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Rho and ROCK Signaling in VEGF‐Induced Microvascular Endothelial Hyperpermeability
Author(s) -
SUN HENGRUI,
BRESLIN JEROME W.,
ZHU JUN,
YUAN SARAH Y.,
WU MACK H.
Publication year - 2006
Publication title -
microcirculation
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.793
H-Index - 83
eISSN - 1549-8719
pISSN - 1073-9688
DOI - 10.1080/10739680600556944
Subject(s) - rhoa , stress fiber , myosin light chain kinase , phosphorylation , microbiology and biotechnology , rock1 , rho associated protein kinase , vascular permeability , chemistry , vascular endothelial growth factor , endothelial stem cell , actin , actin cytoskeleton , biophysics , cytoskeleton , signal transduction , biology , focal adhesion , biochemistry , cell , cancer research , endocrinology , in vitro , vegf receptors
ABSTRACT Objectives: Vascular endothelial growth factor (VEGF) plays an important role in the regulation of microvascular permeability under various physiological and pathological conditions. The authors tested the hypothesis that the small GTPase Rho and its downstream effector ROCK (Rho‐associated coiled‐coil‐containing protein kinase) mediate VEGF‐induced increases in venular permeability. They also investigated myosin light chain (MLC) phosphorylation and actin polymerization, two well‐characterized targets of the Rho‐ROCK pathway that are implicated in the regulation of endothelial barrier function. Methods: The apparent permeability coefficient of albumin ( P a ) was measured in intact isolated porcine coronary venules and in cultured coronary venular endothelial cell (CVEC) monolayers. RhoA activation was determined using a Rhotekin‐agarose pull down assay. MLC phosphorylation was evaluated by immunoblotting with phospho‐specific antibodies, and endothelial cellular F‐actin was viewed using fluorescence microscopy. Results: VEGF increased P a in both isolated coronary venules and CVEC monolayers. The hyperpermeability response occurred in a similar time course to that of Rho activation, MLC phosphorylation, and actin stress fiber formation. Selective blockage of ROCK with Y27632 dose‐dependently inhibited VEGF‐induced venular hyperpermeability. Moreover, inhibition of either Rho with exoenzyme C3 or ROCK with Y‐27632 attenuated VEGF‐induced increases in permeability, MLC phosphorylation, and actin‐stress fiber formation in CVEC monolayers. Conclusions: Collectively, these findings suggest that the Rho–ROCK signal pathway contributes to VEGF‐induced hyperpermeability. Myosin light‐chain phosphorylation and actin stress fiber formation occur concomitantly with the increase in permeability upon VEGF stimulation.