Activation of stress signaling pathways by the end product of lipid peroxidation - 4-hydroxy-2-nonenal is a potential inducer of intracellular peroxide production

In the present study, we studied the signal transduction mechanism that is involved in the expression of c-Jun protein evident after exposure of rat l iver epithelial RL34 cells to the major end product of oxidized fatty acid metabolism, 4-hydroxy-2-nonenal (HNE). HNE treatment of the cells resulted in depletion of intracellular glutathione (GSH) and in the formation of pro tein-bound HNE in plasma membrane. In addition, HNE strongly induced intrac ellular peroxide production, suggesting that HNE exerted oxidative stress o n the cells. Potent expression of c-Jun occurred within 30 min of HNE treat ment, which was accompanied by a time-dependent increase in activator prote in-1 (AP-1) DNA binding activity. We found that HNE caused an immediate inc rease in tyrosine phosphorylation in RL34 cells, In addition, HNE strongly induced phosphorylation of c-Jun N-terminal kinases (JNK) and p38 mitogen-a ctivated protein kinases and also moderately induced phosphorylation of ext racellular signal-regulated kinases. The phosphorylation of JNK was accompa nied by a rapid and transient increase in JNK and p38 activities, whereas c hanges in the activity of extracellular signal-regulated kinase were scarce ly observed. GSH depletion by L-buthionine-S,R-sulfoximine, a specific inhi bitor of GSH biosynthesis, only slightly enhanced peroxide production and J NK activation, suggesting that HNE exerted these effects independent of GSH depletion. This and the findings that (i) HNE strongly induced intracellul ar peroxide production, (ii) HNE-induced JNK activation was inhibited by pr etreatment of the cells with a thiol antioxidant, N-acetylcysteine, and (ii i) H2O2 significantly activated JNK support the hypothesis that pro-oxidant s play a crucial role in the HNE-induced activation of stress signaling pat hways. In addition, we found that, among the inhibitors of tyrosine kinases , cyclooxygenase, and Ca2+ influx, only quercetin exerted a significant inh ibitory effect on HNE-induced JNK activation. In light of the JNK-dependent induction of c-jun transcription and the AP-1-induced transcription of xen obiotic-metabolizing enzymes, these data may show a potential critical role for JNK in the induction of a cellular defense program against toxic produ cts generated from lipid peroxidation.