-b level significantly decreased, suppression of inflammation factor IL-10 levels increased significantly. Compared with the control group mi R-155 antagomir local injection had no obvious effect on wound healing. Results suggest that wound local low expression of mi R-155 can reduce inflammation and had no obvious effect on wound healing.6. wound healing after in C57 mice mi R-155 antagomir local injections the Col1, Col3, a-SMA expression level decreased obviously, Masson staining of collagen prompt local collagen arrangement more neatly and sparse. Show that mi R-155 antagomir can reduce the degree of fibrosis after wound healing, improve the quality of wound healing. Conclusion:Wound local high expression of mi R-155 can promote keratinocytes migration and accelerate wound healing, and wound inflammation and fibrosis degree enhancement after wound healing, MMP-2 play an important role in the mi R-155 induced keratinocytes migration faster. Wound local lower expression of mi R-155 can inhibit inflammatory reaction and relieve degree of fibrosis after wound healing.