Neferine, a bisbenzylisoquinoline alkaloid, offers protection against cobalt chloride-mediated hypoxia-induced oxidative stress in muscle cells

BACKGROUND: Neferine, a bisbenzylisoquinoline alkaloid, isolated from Nelumbo nucifera has a wide range of biological activities. Cobalt chloride (CoCl(2)) was known to mimic hypoxic condition. In the present study, we assessed the cytoprotective effect of neferine against CoCl(2)-induced oxidative stress in muscle cells. METHODS: Rhabdomyosarcoma cells were exposed to different concentrations of CoCl(2), and the IC(50) value was determined using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. Lactate dehydrogenase and NO assays were performed in order to determine the cytotoxic effect of CoCl(2). Reactive oxygen species generation and cellular antioxidant status were determined for evaluating oxidative stress. For analyzing the effect of neferine on CoCl(2)-induced apoptosis, propidium iodide staining was performed. RESULTS: The results of the present study indicate that CoCl(2) induces cell death in a dose-dependent manner. Neferine pretreatment at 700 nM concentration offers better cytoprotection in the cells exposed to CoCl(2). Lactate dehydrogenase and NO release in the culture medium were restored after neferine pretreatment. CoCl(2) triggers time-dependent reactive oxygen species generation in muscle cells. Further, results of propidium iodide staining, mitochondrial membrane potential, and intracellular calcium accumulation confirm that neferine offers protection against CoCl(2)-induced hypoxic injury. Depleted activities of antioxidants such as superoxide dismutase, catalase, glutathione peroxidase, and glutathione S-transferase due to CoCl(2) exposure were also reinstated in the group that received neferine pretreatment. CONCLUSION: Our study suggests that neferine from N. nucifera offers protection to muscle cells by counteracting the oxidative stress induced by CoCl(2).