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Am J Physiol Lung Cell Mol Physiol 282: L782-L795, 2002. First published November 16, 2001; doi:10.1152/ajplung.00206.2001
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Vol. 282, Issue 4, L782-L795, April 2002

NADPH oxidase promotes NF-kappa B activation and proliferation in human airway smooth muscle

Sukhdev S. Brar1, Thomas P. Kennedy1, Anne B. Sturrock2, Thomas P. Huecksteadt2, Mark T. Quinn3, Thomas M. Murphy4, Pasquale Chitano4, and John R. Hoidal2

1 Department of Internal Medicine, Cannon Research Center, Carolinas Medical Center, Charlotte 28232; 4 Division of Pulmonary Diseases, Department of Pediatrics, Duke University Medical Center, Durham, North Carolina 27710; 2 Division of Respiratory, Critical Care and Occupational (Pulmonary) Medicine, University of Utah, Salt Lake City, Utah 84132; and 3 Department of Veterinary Molecular Biology, Montana State University, Bozeman, Montana 59717

Evidence is rapidly accumulating that low-activity-reduced nicotinamide adenine dinucleotide phosphate (NADPH) oxidases homologous to that in phagocytic cells generate reactive oxygen species as signaling intermediates in both endothelium and vascular smooth muscle. We therefore explored the possibility of such an oxidase regulating growth of airway smooth muscle (AWSM). Proliferation of human AWSM cells in culture was inhibited by the antioxidants catalase and N-acetylcysteine, and by the flavoprotein inhibitor diphenylene iodonium (DPI). Membranes prepared from human AWSM cells generated superoxide anion (O<UP><SUB>2</SUB><SUP>−</SUP></UP>) measured by superoxide dismutase-inhibitable lucigenin chemiluminescence, with a distinct preference for NADPH instead of reduced nicotinamide adenine dinucleotide as substrate. Chemiluminescence was also inhibited by DPI, suggesting the presence of a flavoprotein containing oxidase generating O<UP><SUB>2</SUB><SUP>−</SUP></UP> as a signaling molecule for cell growth. Examination of human AWSM cells by reverse transcriptase-polymerase chain reaction consistently demonstrated transcripts with sequences identical to those reported for p22phox. Transfection with p22phox antisense oligonucleotides reduced human AWSM proliferation. Inhibition of NADPH oxidase activity with DPI prevented serum-induced activation of nuclear factor-kappa B (NF-kappa B), and overexpression of a superrepressor form of the NF-kappa B inhibitor Ikappa Balpha significantly reduced human AWSM growth. These findings suggest that an NADPH oxidase containing p22phox regulates growth-factor responsive human AWSM proliferation, and that the oxidase signals in part through activation of the prototypical redox-regulated transcription factor NF-kappa B.

superoxide anion; diphenylene iodonium; p22phox


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