Published Online
on February 24, 2003

A more recent version of this article appeared on March 25, 2003

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Submitted on September 3, 2002
Revised on November 27, 2002
Accepted on December 3, 2002

Small GTP-Binding Protein Rac Is an Essential Mediator of Vascular Endothelial Growth Factor-Induced Endothelial Fenestrations and Vascular Permeability

Anna Eriksson PhD, Renhai Cao PhD, Joy Roy MD, PhD, Katerina Tritsaris PhD, Claes Wahlestedt MD, PhD, Steen Dissing PhD, Johan Thyberg MD, PhD, and Yihai Cao MD, PhD^*

From the Microbiology and Tumor Biology Center (A.E., R.C., Y.C.), the Center for Genomics and Bioinformatics (C.W.), and the Department of Cell and Molecular Biology (J.T.), Karolinska Institute, Stockholm, Sweden; the Department of Surgical Sciences (J.R.), Karolinska Hospital, Stockholm, Sweden; and the Department of Medical Physiology (K.T., S.D.), The Panum Institute, University of Copenhagen, Copenhagen, Denmark.

^* To whom correspondence should be addressed. E-mail: yihai.cao{at}mtc.ki.se.

Background--Vascular endothelial growth factor/vascular permeability factor (VEGF/VPF) induces both angiogenesis and vascular permeability. Although its angiogenic activity has been well characterized, the signaling pathways of VEGF-induced permeability remain poorly understood.

Methods and Results--Using the mouse corneal micropocket assay, Miles assay, and a combination of cytochemical, electron microscopic, and biochemical assays, we demonstrate that VEGF-induced vascular leakage partly can be separated from its angiogenic activity. VEGF but not FGF-2 induced capillaries with a highly fenestrated endothelium, a feature linked with increased vascular permeability. A cell-permeable Rac antagonist (TAT-RacN17) converted VEGF-induced, leaky vascular plexuses into well-defined vascular networks. In addition, this Rac mutant blocked formation of VEGF-induced endothelial fenestrations and vascular permeability but only partially inhibited angiogenesis. Studies on endothelial cell cultures further revealed that VEGF stimulated phosphorylation of VEGF receptor-2 (VEGFR-2), leading to activation of Rac as well as increased phosphorylation of phospholipase C (PLC), protein kinase B (Akt), endothelial nitric oxide synthase (eNOS), and extracellular regulated kinase (Erk1/2). We further found that phosphatidylinositol-3-OH kinase (PI3K) acted upstream of Rac and Akt-eNOS in VEGF/VEGFR-2 signaling.

Conclusions--Our findings indicate that the small GTP-binding protein Rac is a key component in mediation of VEGF-induced vascular permeability but less so in neovascularization. This may have conceptual implications for applying Rac antagonists in treatment and prevention of VEGF-induced vascular leakage and edema in connection with ischemic disorders.

Key words: vasculature • receptors • endothelium • ischemia • proteins

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