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Articles in PresS, published online ahead of print November 16, 2001
Am J Physiol Lung Cell Mol Physiol, 10.1152/ajplung.00206.2001
Submitted on June 6, 2001
Accepted on November 10, 2001
B Activation and Proliferation in Human Airway Smooth Muscle
1 Internal Medicine Research, Carolinas Medical Center, Charlotte, NC, USA
2 Division of Respiratory, Critical Care and Occupational (Pulmonary) Medicine, University of Utah, Salt Lake City, UT, USA
3 Montana State University, Veterinary Molecular Biology, Bozeman, MT, USA
4 Division of Pulmonary Diseases, Dept. of Pediatrics, Duke University Medical Center, Durham, NC, USA
* To whom correspondence should be addressed. E-mail: tkennedy{at}carolinas.org.
Evidence is rapidly accumulating that low-activity NAD(P)H oxidases homologous to that in phagocytic cells generate reactive oxygen species (ROS) 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. Proliferation of human airway smooth muscle (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, measured by SOD-inhibitable lucigenin chemiluminescence, with a distinct preference for NADH instead of NADPH as substrate. Chemiluminescence was also inhibited by DPI, suggesting the presence of a flavoprotein containing oxidase generating superoxide as a signaling molecule for cell growth. Examination of human AWSM cells by RT-PCR consistently demonstrated transcripts with sequences identical to those reported for p22phox. Transfection with p22phox antisense oligonucleotides reduced human AWSM proliferation. Inhibition of NAD(P)H oxidase activity with DPI prevented serum-induced activation of nuclear factor- B (NF-B), and over-expression of a superrepressor for of the NF-B inhibitor IB-
significantly reduced human AWSM growth. These findings suggest that an NAD(P)H 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-B.
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