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(Arteriosclerosis, Thrombosis, and Vascular Biology. 2007;27:2319.)
© 2007 American Heart Association, Inc.

Vascular Biology

Important Role of Nox4 Type NADPH Oxidase in Angiogenic Responses in Human Microvascular Endothelial Cells In Vitro

Srinivasa Raju Datla; Hitesh Peshavariya; Gregory J. Dusting; Kalyankar Mahadev; Barry J. Goldstein; Fan Jiang

From the Bernard O’Brien Institute of Microsurgery, University of Melbourne, Victoria, Australia.

Correspondence to Fan Jiang, Bernard O’Brien Institute of Microsurgery, 42 Fitzroy Street, Fitzroy, Victoria 3065, Australia. E-mail fjiang{at}unimelb.edu.au

Objective— Redox signaling mediated by Nox2-containing NADPH oxidase has been implicated in angiogenic responses both in vitro and in vivo. Because Nox4 type NADPH oxidase is also highly expressed in endothelial cells, we studied the role of Nox4 in angiogenic responses in human endothelial cells in culture.

Methods and Results— Inhibition of Nox4 expression by small interfering RNA reduced angiogenic responses as assessed by the tube formation and wound healing assays, in both human microvascular and umbilical vein endothelial cells. Overexpression of wild-type Nox4 enhanced, whereas expression of a dominant negative form of Nox4 suppressed the angiogenic responses in endothelial cells. These effects were mimicked by exogenous H₂O₂ and the antioxidant compound ebselen, respectively. Overexpression of Nox4 enhanced receptor tyrosine kinase phosphorylation and the activation of extracellular signal-regulated kinase (Erk). Inhibition of the Erk pathway reduced the endothelial angiogenic responses. Nox4 expression also promotes proliferation and migration of endothelial cells, and reduced serum deprivation–induced apoptosis.

Conclusions— Nox4 type NADPH oxidase promotes endothelial angiogenic responses, at least partly, via enhanced activation of receptor tyrosine kinases and the downstream Erk pathway.

In vitro angiogenic responses were inhibited by Nox4 small interfering RNA or by dominant negative Nox4. Wild-type Nox4 overexpression enhanced VEGF-stimulated signaling, reduced apoptosis, and enhanced the angiogenic responses. Nox4-dependent redox signaling is an important positive modulator of angiogenic responses in human vascular endothelial cells.

Key Words: angiogenesis • endothelial cell • NADPH oxidase • Nox4 • redox signaling

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