Activation of endothelial NADPH oxidase during normoxic lung ischemia is KATP channel dependent.
Previous studies have shown endothelial cell membrane depolarization and generation of reactive oxygen species (ROS) in endothelial cells with abrupt reduction in shear stress (ischemia). This study evaluated the role of ATP-sensitive potassium (K(ATP)) channels and NADPH oxidase in the ischemic response by using Kir6.2-/- and gp91(phox)-/- mice. To evaluate ROS generation, we subjected isolated perfused mouse lungs labeled with 2',7'-dichlorodihydrofluorescein (DCF), hydroethidine (HE), or diphenyl-1-pyrenylphosphine (DPPP) to control perfusion followed by global ischemia. In wild-type C57BL/6J mice, imaging of subpleural endothelial cells showed a time-dependent increase in intensity for all three fluorescence probes with ischemia, which was blocked by preperfusion with cromakalim (a K(ATP) channel agonist) or diphenyleneiodonium (DPI, a flavoprotein inhibitor). Endothelial cell fluorescence with bis-oxonol, a membrane potential probe, increased during lung ischemia indicating cell membrane depolarization. The change in membrane potential with ischemia in lungs of gp91(phox)-/- mice was similar to wild type, but ROS generation did not occur. Lungs from Kir6.2-/- showed marked attenuation of the change in both membrane potential and ROS production. Thus membrane depolarization during lung ischemia requires the presence of a K(ATP) channel and is required for activation of NADPH oxidase and endothelial ROS generation.
Zhang
Qunwei
Q
Inst. for Environmental Medicine, University of Pennsylvania School of Medicine, 1 John Morgan Bldg., 3620 Hamilton Walk, Philadelphia, PA 19104-6068, USA.
Matsuzaki
Ikuo
I
Chatterjee
Shampa
S
Fisher
Aron B
AB
D000818:Animals; D019806:Cromakalim / Q000494:pharmacology; D042783:Endothelial Cells / Q000201:enzymology* / Q000473:pathology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005456:Fluorescent Dyes / Q000494:pharmacology; D007511:Ischemia / Q000201:enzymology*; D008168:Lung / Q000098:blood supply* / Q000378:metabolism* / Q000473:pathology; D008562:Membrane Glycoproteins / Q000235:genetics / Q000378:metabolism; D008564:Membrane Potentials / Q000187:drug effects; D051379:Mice; D018345:Mice, Knockout; D000074662:NADPH Oxidase 2; D019255:NADPH Oxidases / Q000235:genetics / Q000378:metabolism*; D009861:Onium Compounds / Q000494:pharmacology; D010994:Pleura / Q000098:blood supply / Q000378:metabolism / Q000473:pathology; D024661:Potassium Channels, Inwardly Rectifying / Q000819:agonists / Q000235:genetics / Q000378:metabolism*; D017382:Reactive Oxygen Species / Q000378:metabolism; D014665:Vasodilator Agents / Q000494:pharmacology; D000099161:Kcnj11 Channel
D019806:Cromakalim; D004791:Enzyme Inhibitors; D005456:Fluorescent Dyes; D008562:Membrane Glycoproteins; D000074662:NADPH Oxidase 2; D019255:NADPH Oxidases; D009861:Onium Compounds; D024661:Potassium Channels, Inwardly Rectifying; D017382:Reactive Oxygen Species; D014665:Vasodilator Agents; D000099161:Kcnj11 Channel; C007517:diphenyleneiodonium; C488842:Cybb protein, mouse