The c-jun N-terminal kinases (JNK) / mitogen-activated protein kinase (MAPK) is responsible for the protection of tanshinol (Danshensu) upon H2 O2 -induced L6 rat myoblast cell injury

ABSTRACT

Background: Promoting L6 rat myoblast cell (L6 RMC) survival in the pro-apoptotic environment is critical to myoblastcell replacement for skeletal muscle degenerative disease therapy. Tanshinol (Danshensu), one of the principal bioactivecomponents in salvia miltiorrhiza bunge, has been used widely in skeletal muscle system (SMS) diseases treatment andserves as an antioxidant to protect myoblast cells against oxidative stress. The present study was undertaken to investigatethe protective effects of Tanshinol on L6 RMC injury induced by hydrogen peroxide (H2O2). After challenge with 100 µMH2O2 for 1h, loss of cell viability and excessive apoptotic cell death were observed in cultured L6 RMC, tanshinol treatmentconferred protective effects against the loss of cellular viability in a concentration-dependent manner.Materials, Methods & Results: L6 rat myoblast cells (L6 RMC) were maintained DMEM containing 4.5 g/L glucose andsupplemented with 10% (v/v) fetal bovine serum (FBS), 100 U/mL penicillin and 100 µg/mL streptomycin on tissue culturefl asks in a 37°C humidifi ed atmosphere of 95% air and 5% CO2. Cells were subcultured every 2-3 days. The cells werestained with SABC-Cy3 fl uorescence and measured by counting the nuclei. L6 RMC viability was determined by MTTassay in 96-well plates. The cells were counter-stained with H & E staining and Hoechst 33342 fl uorescent staining. Cellcycle phase distribution and apoptosis rates were detected by fl ow cytometry. Western blot were repeated three times, andqualitatively similar results were obtained. L6 RMC were pretreated with Tanshinol 375, 187.5, 93.75 mg/mL for 24 h,followed by treatment of 1000, 500, 100 µM H2O2 for 1h (P < 0.05) with viability by MTT method. Tanshinol 187.5 mg/mL pretreatment was protected in L6 RMC from H2O2-induced apoptosis with H&E staining and Hoechst 33342 fl uorescent...(AU)