Submitted on March 23, 2007
Accepted on August 15, 2007
CONSTITUTIVE NADPH OXIDASE AND INCREASED MITOCHONDRIAL RESPIRATORY CHAIN ACTIVITY REGULATE CHEMOKINE GENE EXPRESSION
Linda A. Tephly1 and A. Brent Carter1*
1 Internal Medicine, University of Iowa, Iowa City, Iowa, United States
* To whom correspondence should be addressed. E-mail: brent-carter{at}uiowa.edu.
Alveolar macrophages, which generate high levels of ROS, especially O2.-, 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 ask if O2.- generation in alveolar macrophages had a role in the expression of chemokines. Specifically, we hypothesized that O2.- generation is necessary for chemokine expression in alveolar macrophages after TNF-
stimulation. We found that alveolar macrophages have high constitutive NADPH oxidase activity that was not increased by TNF-, but TNF- increased the activity of the mitochondrial respiratory chain. In addition, the mitochondrial respiratory chain increased O2.- generation if the NADPH oxidase was inhibited. O2.- generation was necessary for MIP-2 gene expression because inhibition of the NADPH oxidase, the mitochondrial respiratory chain, or over expression of Cu,Zn-SOD significantly inhibited expression of MIP-2. TNF- activated the ERK MAP kinase, and ERK activity was essential for chemokine gene expression. In addition, over expression of the MEK1
ERK pathway significantly increased IL-8 expression, and a small interfering RNA to the NADPH oxidase inhibited ERK and TNF--induced chemokine expression. Collectively, these results suggest that in alveolar macrophages, O2.- generation mediates chemokine expression after TNF- stimulation in an ERK-dependent manner.
