Long-term cyclic strain downregulates endothelial Nox4.
Endothelial cells in vivo are constantly exposed to mechanical forces such as cyclic strain. In endothelial cells, Nox4-containing NAD(P)H oxidase complexes have been identified as major sources of superoxide anion (.O(2)(-)) formation. In this study, we analyzed the effect of cyclic strain on endothelial ROS formation by electron paramagnetic resonance spectroscopy, cytochrome c assay, and dihydroethidium fluorescence, on NO formation by Griess reaction and on gene expression by RT-PCR and Western blot. Primary cultures of human umbilical vein endothelial cells were exposed to 2-18% cyclic strain for up to 24 h using the Flexercell system. Long-term application of 5-12% cyclic strain downregulated Nox4 expression and ROS formation in a time-dependent manner. Downregulation of Nox4 was further confirmed by promoter analysis using dual-luciferase assay. Cu/Zn SOD, MnSOD, and catalase expression was decreased after application of chronic 12% cyclic strain. In contrast, endothelial NO formation and eNOS were increased by cyclic strain. Strain-dependent Nox4 downregulation was abolished by eNOS inhibition with L-NAME. In conclusion, physiological levels of cyclic strain downregulate Nox4 expression and superoxide anion formation. This novel mechanism might contribute to a vasoprotective balance between NO and superoxide anions in response to physiological mechanical stimulation of endothelial cells.
Goettsch
Claudia
C
Division of Vascular Endothelium and Microcirculation, Department of Medicine III, University of Technology Dresden, D-01307 Dresden, Germany.
Goettsch
Winfried
W
Arsov
Alexander
A
Hofbauer
Lorenz C
LC
Bornstein
Stefan R
SR
Morawietz
Henning
H
D000975:Antioxidants / Q000378:metabolism; D002460:Cell Line; D042783:Endothelial Cells / Q000166:cytology / Q000201:enzymology*; D004730:Endothelium, Vascular / Q000166:cytology / Q000201:enzymology; D004791:Enzyme Inhibitors / Q000378:metabolism; D006801:Humans; D007527:Isoenzymes / Q000235:genetics / Q000378:metabolism; D000074663:NADPH Oxidase 4; D019255:NADPH Oxidases / Q000235:genetics / Q000378:metabolism*; D019331:NG-Nitroarginine Methyl Ester / Q000378:metabolism; D009569:Nitric Oxide / Q000378:metabolism; D011401:Promoter Regions, Genetic; D017382:Reactive Oxygen Species / Q000378:metabolism; D013314:Stress, Mechanical; D013482:Superoxide Dismutase / Q000235:genetics / Q000378:metabolism; D013481:Superoxides / Q000378:metabolism
D000975:Antioxidants; D004791:Enzyme Inhibitors; D007527:Isoenzymes; D017382:Reactive Oxygen Species; D013481:Superoxides; D009569:Nitric Oxide; D013482:Superoxide Dismutase; D000074663:NADPH Oxidase 4; D019255:NADPH Oxidases; C498910:NOX4 protein, human; D019331:NG-Nitroarginine Methyl Ester