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Am J Physiol Lung Cell Mol Physiol 286: L37-L48, 2004. First published June 13, 2003; doi:10.1152/ajplung.00116.2003
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Oxidant Signaling in Lung Cells

NAD(P)H oxidase mediates the endothelial barrier dysfunction induced by TNF-{alpha}

Nancy Gertzberg,1 Paul Neumann,1 Victor Rizzo,1 and Arnold Johnson1,2

1Center for Cardiovascular Science, Albany Medical College, and 2Research Service, Stratton Veterans Affairs Medical Center, Albany, New York 1228

Submitted 16 April 2003 ; accepted in final form 5 June 2003

We tested the hypothesis that the NAD(P)H oxidase-dependent generation of superoxide anion () mediates tumor necrosis factor-{alpha} (TNF)-induced alterations in the permeability of pulmonary microvessel endothelial monolayers (PMEM). The permeability of PMEM was assessed by the clearance rate of Evans blue-labeled albumin. The NAD(P)H oxidase subcomponents p47phox and p22phox were assessed by immunofluorescent microscopy and Western blot. The reactive oxygen species was measured by the fluorescence of 6-carboxy-2',7'-dichlorodihydrofluorescein diacetatedi(acetoxymethyl ester), 5 (and 6)-chloromethyl-2',7'-dichlorodihydrofluorescein diacetate-acetyl ester, and dihydroethidium. TNF treatment (50 ng/ml for 4.0 h) induced 1) p47phox translocation, 2) an increase in p22phox protein, 3) increased localization of p47phox with p22phox, 4) generation, and 5) increased permeability to albumin. p22phox antisense oligonucleotide prevented the TNF-induced effect on p22phox, p47phox, , and permeability. The scrambled nonsense oligonucleotide had no effect. The TNF-induced increase in and permeability to albumin was also prevented by the scavenger Cu-Zn superoxide dismutase (100 U/ml). The results indicate that the activation of NAD(P)H oxidase, via the generation of , mediates TNF-induced barrier dysfunction in PMEM.

antisense; edema; mRNA; permeability; transcription


Address for reprint requests and other correspondence: A. Johnson, Mail-stop 151, 113 Holland Ave., Stratton VA Medical Center, Albany, NY 12208 (E-mail: jmurd{at}msn.com).




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