Nox4 as the major catalytic component of an endothelial NAD(P)H oxidase.
BACKGROUND
Recent evidence has suggested that reactive oxygen species are important signaling molecules in vascular cells and play a pivotal role in the development of vascular diseases. The activity of NAD(P)H oxidase has been identified as the major source of reactive oxygen species in vascular endothelial cells. However, the precise molecular structure and the mechanism of activation of the oxidase have remained poorly understood.


RESULTS
Here, we investigated the molecular identities and the superoxide-producing activity of endothelial NAD(P)H oxidase. We found that Nox4, a homologue of gp91phox/Nox2, was abundantly expressed in endothelial cells. The expression of Nox4 in endothelial cells markedly exceeded that of other Nox proteins, including gp91phox/Nox2, and was affected by cell growth. Using electron spin resonance and chemiluminescence, we measured the superoxide production and found that the endothelial membranes had an NAD(P)H-dependent superoxide-producing activity comparable to that of the neutrophil membranes, whereas the activity was not enhanced by the 2 recombinant proteins p47phox and p67phox, in contrast to that of the neutrophil membranes. Downregulation of Nox4 by an antisense oligonucleotide reduced superoxide production in endothelial cells in vivo and in vitro.


CONCLUSIONS
These findings suggest that Nox4 may function as the major catalytic component of an endothelial NAD(P)H oxidase.
Ago
Tetsuro
T
Department of Medicine and Clinical Science, Graduate School of Medical Sciences, Kyushu University, Maidashi 3-1-1, Higashi-ku, Fukuoka 812-8582, Japan. agou@intmed2.med.kyushu-u.ac.jp
Kitazono
Takanari
T
Ooboshi
Hiroaki
H
Iyama
Teruaki
T
Han
Youn Hee
YH
Takada
Junichi
J
Wakisaka
Masanori
M
Ibayashi
Setsuro
S
Utsumi
Hideo
H
Iida
Mitsuo
M
D000818:Animals; D002384:Catalysis; D002462:Cell Membrane / Q000201:enzymology; D002478:Cells, Cultured; D004730:Endothelium, Vascular / Q000201:enzymology*; D005786:Gene Expression Regulation; D006801:Humans; D008297:Male; D000074663:NADPH Oxidase 4; D019255:NADPH Oxidases / Q000032:analysis* / Q000737:chemistry* / Q000378:metabolism* / Q000502:physiology; D010750:Phosphoproteins / Q000502:physiology; D021122:Protein Subunits / Q000032:analysis / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D013481:Superoxides / Q000378:metabolism
D010750:Phosphoproteins; D021122:Protein Subunits; D012333:RNA, Messenger; C079789:neutrophil cytosol factor 67K; D013481:Superoxides; D000074663:NADPH Oxidase 4; D019255:NADPH Oxidases; C498910:NOX4 protein, human; C498911:Nox4 protein, rat; C060505:neutrophil cytosolic factor 1