Dynamics of Vascular Endothelial-Cadherin and β-Catenin Localization by Vascular Endothelial Growth Factor-Induced Angiogenesis in Human Umbilical Vein Cells

The adherens junctional molecule, vascular endothelial cadherin (VE-cadherin), functions to maintain adherens junction stability and to suppress apoptosis of endothelial cells by forming a complex with vascular endothelial growth factor (VEGF) receptor 2 and members of the armadillo family of cytoplasmic proteins. In order to investigate the dynamics of the association of VE-cadherin with adherens junctions during the initial stages of angiogenesis, human umbilical cord endothelial cells (HUVECs) were stimulated with VEGF to undergo angiogenesis in type-I collagen three-dimensional culture. In confluent monolayers of HUVECs, VE-cadherin and its signaling partner, β-catenin, as well as the paracellular transmembrane adhesion molecule platelet–endothelial cell adhesion molecule (PECAM-1), were all present in regions of cell–cell contact. Within 3 h of stimulation of angiogenesis, VE-cadherin and β-catenin were lost from these regions. In contrast, the distribution pattern of PECAM-1 did not alter. After 6 h the majority of endothelial cells had migrated to form a network of capillary cords with cell–cell contacts that contained all three molecules. By metabolic labeling of HUVECs it was found that de novo synthesis of VE-cadherin was not essential for the formation of new adherens junctions. Coimmunoprecipitation and immunoblotting experiments showed that the VE-cadherin and β-catenin remained associated after they were lost from adherens junctions. Detergent extraction of cells with Triton X-100 indicted that the majority of VE-cadherin and β-catenin was Triton soluble, indicating that they are only weakly associated with the actin-based cytoskeleton.