Protective effect of ATP on skeletal muscle satellite cells damaged by H₂O₂

Abstract

This study investigated the protective effect of ATP on skeletal muscle satellite cells damaged by H₂O₂ in neonatal rats and the possible mechanism. The skeletal muscle satellite cells were randomly divided into four groups: normal group, model group (cells treated with 0.1 mmol/L H₂O₂ for 50 s), protection group (cells treated with 16, 8, 4, 2, 1, 0.5, or 0.25 mmol/L ATP for 24 h, and then with 0.1 mmol/L H₂O₂ for 50 s), proliferation group (cells treated with 16, 8, 4, 2, 1, 0.5, or 0.25 mmol/L ATP for 24 h). MTT assay, FITC+PI+DAPI fluorescent staining, Giemsa staining and immunofluorescence were performed to examine cell viability and apoptosis, and apoptosis-related proteins. The results showed that the survival rate of skeletal muscle satellite cells was decreased and the apoptosis rate was increased after H₂O₂ treatment (P<0.01). Different doses of ATP had different effects on skeletal muscle satellite cells damaged by H₂O₂: the survival rate of muscle satellite cells treated with ATP at 4, 2, or 1 mmol/L was increased. The protective effect was most profound on cells treated with 2 mmol/L ATP. Immunofluorescence showed that ATP could increase the number of Bcl-2-positive cells (P<0.01) and decrease the number of the Bax-positive cells (P<0.01). It was concluded that ATP could protect skeletal muscle satellite cells against H₂O₂ damage in neonatal rats, which may be attributed to the up-regulation of the expression of Bcl-2 and down-regulation of Bax, resulting in the suppression of apoptosis.

Keywords

adenosine triphosphate / hydrogen peroxide / damage / protection / skeletal muscle satellite cells / Bcl-2 / Bax

References

[1] BurnstockG. Purinergic signaling: past, present and future. Braz J Med Biol Res, 2009, 42(1): 3-8 PMID: 18853040

[2] ErlingeD, BurnstockG. P2 receptors in cardiovascular regulation and disease. Purinergic Signal, 2008, 4(1): 1-20 PMCID: 2245998 PMID: 18368530

[3] LiFH, GongHM, FengSB, et al. . Research progress of skeletal muscle satellite cells. Zhongguo Xumu yu Shouyi Kexue (Chinese), 2010, 37(10): 25-28

[4] ChargeSB, RudnickiMA. Cellular and molecular regulation of muscle regeneration. Physiol Rev, 2004, 84(3): 209-238 PMID: 14715915

[5] RytenM, YangSY, DunnPM, et al. . Purinoceptor expression in regenerating skeletal muscle in the mdx mouse model of muscular dystrophy and in satellite cell cultures. FASEB J, 2004, 18(12): 1404-1406 PMID: 15231720

[6] PallafacchinaG, FrançoisS, RegnaultB, et al. . An adult tissue-specific stem cell in its niche: a gene profiling analysis of in vivo quiescent and activated muscle satellite cells. Stem Cell Res, 2010, 4(1): 77-91 PMID: 19962952

[7] GharaibehB, Chun-LansingerY, HagenT, et al. . Biological approaches to improve skeletal muscle healing after injury and disease. Birth Defects Res C: Embryo Today, 2012, 96(2): 82-94

[8] SassoliC, Zecchi-OrlandiniS, FormigliL. Trophic actions of bone marrow-derived mesenchymal stromal cells for muscle repair/regeneration. Cells, 2012, 1(5): 832-850 PMCID: 3901134 PMID: 24710532

[9] YinH, PriceF, ArudnickiM. Satellite cells and the muscle stem cell niche. Physiol Rev, 2013, 93(1): 23-67 PMCID: 4073943 PMID: 23303905

[10] BestTM, GharaibehB, HuardJ. Stem cells, angiogenesis and muscle healing: a potential role in massage therapies?. Postgrad Med J, 2013, 89(5): 666-670 PMID: 24129034

[11] YanS, McfarlandDC, VellemanSG. Fibroblast growth factor 2 and protein kinase C alpha are involved in syndecan-4 cytoplasmic domain modulation of turkey myogenic satellite cell proliferation. Comp Biochem Physio Part A, 2012, 161(1): 44-52

[12] SciancaleporeM, LuinE, ParatoG, et al. . Reactive oxygen species contribute to the promotion of the ATP-mediated proliferation of mouse skeletal myoblasts. Free Radic Biol Med, 2012, 53(7): 1392-1398 PMID: 22917975

[13] MartinelloT, BaldoinMC, MorbiatoL, et al. . Extracellular ATP signaling during differentiation of C2C12 skeletal muscle cells: role in proliferation. Mol Cell Biochem, 2011, 351(1–2): 183-196 PMID: 21308481

[14] TanakaS, SaitoK, ReedJC. Structure-function analysis of the Bcl-2 oncoprotein. Addition of a heterologous transmembrane domain to portions of the Bcl-2 beta protein restores function as a regulator of cell survival. Biol Chem, 1993, 268(15): 10920-10926

[15] ZhengY, QinH, FrankSJ, et al. . A CK2-dependent mechanism for activation of the JAK-STAT signaling pathway. Blood, 2011, 118(1): 156-166 PMCID: 3139382 PMID: 21527517

[16] ZhaoSF. Muscle exercise and apoptosis in the skeletal muscle. Linchuang Chongjian Zuzhi Gongcheng Yanjiu Zazhi (Chinese), 2010, 14(11): 2030-2034

[17] ZhuDL, WangKL, ChenPL, et al. . Effect of oxymatrine on H₂O₂ resistance in L6 rat myoblast cells. Prog Biochem Biophys, 2012, 39(7): 662-670

[18] ZhuDL, WangKL, ChenPL, et al. . Analysis of ATP on H₂O₂ apoptosis model resistance in rat myoblast cells. Beijing Gongye Daxue Xuebao (Chinese), 2013, 39(2): 314-320