Abstract
BACKGROUND: Skeletal muscle injury is one of the most common sports injuries; if not properly treated or not effective rehabilitation treatment after injury, it can be transformed into chronic cumulative injury. Curcumin, an herbal ingredient, has been found to promote skeletal muscle injury repair and regeneration. The Wnt5a pathway is related to the expression of myogenic regulatory factors, and Ca(2+) promotes the differentiation and fusion process of myoblasts. This study explored the effect and mechanism of curcumin on myoblast differentiation during the repair and regeneration of injured skeletal muscle and its relationship with the Wnt5a pathway and Ca(2+) channel. METHODS: Myogenic differentiation of C2C12 cells was induced with 2% horse serum, and a mouse (male, 10 weeks old) model of acute skeletal muscle injury was established using cardiotoxin (20 μL). In addition, we constructed a Wnt5a knockdown C2C12 cell model and a Wnt5a knockout mouse model. Besides, curcumin was added to the cell culture solution (80 mg/L) and fed to the mice (50 mg/kg). Fluorescence microscopy was used to determine the concentration of Ca(2+). Western blot and RT-qPCR were used to detect the protein and mRNA levels of Wnt5a, CaN, NFAT2, MyoD, Myf5, Pax7, and Myogenin. The expression levels of MyoD, Myf5, Myogenin, MHC, Desmin, and NFAT2 were detected using immunofluorescence techniques. In addition, MyoD expression was observed using immunohistochemistry, and morphological changes in mouse muscle tissue were observed using HE staining. RESULTS: During myoblast differentiation and muscle regeneration, Wnt5a expression was upregulated (P < 0.001) and the Wnt5a signalling pathway was activated. Wnt5a overexpression promoted the expression of MyoD, Myf5, Myogenin, MHC, and Desmin (P < 0.05), and conversely, knockdown of Wnt5a inhibited their expression (P < 0.001). The Wnt5a pathway mediated the opening of Ca(2+) channels, regulated the expression levels of CaN, NFAT2, MyoD, Myf5, Myogenin, MHC, and Desmin (P < 0.01) and promoted the differentiation of C2C12 myoblasts and the repair and regeneration of injured skeletal muscle. The expression of Wnt5a, CaN, NFAT2, MyoD, Myogenin, Myf5, and MHC in C2C12 myoblast was significantly increased after curcumin intervention (P < 0.05); however, their expression decreased significantly after knocking down Wnt5a on the basis of curcumin intervention (P < 0.05). Similarly, in Wnt5a knockout mice, the promotion of muscle regeneration by curcumin was significantly attenuated. CONCLUSIONS: Curcumin can activate the Wnt5a signalling pathway and mediate the opening of Ca(2+) channels to accelerate the myogenic differentiation of C2C12 cells and the repair and regeneration of injured skeletal muscle.