NADPH Oxidase Activity Selectively Modulates Vascular Endothelial Growth Factor Signaling Pathways

Katherine C Spokes,Md Ruhul Abid,William C Aird,Shou-Ching Shih

doi:10.1074/jbc.m702175200

Katherine C Spokes, Md Ruhul Abid + Show 2 more

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https://doi.org/10.1074/jbc.m702175200

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Abstract

Vascular endothelial growth factor (VEGF) and reactive oxygen species (ROS) play critical roles in vascular physiology and pathophysiology. We have demonstrated previously that NADPH oxidase-derived ROS are required for VEGF-mediated migration and proliferation of endothelial cells. The goal of this study was to determine the extent to which VEGF signaling is coupled to NADPH oxidase activity. Human umbilical vein endothelial cells and/or human coronary artery endothelial cells were transfected with short interfering RNA against the p47(phox) subunit of NADPH oxidase, treated in the absence or presence of VEGF, and assayed for signaling, gene expression, and function. We show that NADPH oxidase activity is required for VEGF activation of phosphoinositide 3-kinase-Akt-forkhead, and p38 MAPK, but not ERK1/2 or JNK. The permissive role of NADPH oxidase on phosphoinositide 3-kinase-Akt-forkhead signaling is mediated at post-VEGF receptor levels and involves the nonreceptor tyrosine kinase Src. DNA microarrays revealed the existence of two distinct classes of VEGF-responsive genes, one that is ROS-dependent and another that is independent of ROS levels. VEGF-induced, thrombomodulin-dependent activation of protein C was dependent on NADPH oxidase activity, whereas VEGF-induced decay-accelerating factor-mediated protection of endothelial cells against complement-mediated lysis was not. Taken together, these findings suggest that NADPH oxidase-derived ROS selectively modulate some but not all the effects of VEGF on endothelial cell phenotypes.

Highlights

Has been implicated in wound repair, angiogenesis of ischemic tissue, tumor growth, microvascular permeability, vascular protection, and hemostasis [1,2,3,4,5,6,7,8]
We previously reported that Vascular endothelial growth factor (VEGF) induces NADPH oxidase activity and that NADPH oxidase activity is required for VEGFmediated migration and proliferation of endothelial cells [21, 22]
NADPH Oxidase Activity Is Abrogated by Down-regulation of p47phox Expression in Primary Human Endothelial Cells—To inhibit NADPH oxidase in endothelial cells, HCAEC or human umbilical vein endothelial cells (HUVEC) were transfected with antisense or siRNA against p47phox

Summary

Introduction

Has been implicated in wound repair, angiogenesis of ischemic tissue, tumor growth, microvascular permeability, vascular protection, and hemostasis [1,2,3,4,5,6,7,8]. We previously reported that VEGF induces NADPH oxidase activity and that NADPH oxidase activity is required for VEGFmediated migration and proliferation of endothelial cells [21, 22] These results were confirmed and expanded upon by other groups. We showed that NADPH oxidase-derived ROS are required for VEGF stimulation of manganese superoxide dismutase, activation of NF-␬B, and inactivation of FKHR in endothelial cells [44]. We show that NADPH oxidase activity is required for some, but not all, downstream effects of VEGF in endothelial cells These data argue against a global sensitivity of VEGF signal transduction pathways to the redox state of the cell, and suggest that therapeutic modulation of ROS will selectively influence the effect of VEGF on endothelial cell phenotypes

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