Control of JNK for an activation of NADPH oxidase in LPS-stimulated BV2 microglia

Abstract

NADPH oxidase is a main regulator for H2O2 productivity in neuroinflammatory cells, including microglia, under various CNS diseases and its activity is controlled by mitogen-activated protein kinases (MAPKs), such as extracellular signal-regulated kinase 1/2 (ERK1/2), p38 MAPK, and c-Jun N-terminal kinase (JNK). However, little is known about the link between NADPH oxidase-driven H2O2 productivity and JNK in microglia. The purpose of this study is to uncover the link using lipopolysaccharide (LPS)-stimulated BV2 microglia. LPS-stimulated BV2 microglia produced H2O2 that was decreased by NADPH oxidase inhibitors, including 4-(2-aminoethyl)benzenesulfonylfluoride and diphenyleneiodonium chloride. In addition, NADPH oxidase was activated in LPS-stimulated BV2 cells. These results suggest that NAPDH oxidase is a main factor for H2O2 productivity in LPS-stimulated BV2 microglia. Based on a semi-quantitative PCR analysis, two of NADPH oxidase components, p47(phox) and gp91(phox), were involved in the activation of NADPH oxidase because transcriptional levels of both components were upregulated by LPS. Role of JNK in NADPH oxidase-regulated H2O2 productivity was pursued using specific inhibitors, including SP600125 and JNK inhibitory peptide (JIP). Inhibition of the JNK pathways significantly reduced H2O2 productivity, which was closely related to the attenuation of NADPH oxidase activation and the upregulation of components. We conclude that JNK pathways are involved in NADPH oxidase-mediated H2O2 productivity in BV2 microglia.