Potential Role of the JNK/SAPK Signal Transduction Pathway in the Induction of iNOS by TNF-α

Nitric oxide production by macrophages is principally regulated by the calcium-independent enzyme, inducible nitric oxide synthase (iNOS). Both lipopolysaccharide and TNF-α synergize with IFN-γ in the expression of iNOS with subsequent production of nitric oxide. Previous work has shown that IL-4 downregulates iNOS and nitric oxide expression by macrophages stimulated with LPS and IFN-γ. In this study, we found that IL-4 also downregulated iNOS and nitric oxide expression induced by IFN-γ and TNF-α and in mouse macrophages. Because various members of the mitogen-activated protein kinases and their upstream kinases have been shown to directly or indirectly activate a number of transcription factors including AP-1 and NFκB, we examined the effects of IL-4 on TNF-α activation of the MAPKs. Our results show that IL-4 modestly inhibited JNK/SAPK and ERK activation by TNF-α. Previously, we showed that selective pharmacologic inhibition of the ERK and/or p38mapkpathway did not affect NO−2expression. Treatment of cells with the chloride channel blocker 5-nitro-2-(3-phenylpropylamino) benzoic acid (NPPB) showed a dose–response inhibition of NO−2expression. NPPB was also found to inhibit ERK and JNK/SAPK activation but not p38mapkwith TNF-α stimulation. The discordance between the marked degree of inhibition of iNOS transcript by IL-4 and the modest inhibition of JNK/SAPK and ERK suggests that the mechanism by which IL-4 inhibits iNOS transcription appears more complex than a mere inhibition of these MAPKs.