The Effects of Quercetin on Oxidative Stress-Induced Cell Death in C2C12 Muscle Cells.

Abstract

PURPOSE: Reactive oxygen species (ROS) are unstable molecules that alter the structure and function of proteins, lipids, and DNA and are produced in both resting and contracting skeletal muscle. The purpose of this research was to examine the dosedependent antioxidant effects of quercetin, a natural antioxidant, to attenuate oxidative stress in skeletal muscle cells. METHODS: C2C12 mouse muscle cells (myoblasts) were grown in 96 well microplates and allowed to settle for 24hrs. 1.15mM H2O2 was empirically determined to elicit severe oxidative stress-induced cell death. Cells were divided into two groups: Group 1 (G1) was pre-conditioned with 1.0-100μm quercetin (dissolved in dimethyl sulfoxide (DMSO)) for 18hrs then challenged with H2O2 for 6hrs. Group 2 (G2) was preconditioned with quercetin for 18hrs then co-cultured with quercetin and H2O2 for 6hrs. Culture media was changed between pre-conditioning and stress conditions to avoid residual effects of quercetin in culture media. Cell viability was assessed using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) Assay and expressed as % of negative controls. A one-way analysis of variance (ANOVA) and post-hoc tests were performed with the Statistical Analysis Software System (SAS). RESULTS: DMSO (0.01-1.4% vol/vol) did not affect cell viability in C2C12 cells (p=0.6). Dramatic cell death (86%, p<0.01) was observed in cells treated with H2O2 in the absence of quercetin. Quercetin at [5-100μm] significantly attenuated oxidative stress induced-cell death compared with controls (44-20% vs. 86%, p<0.01, respectively) in G1 and G2 (32-14% vs. 86%, p<0.01). Quercetin at [5-10μm] improved cell viability in a dose dependent manner but significant differences were not observed among higher treatment levels (25-100μm). No significant differences were observed between treatment levels x group (G1 vs. G2, p>0.06), indicating that preconditioning with quercetin alone was as effective as preconditioning and concomitant treatment with quercetin and H2O2. Non-significant differences between experimental groups suggest that quercetin may exert its effects at the cellular-level. CONCLUSIONS: These findings demonstrate quercetin is a potent antioxidant to rescue muscle cells from severe oxidative stress-induced cell death.