Activation of p38 MAPK induced peroxynitrite generation in LPS plus IFN-γ-stimulated rat primary astrocytes via activation of iNOS and NADPH oxidase

We have shown that immunostimulated astrocytes produce excess nitric oxide (NO) and eventually peroxynitrite (ONOO−) that was closely associated with the glucose deprivation-potentiated death of astrocytes. The present study shows that activated p38 MAPK regulates ONOO− generation from lipopolysaccharide (LPS) plus interferon-γ (IFN-γ)-stimulated astrocytes. LPS + IFN-γ-induced p38 MAPK activation and ONOO− generation were attenuated by SB203580 or SKF-86002, specific inhibitors of p38 MAPK. ONOO− generation was blocked by NADPH oxidase inhibitor, diphenyleneiodonium chloride, and nitric oxide synthase (NOS) inhibitor, Nω-nitro-l-arginine methyl ester, suggesting both enzymes are involved in ONOO− generation. Inhibition of p38 MAPK suppressed LPS + IFN-γ-induced NO production through down-regulating inducible form of NOS expression. It also suppressed LPS + IFN-γ-induced NADPH oxidase activation and eventually, the inducible form of superoxide production. Transfection with dominant negative vector of p38α reduced LPS + IFN-γ-induced ONOO− generation through blocking both iNOS-derived NO production and NADPH oxidase-derived O2− production. Our results suggest that activated p38 MAPK may serve as a potential signaling molecule in ONOO− generation through dual regulatory mechanisms, involving iNOS induction and NADPH oxidase activation.