Role of the small GTPase Rap1 for integrin activity regulation in endothelial cells and angiogenesis

Blood ◽  
2009 ◽  
Vol 113 (2) ◽  
pp. 488-497 ◽  
Author(s):  
Guillaume Carmona ◽  
Stephan Göttig ◽  
Alessia Orlandi ◽  
Jürgen Scheele ◽  
Tobias Bäuerle ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Growth Factor ◽  
Umbilical Vein ◽  
Critical Role ◽  
Limb Ischemia ◽  
Small Gtpase ◽  
And Migration ◽  
Umbilical Vein Endothelial Cells ◽  
Endothelial Growth Factor

Abstract Ras-associated protein 1 (Rap1), a small GTPase, attracted attention because of its involvement in several aspects of cell adhesion, including integrin- and cadherin-mediated adhesion. Yet, the role of Rap1 genes and of Rap1 effectors for angiogenesis has not been investigated. Human umbilical vein endothelial cells (HUVECs) express Rap1a and Rap1b mRNA. To determine the contribution of Rap1 activity for angiogenesis, we overexpressed Rap1GAP1, a GTPase-activating protein that inhibits Rap1 activity. Overexpression of Rap1GAP1 significantly blocked angiogenic sprouting and tube-forming activity of HUVECs as well as migration and integrin-dependent adhesion. Silencing of Rap1a, Rap1b, or both significantly blocked HUVECs sprouting under basal and basic fibroblast growth factor-stimulated conditions and reduced HUVEC migration and integrin-dependent adhesion. We found that Rap1a and Rap1b are essential for the conformational activation of β1-integrins in endothelial cells. Furthermore, silencing of Rap1a and Rap1b prevented phosphorylation of tyrosine 397 in focal adhesion kinase (FAK) and vascular endothelial growth factor-induced Akt1-activation. Rap1a−/−-deficient and Rap1a+/− heterozygote mice displayed reduced neovascularization after hind limb ischemia compared with wild-type mice. Silencing of RAPL significantly blocked the Rap1-induced sprouting of HUVECs, suggesting that the angiogenic activity of Rap1 is partly mediated by RAPL. Our data demonstrate a critical role of Rap1 in the regulation of β1-integrin affinity, adhesion, and migration in endothelial cells and in postnatal neovascularization.

Platelet-Derived Exosomes Affect the Proliferation and Migration of Human Umbilical Vein Endothelial Cells Via miR-126

2019 ◽  
Vol 17 (4) ◽  
pp. 379-387 ◽  
Author(s):  
Yan Sun ◽  
Xiao-li Liu ◽  
Dai Zhang ◽  
Fang Liu ◽  
Yu-jing Cheng ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Growth Factor ◽  
Real Time ◽  
Umbilical Vein ◽  
Angiogenic Factors ◽  
Healthy Donors ◽  
And Migration ◽  
Umbilical Vein Endothelial Cells ◽  
Tgf Β1 ◽  
Proliferation And Migration

Background:Intraplaque angiogenesis, the process of generating new blood vessels mediated by endothelial cells, contributes to plaque growth, intraplaque hemorrhage, and thromboembolic events. Platelet-derived Exosomes (PLT-EXOs) affect angiogenesis in multiple ways. The ability of miR-126, one of the best-characterized miRNAs that regulates angiogenesis, carried by PLT-EXOs to influence angiogenesis via the regulation of the proliferation and migration of endothelial cells is unknown. In this study, we aimed to investigate the effects of PLT-EXOs on angiogenesis by Human Umbilical Vein Endothelial Cells (HUVECs).Methods:We evaluated the levels of miR-126 and angiogenic factors in PLT-EXOs from Acute Coronary Syndrome (ACS) patients and healthy donors by real-time Polymerase Chain Reaction (PCR) and western blotting. We incubated HUVECs with PLT-EXOs and measured cell proliferation and migration with the Cell Counting Kit-8 assay and scratch assay, respectively. We also investigated the expression of miR-126 and angiogenic factors in HUVECs after exposure to PLT-EXOs by western blotting and real-time PCR.Results:PLT-EXOs from ACS patients contained higher levels of miR-126 and angiogenic factors, including Vascular Endothelial Growth Factor (VEGF), basic Fibroblast Growth Factor (bFGF), and Transforming Growth Factor Beta 1 (TGF-β1), than those from healthy donors (p<0.05). Moreover, the levels of exosomal miR-126 and angiogenic factors were increased after stimulation with thrombin (p<0.01). HUVEC proliferation and migration were promoted by treatment with activated PLT-EXOs (p<0.01); they were accompanied by the over-expression of miR-126 and angiogenic factors, including VEGF, bFGF, and TGF-β1 (p<0.01).Conclusion:Activated PLT-EXOs promoted the proliferation and migration of HUVECs, and the overexpression of miR-126 and angiogenic factors, thereby elucidating potential new therapeutic targets for intraplaque angiogenesis.

MODULATION of PROSTACYCLIN PRODUCTION BY HUMAN UMBILICAL VEIN ENDOTHELIAL CELLS WITH ECGF/HEPARIN MEDIUM : ROLE OF CELLULAR DENSITY AT CONFLUENCE

1987 ◽  
Author(s):  
O BOUTHERIN-FALSON ◽  
N BLAES
Keyword(s):  
Endothelial Cells ◽  
Endothelial Cell ◽  
Growth Factor ◽  
Umbilical Vein ◽  
Calf Serum ◽  
Lower Amount ◽  
Endothelial Cell Proliferation ◽  
Human Umbilical Vein ◽  
Umbilical Vein Endothelial Cells

Prostacyclin (PGI2) is a major product of arachidonic acid metabolism in vascular endothelial cells. In addition to the role of exogenous agents, its production could be modulated by culture conditions : proliferative state, medium renewal, subcultivation... The use of endothelial cell growth factor (ECGF) associated with heparin has been shown to improve human endothelial cell proliferation. Here we report that human umbilical vein endothelial cells (HUVEC) grown in that medium produce less prostacyclin than without growth factor.HUVEC were cultured in RPMI-199 1:1 + 20% fetal calf serum, added or not with ECGF (Bovine hypothalamus extract BTI Cambridge, 24 ug/ml) and heparin (from porcine intestinal mucosa, Signa, 90 ug/ml). After 4 days in culture, medium was removed and replaced by Tyrode Hepes buffer and basal production was measured after 20 min. Cells were then submitted to 5 min thrombin to assess PGI2 production in stimulated conditions. PGI2 production was estimated by specific radioimmunoassay for 6 keto PGFjalpha. For each point, cell number in the culture was counted after Trypsin EDTA treatment. In the present study, cells grown in ECGF-heparin medium produce lower amount of PGI2, compared to heparin or control medium. This result was observed in both basal and stimulated conditions. For each medium (ECGF-heparin, heparin, control), correlations between PGI2 production per cell and log cell density were shown to be significantly negative.These observations suggest that ECGF effect on PGI2 production could be a consequence of its growth factor activity, notably by the fact that it leads to an endothelial monolayer made of more numerous cells. Since it is now suggested by a number of clinical observations that PGI2 is rather produced in pathological conditions, culture models showing a weak production of PGI2 appear in that connection doser to the physiological conditions.

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