TY - JOUR
T1 - Novel role of gp91phox-containing NAD(P)H oxidase in vascular endothelial growth factor-induced signaling and angiogenesis
AU - Ushio-Fukai, Masuko
AU - Tang, Yan
AU - Fukai, Tohru
AU - Dikalov, Sergey I.
AU - Ma, Yuxian
AU - Fujimoto, Mitsuaki
AU - Quinn, Mark T.
AU - Pagano, Patrick J.
AU - Johnson, Chad
AU - Alexander, R. Wayne
PY - 2002/12
Y1 - 2002/12
N2 - Vascular endothelial growth factor (VEGF) induces angiogenesis by stimulating endothelial cell proliferation and migration, primarily through the receptor tyrosine kinase VEGF receptor2 (Flk1/KDR). Reactive oxygen species (ROS) derived from NAD(P)H oxidase are critically important in many aspects of vascular cell regulation, and both the small GTPase Rac1 and gp91phox are critical components of the endothelial NAD(P)H oxidase complex. A role of NAD(P)H oxidase in VEGF-induced angiogenesis, however, has not been defined. In the present study, electron spin resonance spectroscopy is utilized to demonstrate that VEGF stimulates O2.- production, which is inhibited by the NAD(P)H oxidase inhibitor, diphenylene iodonium, as well as by overexpression of dominant-negative Rac1 (N17Rac1) and transfection of gp91phox antisense oligonucleotides in human umbilical vein endothelial cells (ECs). Antioxidants, including N-acetylcysteine (NAC), various NAD(P)H oxidase inhibitors, and N17Rac1 significantly attenuate not only VEGF-induced KDR tyrosine phosphorylation but also proliferation and migration of ECs. Importantly, these effects of VEGF are dramatically inhibited in cells transfected with gp91phox antisense oligonucleotides. By contrast, ROS are not involved in mediating these effects of sphingosine 1-phosphate (SIP) on ECs. Sponge implant assays demonstrate that VEGF-, but not S1P-, induced angiogenesis is significantly reduced in wild-type mice treated with NAC and in gp91phox-/- mice, suggesting that ROS derived from gp91phox-containing NAD(P)H oxidase play an important role in angiogenesis in vivo. These studies indicate that VEGF-induced endothelial cell signaling and angiogenesis is tightly controlled by the reduction/oxidation environment at the level of VEGF receptor and provide novel insights into the NAD(P)H oxidase as a potential therapeutic target for angiogenesis-dependent diseases.
AB - Vascular endothelial growth factor (VEGF) induces angiogenesis by stimulating endothelial cell proliferation and migration, primarily through the receptor tyrosine kinase VEGF receptor2 (Flk1/KDR). Reactive oxygen species (ROS) derived from NAD(P)H oxidase are critically important in many aspects of vascular cell regulation, and both the small GTPase Rac1 and gp91phox are critical components of the endothelial NAD(P)H oxidase complex. A role of NAD(P)H oxidase in VEGF-induced angiogenesis, however, has not been defined. In the present study, electron spin resonance spectroscopy is utilized to demonstrate that VEGF stimulates O2.- production, which is inhibited by the NAD(P)H oxidase inhibitor, diphenylene iodonium, as well as by overexpression of dominant-negative Rac1 (N17Rac1) and transfection of gp91phox antisense oligonucleotides in human umbilical vein endothelial cells (ECs). Antioxidants, including N-acetylcysteine (NAC), various NAD(P)H oxidase inhibitors, and N17Rac1 significantly attenuate not only VEGF-induced KDR tyrosine phosphorylation but also proliferation and migration of ECs. Importantly, these effects of VEGF are dramatically inhibited in cells transfected with gp91phox antisense oligonucleotides. By contrast, ROS are not involved in mediating these effects of sphingosine 1-phosphate (SIP) on ECs. Sponge implant assays demonstrate that VEGF-, but not S1P-, induced angiogenesis is significantly reduced in wild-type mice treated with NAC and in gp91phox-/- mice, suggesting that ROS derived from gp91phox-containing NAD(P)H oxidase play an important role in angiogenesis in vivo. These studies indicate that VEGF-induced endothelial cell signaling and angiogenesis is tightly controlled by the reduction/oxidation environment at the level of VEGF receptor and provide novel insights into the NAD(P)H oxidase as a potential therapeutic target for angiogenesis-dependent diseases.
KW - Angiogenesis
KW - Endothelial cells
KW - NAD(P)H oxidase
KW - Reactive oxygen species
KW - Vascular endothelial growth factor
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U2 - 10.1161/01.RES.0000046227.65158.F8
DO - 10.1161/01.RES.0000046227.65158.F8
M3 - Article
C2 - 12480817
AN - SCOPUS:0036954803
SN - 0009-7330
VL - 91
SP - 1160
EP - 1167
JO - Circulation research
JF - Circulation research
IS - 12
ER -