NAD(P)H oxidases in rat basilar arterial endothelial cells.
OBJECTIVE
Reactive oxygen species (ROS) may play a critical role in the regulation of vascular tone and development of vascular diseases, such as stroke. NAD(P)H oxidase is a major source of ROS in vascular cells, including endothelial cells. It has been considered that Nox2 and Nox4 are exclusively expressed among Nox homologues in the endothelial cells of noncerebral blood vessels. However, the precise molecular identity of the NAD(P)H oxidase in the endothelial cells of the cerebral arteries is not fully understood. We examined the expression of Nox homologues and their activation mechanism in the endothelial cells of the cerebral arteries.


METHODS
We isolated and cultured basilar artery endothelial cells (BAECs) of Sprague-Dawley rats. Expression of NAD(P)H oxidase was examined by reverse-transcription-polymerase chain reaction (RT-PCR) and immunohistological staining.


RESULTS
RT-PCR disclosed abundant expression of Nox4 with marginal Nox2 in BAEC. In addition, Nox1 was expressed highly both at mRNA and protein levels in BAECs. Immunohistological staining also showed the prominent expression of Nox1 in the endothelial cells of the basilar artery. With respect to the cytosolic components of NAD(P)H oxidases, BAECs expressed p67phox and, to a lesser extent, p47phox, Noxo1, and Noxa1. Both NADH and NADPH induced superoxide production of the BAEC membranes. The phagocyte-type cytosolic components, p47phox and p67phox, significantly enhanced the NADH-induced superoxide production of the BAEC membranes, whereas the components failed to increase the NADPH-induced superoxide production.


CONCLUSIONS
Nox1 is highly expressed in the endothelial cells of the cerebral arteries along with Nox2 and Nox4, and the endothelial NAD(P)H oxidase of the cerebral arteries may have a unique activation mechanism by the phagocyte-type cytosolic components.
Ago
Tetsuro
T
Department of Medicine and Clinical Science, Graduate School of Medical Sciences, Kyushu University, Fukuoka, Japan. agou@intmed2.med.kyushu-u.ac.jp
Kitazono
Takanari
T
Kuroda
Junya
J
Kumai
Yasuhiro
Y
Kamouchi
Masahiro
M
Ooboshi
Hiroaki
H
Wakisaka
Masanori
M
Kawahara
Tsukasa
T
Rokutan
Kazuhito
K
Ibayashi
Setsuro
S
Iida
Mitsuo
M
D000818:Animals; D001488:Basilar Artery / Q000166:cytology / Q000201:enzymology*; D042783:Endothelial Cells / Q000166:cytology / Q000201:enzymology; D004730:Endothelium, Vascular / Q000166:cytology / Q000201:enzymology*; D008297:Male; D009247:NADH, NADPH Oxidoreductases / Q000235:genetics / Q000378:metabolism; D000074624:NADPH Oxidase 1; D019255:NADPH Oxidases / Q000378:metabolism*; D010750:Phosphoproteins / Q000096:biosynthesis / Q000235:genetics; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D013481:Superoxides / Q000378:metabolism
D010750:Phosphoproteins; D012333:RNA, Messenger; C079789:neutrophil cytosol factor 67K; D013481:Superoxides; D009247:NADH, NADPH Oxidoreductases; D000074624:NADPH Oxidase 1; D019255:NADPH Oxidases; C000614712:NOX1 protein, rat; C060505:neutrophil cytosolic factor 1