Involvement of RhoA/Rho Kinase Signaling in VEGF-Induced Endothelial Cell Migration and Angiogenesis In Vitro

Amerongen, Geerten P. van Nieuw; Koolwijk, Pieter; Versteilen, Amanda; Hinsbergh, Victor W.M. van

doi:10.1161/01.atv.0000054198.68894.88

Cited by 298 publications

(244 citation statements)

References 59 publications

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“…We found that RhoA transiently becomes activated 5-15 min after VEGF treatment. This data confirmed work by other groups [18,19] that demonstrated a rapid and transient increase of RhoA activity in the first few minutes after VEGF treatment. Cdc42 was also activated 5-15 min after stimulation with VEGF; this activity declined and elevated again around 60 min.…”

Section: Vegf Activates Rac1 Through Vegfr-2 In Endothelial Cellssupporting

confidence: 92%

VEGF-induced Rac1 activation in endothelial cells is regulated by the guanine nucleotide exchange factor Vav2

Garrett

Buul

Burridge

2007

Experimental Cell Research

152

132

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Vascular endothelial growth factor (VEGF) signaling is critical for both normal and diseaseassociated vascular development. Dysregulated VEGF signaling has been implicated in ischemic stroke, tumor angiogenesis, and many other vascular diseases. VEGF signals through several effectors, including the Rho family of small GTPases. As a member of this family, Rac1 promotes VEGF-induced endothelial cell migration by stimulating the formation of lamellipodia and membrane ruffles. To form these membrane protrusions, Rac1 is activated by guanine nucleotide exchange factors (GEFs) that catalyze the exchange of GDP for GTP. The goal of this study was to identify the GEF responsible for activating Rac1 in response to VEGF stimulation. We have found that VEGF stimulates biphasic activation of Rac1 and for these studies we focused on the peak of activation that occurs at 30 min. Inhibition of VEGFR-2 signaling blocks VEGF-induced Rac1 activation. Using a Rac1 nucleotide-free mutant (G15ARac1), which has a high affinity for binding activated GEFs, we show that the Rac GEF Vav2 associates with G15ARac1 after VEGF stimulation. Additionally, we show that depleting endothelial cells of endogenous Vav2 with siRNA prevents VEGF-induced Rac1 activation. Moreover, Vav2 is tyrosine phosphorylated upon VEGF treatment, which temporally correlates with Rac1 activation and requires VEGFR-2 signaling and Src kinase activity. Finally, we show that depressing Vav2 expression by siRNA impairs VEGF-induced endothelial cell migration. Taken together, our results provide evidence that Vav2 acts downstream of VEGF to activate Rac1.

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Section: Vegf Activates Rac1 Through Vegfr-2 In Endothelial Cellssupporting

confidence: 92%

VEGF-induced Rac1 activation in endothelial cells is regulated by the guanine nucleotide exchange factor Vav2

Garrett

Buul

Burridge

2007

Experimental Cell Research

152

132

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“…In addition, as a major angiogenesis factor, in VEGF-stimulated cells, RhoA signaling is necessary for cell migration (25). Previous experiments indicated that RhoA critically regulates angiogenesis driven by VEGF and constitutively active RhoA can complement VEGF to increase angiogenesis by >100%, whereas dominant negative RhoA markedly inhibits VEGF-driven neovascularization (26). However, in this study, we demonstrate that the overexpression of RhoA is sufficient to induce migration and increase motility of human umbilical vein endothelial cells, in the absence of any exogenous stimulation, such as growth factors or cytokines.…”

Section: Discussionmentioning

confidence: 99%

The effect of RhoA on human umbilical vein endothelial cell migration and angiogenesis in vitro

Zhao

Xu²,

Zhou³

et al. 2006

Oncol Rep

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Abstract. The mechanisms that control the morphologic organization of endothelial cells (ECs) into new blood vessels are not well understood. Recent studies revealed that the small G proteins of the Rho family are key regulators of cell migration, involving reorganization of the actin cytoskeleton, cell migration and the regulation of gene transcription. We hypothesized that RhoA GTPase, a member of the Rho family, may play an important role in EC organization during angiogenesis, the process of new vessel formation in preexisting tissues. To test this hypothesis, we investigated the effects of RhoA on human umbilical vein endothelial (HUVE) cell migration and angiogenesis in vitro, by stably transfecting HUVE cells with sense RhoA expression plasmid through the Lipofect-2000 system. Wound assay in vitro and 3-dimensional cell culture were used to detect the migration and angiogenesis capacity of HUVE cells. The morphological changes of transfected cells were revealed under confocal and phase contrast microscopy. Our results demonstrated that the increased expression of RhoA in HUVE cells significantly enhanced the morphogenetic changes and cytoskeletal reorganization of the transfected cells, and also enhanced cell migration and angiogenic capacity in vitro, suggesting that RhoA plays an important role in the process of HUVE cell migration and angiogenesis in vitro.

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“…For instance, it has been demonstrated that VEGF increases RhoA activity along with its recruitment to the cell membrane; at the same time, RhoA signaling has been implicated in VEGF-mediated angiogenesis. 8 In addition, RhoA activation increases tyrosine phosphorylation of VEGF receptor 2 (VEGFR-2). 9 Although RhoA signaling has been implicated in the regulation of angiogenesis, a comprehensive analysis of the specific mechanisms by which it mediates VEGF-induced angiogenesis still needs to be done.…”

Section: Introductionmentioning

confidence: 99%

Ras homolog gene family, member A promotes p53 degradation and vascular endothelial growth factor‐dependent angiogenesis through an interaction with murine double minute 2 under hypoxic conditions

Xue

Cui

et al. 2012

Cancer

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BACKGROUND: Tumor neovascularization (TNV) is a common pathologic basis for malignant growth and metastasis. However, the mechanism of TNV pathogenesis is not fully understood. Ras homolog gene family, member A (RhoA), a Rho guanosine triphosphatase (GTPase) family member, may be involved in a hypoxia-induced vascular endothelial growth factor (VEGF) pathway that regulates TNV angiogenesis through an unclear mechanism. METHODS: The regulation of RhoA on p53, the p53 binding protein homolog murine double minute 2 (MDM2), and VEGF was analyzed in hypoxic MCF-7 cells using Western blot analysis, real-time polymerase chain reaction (PCR) analysis, coimmunoprecipitation, and immunofluorescence staining assays. Changes in proliferation, invasion, migration, stress fiber formation, and tube formation were detected in an MCF-7 human umbilical vein endothelial cell (HUVEC) coculture system. Correlations of RhoA expression with MDM2, wild-type p53 (wt-p53), and VEGF expression in breast cancer tissues and relations between RhoA and breast cancer clinical features were analyzed by immunohistochemistry. RESULTS: Activated RhoA down-regulated p53 protein, which increased VEGF expression in hypoxic MCF-7 cells; whereas p53 messenger RNA levels were not altered. In addition, the ubiquitin-mediated degradation of p53 was enhanced by active RhoA. RhoA and MDM2 colocalized in the cytoplasm of hypoxic MCF-7 cells and interacted with each other physically. Furthermore, nutlin-3, a specific MDM2 inhibitor, was capable of reducing activated RhoA-induced p53 protein stability and attenuating VEGF accumulation. In an MCF-7-HUVEC coculture system, nutlin-3 effectively inhibited HUVEC proliferation, invasion, migration, stress fiber formation, and tube formation mediated by activated RhoA under hypoxic conditions. Data from 129 clinical breast cancer specimens with wt-p53 revealed that high RhoA expression was correlated with high MDM2 expression, low wt-p53 expression, and high VEGF expression. CONCLUSIONS: The current data suggested that activated RhoA promotes VEGF expression and hypoxia-induced angiogenesis through the up-regulation of MDM2 to decrease p53 stability. Cancer 2012;118:4105-16.

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Involvement of RhoA/Rho Kinase Signaling in VEGF-Induced Endothelial Cell Migration and Angiogenesis In Vitro

Cited by 298 publications

References 59 publications

VEGF-induced Rac1 activation in endothelial cells is regulated by the guanine nucleotide exchange factor Vav2

VEGF-induced Rac1 activation in endothelial cells is regulated by the guanine nucleotide exchange factor Vav2

The effect of RhoA on human umbilical vein endothelial cell migration and angiogenesis in vitro

Ras homolog gene family, member A promotes p53 degradation and vascular endothelial growth factor‐dependent angiogenesis through an interaction with murine double minute 2 under hypoxic conditions

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