Induction of endogenous antioxidants and phase 2 enzymes by alpha-lipoic acid in rat cardiac H9C2 cells: protection against oxidative injury.

Alpha-lipoic acid (LA) has recently been reported to exert protective effects on various forms of oxidative cardiac disorders. However, the mechanisms underlying LA-mediated cardioprotection remain to be investigated. This study was undertaken to determine whether LA treatment could increase endogenous antioxidants and phase 2 enzymes in cultured cardiomyocytes, and whether such increased cellular defenses could afford protection against oxidative cardiac cell injury. Incubation of rat cardiac H9C2 cells with low micromolar concentrations of LA resulted in a significant induction of a scope of cellular antioxidants and phase 2 enzymes in a concentration- and/or time-dependent fashion. These include catalase, reduced glutathione, glutathione reductase, glutathione S-transferase, and NAD(P)H:quinone oxidoreductase-1 (NOQ1). Induction of catalase and NOQ1 was most dramatic among the above LA-inducible antioxidants and phase 2 enzymes. To further investigate the protective effects of the LA-induced cellular defenses on oxidative cardiac cell injury, H9C2 cells were pretreated with LA (25-100 microM) for 72h and then exposed to xanthine oxidase (XO)/xanthine, a system that generates reactive oxygen species (ROS), for another 24h. We observed that LA pretreatment of H9C2 cells led to a marked protection against XO/xanthine-mediated cytotoxicity, as detected by 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium reduction assay. The cytoprotective effects also exhibited a LA concentration-dependent fashion. Moreover, the LA pretreatment resulted in a great inhibition of intracellular accumulation of ROS in H9C2 cells following incubation with XO/xanthine. Taken together, this study demonstrates for the first time that a number of endogenous antioxidants and phase 2 enzymes in cultured cardiomyocytes can be induced by LA at low micromolar concentrations, and that the LA-mediated elevation of cellular defenses is accompanied by a markedly increased resistance to ROS-elicited cardiac cell injury. The results of this study have important implications for the cardioprotective effects of LA.