Nitric oxide protects cultured rat hepatocytes from tumor necrosis factor-α-induced apoptosis by inducing heat shock protein 70 expression

Nitric oxide (NO) and tumor necrosis factor-α (TNFα) play important roles in the pathogenesis of liver disease during acute inflammation. The present study was designed to elucidate the effect of NO pre-exposure on TNFα-induced hepatotoxicity. Pretreatment of primary cultures of rat hepatocytes with the NO donor S-nitroso-N-acetylpenicillamine (SNAP) induced the expression of heat shock protein 70 (HSP70) mRNA and protein, which was associated with thermotolerance and cytoprotection from TNFα+actinomycin D- induced hepatotoxicity and apoptosis. SNAP transiently changed the intracellular redox state by inducing glutathione (GSH) oxidation associated with the formation of S-nitrosoglutathione (GSNO). HSP70 mRNA was also induced by the GSH-oxidizing agent diamide and the GSH-conjugating agent N- ethylmaleimide, suggesting that NO induces HSP70 expression through GSH oxidation. The protective effect of SNAP pretreatment on TNFα-induced apoptosis correlated with the level of HSP70 expression. SNAP pretreatment inhibited reactive oxygen intermediate generation and lipid peroxidation effects that were reversed by blocking HSP70 expression using an antisense oligonucleotide to HSP70. Finally, endogenous NO formation, induced in hepatocytes stimulated with interferon-γ and interleukin-1β, led to the formation of GSNO and GSSG, induced HSP70, and attenuated TNFα-mediated cytotoxicity. These findings demonstrated that NO can induce resistance to TNFα-induced hepatotoxicity, possibly through the stimulation of HSP70 expression.