Negative Regulation of Vascular Endothelial Growth Factor Receptor-2-Mediated Angiogenesis by Phosphatidylinositol 3-Kinase

Objective: To test the hypothesis that vascular endothelial growth factor receptor-2 (VEGFR-2)-induced angiogenesis via phospholipase Cy (PLC ψ ) may be negatively regulated by phosphatidylinositol 3-kinase (PI3K). Design: Experimental cell lines study. Setting: School of Experimental Medicine, College of Medicine and Dentistry, University of Birmingham, Birmingham, UK. Method: In vitro angiogenesis was assessed in porcine aortic endothelial cells stably expressing human VEGFR-2 (PAEC-hVEGFR-2) using pharmacological and molecular biology tools. In addition, endothelial cell lysates were im-munoblotted to assess the levels of active PLCy. Result: Incubation of PAEC-hVEGFR-2 cells with the growth factor VEGF caused a partial capillary- like tube network formation in endothelial cells plated on growth-factor reduced MType equation here.atrigel that was abrogated with U73122, a PLC ψ inhibitor. Preincubation of these endothelial cells with the PI3K inhibitor, LY294002 augmented VEGF-induced PLC ψ phosphorylation and capillary network. In addition, adenoviral gene transfer of the endogenous inhibitor of PI3K cell signalling, phosphatase and tensin homolog (PTEN) into PAEC-hVEGFR-2 caused a profound increase in VEGF-induced angiogenesis similar to the effect of the pharmacological inhibitor LY294002 that was not observed in the control group. Conclusion: We conclude that inhibition of PI3K significantly augmented VEGFR-2 induced angiogenesis, possibly due to a competition between PI3K and PLC ψ enzymes on the same shared substrate.