Membrane depolarization and NADPH oxidase activation in aortic endothelium during ischemia reflect altered mechanotransduction.
We previously showed that "ischemia" (abrupt cessation of flow) leads to rapid membrane depolarization and increased generation of reactive oxygen species (ROS) in lung microvascular endothelial cells. This response is not associated with anoxia but, rather, reflects loss of normal shear stress. This study evaluated whether a similar response occurs in aortic endothelium. Plasma membrane potential and production of ROS were determined by fluorescence microscopy and cytochrome c reduction in flow-adapted rat or mouse aorta or monolayer cultures of rat aortic endothelial cells. Within 30 s after flow cessation, endothelial cells that had been flow adapted showed plasma membrane depolarization that was inhibited by pretreatment with cromakalim, an ATP-sensitive K(+) (K(ATP)) channel agonist. Flow cessation also led to ROS generation, which was inhibited by cromakalim and the flavoprotein inhibitor diphenyleneiodonium. Aortic endothelium from mice with "knockout" of the K(ATP) channel (K(IR)6.2) showed a markedly attenuated change in membrane potential and ROS generation with flow cessation. In aortic endothelium from mice with knockout of NADPH oxidase (gp91(phox)), membrane depolarization was similar to that in wild-type mice but ROS generation was absent. Thus rat and mouse aortic endothelial cells respond to abrupt flow cessation by K(ATP) channel-mediated membrane depolarization followed by NADPH oxidase-mediated ROS generation, possibly representing a cell-signaling response to altered mechanotransduction.
Institute for Environmental Medicine, University of Pennsylvania School of Medicine, 1 John Morgan Bldg., Philadelphia, PA 19104-6068, USA.
Ikuo
I
Matsuzaki
Shampa
S
Chatterjee
Kris
K
Debolt
Yefim
Y
Manevich
Qunwei
Q
Zhang
Aron B
AB
Fisher
D000818:Animals; D001011:Aorta / Q000201:enzymology / Q000503:physiopathology*; D002462:Cell Membrane / Q000378:metabolism; D002478:Cells, Cultured; D045304:Cytochromes c / Q000378:metabolism; D042783:Endothelial Cells / Q000201:enzymology; D004730:Endothelium, Vascular / Q000201:enzymology / Q000503:physiopathology*; D004789:Enzyme Activation; D007511:Ischemia / Q000201:enzymology / Q000503:physiopathology*; D008297:Male; D040542:Mechanotransduction, Cellular; D008564:Membrane Potentials; D051379:Mice; D018345:Mice, Knockout; D019255:NADPH Oxidases / Q000378:metabolism*; D024661:Potassium Channels, Inwardly Rectifying / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D017382:Reactive Oxygen Species / Q000378:metabolism; D013314:Stress, Mechanical; D000099161:Kcnj11 Channel
D045304:Cytochromes c; D019255:NADPH Oxidases; D024661:Potassium Channels, Inwardly Rectifying; D017382:Reactive Oxygen Species; D000099161:Kcnj11 Channel