Effect Of Sarcolipin-mediated Cell Transdifferentiation In Senescence-associated Skeletal Muscle Fibrosis

Objective:To explore the role of SLN in senescence-related skeletal muscle fibtoric changes.Methods:Firstly,C2C12 myoblasts were induced to senescence with D-gal at different concentrations(0,10,20,40 g / L),cell proliferation and apoptosis were detected by CCK8 kit and flow cytometry,respectively.The senescent cells were stained blue by ?-galactosidase kit,and the senescence-related protein expression of P16 and P53 were detected by Western Blot.Secondly,Western blot and qPCR were used to detect the expression of fibrotic indicators TGF-?1,Col1 and Col3 in the senescence group and control group;the expression of SLN was detected using a laser scanning confocal microscopy(LSCM)and qPCR,and the changes of intracellular calcium ions was also measured.Finally,the transfection efficiency of recombinant adenovirus containing SLN gene(Ad-SLN)and recombinant adenovirus containing green fluorescent protein gene(Ad-GFP)in C2C12 cells was observed with an inverted fluorescence microscope,the protein expression levels of Col1 and Col3,transdifferentiation-related indicators ?-SMA and TGF-?1 were detected by qPCR and Western blot.Results : D-gal successfully induced the senescence of C2C12 myoblasts.CCK8 and flow cytometry showed that the cell proliferation activity decreased significantly at the concentration of 20 g / L of D-gal.The number of late apoptotic cells were increased,and remained cells were in good condition;?-Galactosidase staining showed that the number of senescent cells were increased significantly,and the protein expression of senescence-related protein P16 and P53 were increased also;the fibrotic indicators TGF-?1 and Col1,Col3 mRNA and protein expression levels were increased in senescence group;SLN expression also increased significantly in senescence group.The intracellular calciumion was increased in D-gal group.After Ad-SLN and Ad-GFP successfully transfected into C2C12 cells,SLN mRNA expression was reduced,the expression of myofibroblast cell marker ?-SMA protein was significantly reduced,and fibrotic indicators TGF-?1 and collagen protein expression were also suppressed.Conclusion : SLN promotes myoblasts to change into fibrotic phenotype by regulating cell transdifferentiation.Decreasing SLN expression can inhibit myoblasts into myofibroblasts and alleviate skeletal muscle fibrosis.