Urokinase plasminogen activator stimulates vascular smooth muscle cell proliferation via redox-dependent pathways.
OBJECTIVE
We showed previously that increased urokinase plasminogen activator (uPA) expression contributes to vascular smooth muscle cell (VSMC) proliferation and neointima formation after injury. Proliferation of cultured rat aortic VSMCs induced by uPA was inhibited by the antioxidant ebselen. Because increases in VSMC reactive oxygen species (ROS) contribute to VSMC proliferation, we hypothesized that uPA increases ROS generation by regulating expression or activity of cellular oxidases.


RESULTS
uPA stimulated ROS production to levels equivalent to angiotensin II as measured by electron spin resonance and fluorescent redox indicators (dichlorofluorescein diacetate, lucigenin, and hydroethidine). The increase in ROS was biphasic, with the first peak at 30 minutes and the second peak at 4 hours. uPA increased expression of the NAD(P)H oxidases Nox1 and Nox4 as measured by RT-PCR and Western blot analysis. Knockdown of Nox1 and Nox4 expression with small interfering RNA showed that both isoforms (Nox1>Nox4) contributed significantly to uPA-stimulated ROS production and VSMC proliferation. Transfection of VSMCs with uPA cDNA to increase endogenous uPA expression enhanced ROS production dramatically, suggesting that autocrine uPA production may be an important mechanism for uPA-mediated VSMC events.


CONCLUSIONS
These data show that uPA is an autocrine VSMC growth factor that increases ROS generated by both Nox1 and Nox4 oxidases.
Menshikov
Mikhail
M
Cardiology Research Center, Moscow, Russia.
Plekhanova
Olga
O
Cai
Hua
H
Chalupsky
Karel
K
Parfyonova
Yelena
Y
Bashtrikov
Pavel
P
Tkachuk
Vsevolod
V
Berk
Bradford C
BC
D000804:Angiotensin II / Q000378:metabolism; D000818:Animals; D001011:Aorta / Q000166:cytology / Q000502:physiology; D049109:Cell Proliferation; D002478:Cells, Cultured; D008297:Male; D009131:Muscle, Smooth, Vascular / Q000166:cytology / Q000502:physiology*; D009247:NADH, NADPH Oxidoreductases / Q000037:antagonists & inhibitors / Q000378:metabolism; D000074624:NADPH Oxidase 1; D000074663:NADPH Oxidase 4; D019255:NADPH Oxidases / Q000037:antagonists & inhibitors / Q000378:metabolism; D010084:Oxidation-Reduction; D051381:Rats; D017207:Rats, Sprague-Dawley; D017382:Reactive Oxygen Species / Q000378:metabolism; D015398:Signal Transduction; D013997:Time Factors; D014568:Urokinase-Type Plasminogen Activator / Q000235:genetics / Q000378:metabolism*
D017382:Reactive Oxygen Species; D000804:Angiotensin II; D009247:NADH, NADPH Oxidoreductases; D000074624:NADPH Oxidase 1; D000074663:NADPH Oxidase 4; D019255:NADPH Oxidases; C000614712:NOX1 protein, rat; C498911:Nox4 protein, rat; D014568:Urokinase-Type Plasminogen Activator