Cytokine Induction of Inducible Nitric Oxide Synthase in an Oligodendrocyte Cell Line: Role of P38 Mitogen-activated Protein Kinase Activation

The induction of inducible nitric oxide synthase (iNOS) by proinflammatory cytokines was studied in an oligodendrocyte progenitor cell line in relation to mitogen-activated protein kinase (MAPK) activation and cytokine-mediated cytotoxicity. When introduced individually to cultures of CG4 cells, the cytokines, i.e., tumor necrosis factor-alpha (TNF alpha), interleukin-1 (IL-1), and interferon-gamma (IFN gamma), had either minimal (TNF alpha) or no (IL-1 and IFN gamma) detectable stimulatory effect on the production of nitric oxide. However, combinations of these factors, in particular, TNF alpha plus IFN gamma, elicited a strong enhancement of nitric oxide synthesis and, as revealed by western blot and RT-PCR analysis, the expression of iNOS. TNF alpha and IL-1 were able to activate p38 MAPK in a time- and dose-dependent manner and together showed a combinatorial effect. In contrast, IFN gamma neither activated on its own nor enhanced the activation of p38 MAPK in response to TNF alpha and IL-1. However, a specific inhibitor of p38 MAPK, i.e., SB203580, inhibited the induction of iNOS in cytokine combination-treated cells in a dose-dependent manner, thereby suggesting a role for the MAPK cascade in regulating the induction of iNOS gene expression in cytokine-treated cells. Blocking of nitric oxide production by an inhibitor of iNOS, i.e., nitro-L-arginine methyl ester, had a minimal protective effect against cytokine-mediated cytotoxicity that occurred before the elevation of nitric oxide levels, thereby indicating temporal and functional dissociation of nitric oxide production from cell killing.