Abstract 3938: Basement Membrane and VEGF Synergistically Activate Cdc42 and Rac1 to Promote Endothelial Cell Migration and Survival during Vasculogenesis from Embryonic Stem Cells
Vasculogenesis depicts the formation of nascent blood vessels from progenitors both during embryonic development and in response to ischemic insults in adult. The role of vascular endothelial growth factor (VEGF) and its receptors has been well established in endothelial cell proliferation, migration and survival. However, relatively little is known about the extracellular matrix (ECM)-induced signaling pathways that coordinate with VEGF-stimulated responses to regulate the vasculogenetic process. In this study we show that cohesive cell aggregates that express endothelial lineage markers Flk-1, CD34 and PECAM-1 but not VE-cadherin, can arise in differentiated embryoid bodies derived from mouse embryonic stem cells. These endothelial precursors subsequently differentiate into VE-cadherin-positive endothelial cells that migrate to form interconnecting vascular channels. We further show for the first time that basement membrane (BM), a specialized ECM that underlies the endothelium of mature vessels, forms on the surface of the aggregated endothelial precursors. Treatment of isolated endothelial precursors with the BM protein laminin-111 induces the formation of cohesive cell spherules and the BM assembly on their surfaces; the latter markedly enhances VEGF-induced Cdc42 and Rac1 activation. This synergistic effect of BM and VEGF is likely mediated by β1 integrin and integrin-linked kinase because their knockout inhibits Cdc42 and Rac1 activation. Moreover, ablation of Cdc42 in embryoid bodies completely blocks endothelial cell migration and the assembly of vascular networks without affecting endothelial lineage differentiation. Lastly, targeted deletion of the Rac1 gene causes massive apoptosis of the endothelial precursors. These results suggest that BM formation and VEGF signaling synergistically activate the small GTPase Cdc42 and Rac1, thereby promoting endothelial cell migration and survival during vasculogenesis.