KDR activation is crucial for VEGF165-mediated Ca21 mobilization in human umbilical vein endothelial cells

Cunningham, Sonia A., Tuan M. Tran, M. Pia Arrate, Robert Bjercke, and Tommy A. Brock. KDR activation is crucial for VEGF165-mediated Ca21 mobilization in human umbilical vein endothelial cells. Am. J. Physiol. 276 (Cell Physiol. 45): C176–C181, 1999.—We have prepared a polyclonal mouse antibody directed against the first three immunoglobulin-like domains of the kinase insert domain-containing receptor (KDR) tyrosine kinase. It possesses the ability to inhibit binding of the 165-amino acid splice variant of vascular endothelial cell growth factor (VEGF165) to recombinant KDR in vitro as well as to reduce VEGF165 binding to human umbilical vein endothelial cells (HUVEC). These results confirm that the first three immunoglobulin-like domains of KDR are involved in VEGF165 interactions. The anti-KDR antibody is able to completely block VEGF165-mediated intracellular Ca21 mobilization in HUVEC. Therefore, it appears that binding of VEGF165 to the fms-like tyrosine kinase (Flt-1) in these cells does not translate into a Ca21 response. This is further exemplified by the lack of response to placental growth factor (PlGF), an Flt-1-specific ligand. Additionally, PlGF is unable to potentiate the effects of submaximal concentrations of VEGF165. Surprisingly, the VEGF-PlGF heterodimer was also very inefficient at eliciting a Ca21 signaling event in HUVEC. We conclude that KDR activation is crucial for mobilization of intracellular Ca21 in HUVEC in response to VEGF165.