Hy. Sohn et al., The small G-protein Rac mediates depolarization-induced superoxide formation in human endothelial cells, J BIOL CHEM, 275(25), 2000, pp. 18745-18750
Superoxide anions impair nitric oxide-mediated responses and are involved i
n the development of hypertensive vascular hypertrophy, The regulation of t
heir production in the vascular system is, however, poorly understood. We i
nvestigated whether changes in membrane potential that occur in hypertensiv
e vessels modulate endothelial superoxide production, In cultured human umb
ilical vein endothelial cells, changes in membrane potential were induced b
y high potassium buffer, the non-selective potassium channel blocker tetrab
utylammonium chloride (1 mM), and the non-selective cat ion ionophore grami
cidin (1 mu M). Superoxide formation was significantly elevated to a simila
r degree by all three treatments (by similar to 60%, n = 23, p < 0.01), whe
reas hyperpolarization by the K-ATP channel activator Hoe234 (1 mu M) signi
ficantly decreased superoxide formation. Depolarization also induced an inc
reased tyrosine phosphorylation of several not yet identified proteins (90-
110 kDa) and resulted in a significant increase in membrane association of
the small G-protein Rac. Accordingly, the Rac inhibitor Clostridium diffici
le toxin B blocked the effects of depolarization on superoxide formation. T
he tyrosine kinase inhibitor genistein (30 mu M, n = 15) abolished depolari
zation-induced superoxide formation and also prevented depolarization-induc
ed Rac translocation associated with it. It is concluded that depolarizatio
n is an important stimulus of endothelial superoxide production, which invo
lves a tyrosine phosphorylation-dependent translocation of the small G-prot
ein Rac.