Constitutive NADPH oxidase and increased mitochondrial respiratory chain activity regulate chemokine gene expression.
Alveolar macrophages, which generate high levels of reactive oxygen species, especially O(2)(*-), are involved in the recruitment of neutrophils to sites of inflammation and injury in the lung, and the generation of chemotactic proteins triggers this cellular recruitment. In this study, we asked whether O(2)(*-) generation in alveolar macrophages had a role in the expression of chemokines. Specifically, we hypothesized that O(2)(*-) generation is necessary for chemokine expression in alveolar macrophages after TNF-alpha stimulation. We found that alveolar macrophages have high constitutive NADPH oxidase activity that was not increased by TNF-alpha, but TNF-alpha increased the activity of the mitochondrial respiratory chain. In addition, the mitochondrial respiratory chain increased O(2)(*-) generation if the NADPH oxidase was inhibited. O(2)(*-) generation was necessary for macrophage inflammatory protein-2 (MIP-2) gene expression, because inhibition of NADPH oxidase or the mitochondrial respiratory chain or overexpression of Cu,Zn-superoxide dismutase significantly inhibited expression of MIP-2. TNF-alpha activated the ERK MAP kinase, and ERK activity was essential for chemokine gene expression. In addition, overexpression of the MEK1-->ERK pathway significantly increased IL-8 expression, and a small interfering RNA to the NADPH oxidase inhibited ERK- and TNF-alpha-induced chemokine expression. Collectively, these results suggest that in alveolar macrophages, O(2)(*-) generation mediates chemokine expression after TNF-alpha stimulation in an ERK-dependent manner.
Tephly
Linda A
LA
Division of Pulmonary, University of Iowa Roy J. and Lucille A. Carver College of Medicine, Iowa City, IA 52242, USA.
Carter
A Brent
AB
D015153:Blotting, Western; D054426:Chemokine CXCL2 / Q000378:metabolism*; D018925:Chemokines / Q000235:genetics*; D004579:Electron Transport; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism; D005786:Gene Expression Regulation / Q000502:physiology*; D006801:Humans; D006861:Hydrogen Peroxide / Q000494:pharmacology; D016209:Interleukin-8 / Q000378:metabolism; D048369:MAP Kinase Kinase 1 / Q000378:metabolism; D016676:Macrophages, Alveolar / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D008928:Mitochondria / Q000378:metabolism*; D019255:NADPH Oxidases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D016877:Oxidants / Q000494:pharmacology; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D034741:RNA, Small Interfering / Q000494:pharmacology; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013482:Superoxide Dismutase / Q000378:metabolism; D013481:Superoxides / Q000378:metabolism; D014158:Transcription, Genetic; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology
D054426:Chemokine CXCL2; D018925:Chemokines; D016209:Interleukin-8; D016877:Oxidants; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D014409:Tumor Necrosis Factor-alpha; D013481:Superoxides; D006861:Hydrogen Peroxide; D013482:Superoxide Dismutase; D019255:NADPH Oxidases; D048049:Extracellular Signal-Regulated MAP Kinases; D048369:MAP Kinase Kinase 1; C482125:MAP2K1 protein, human