Fibronectin promotes VEGF-induced CD34+ cell differentiation into endothelial cells

Background: Adult endothelial progenitor cells (EPC) may be a useful source for engineering the endothelialization of vascular grafts. However, the optimal factors that promote differentiation of EPCs into endothelium, remain to be elucidated. The goal of this current report was to determine which extracellular matrix (ECM) protein might modulate or enhance the effects of EPCs on differentiation into mature endothelium. Methods: Human EPCs (CD34(+) cells) were cultured in ECM-coated six-well plates in MCDB-131 medium containing vascular endothelial growth factor (VEGF), insulin-like growth factor-1, and basic fibroblast growth factor. After 21 days, differentiated endothelial colonies were confirmed by immunofluorescence for von Willebrand factor (vWF) and vascular-endothelial (VE)-cadherin and mRNA expression of the endothelial markers FIk-1, vWF, and VE-cadherin. Cell migration toward the VEGF-matrix protein combinations was also measured. Results: As judged by positive staining for endothelial markers vWF and VE-cadherin, the combination of VEGF with fibronectin (FN) produced significantly more endothelial colonies (P <.05) than did collagens I or IV or vitronectin. Defined fragments of IN did not enhance VEGF-mediated effects. Fibrinogen produced intermediate stimulation of differentiation. FN also enhanced VEGF-mediated CD34(+) cell migration. Blockade of alpha5beta1, but not alphavbeta3 or alphavbeta5, inhibited both VEGF-mediated CD34(+) cell differentiation and migration. Conclusions: VEGF and IN together significantly promote the migration and differentiation of CD34(+) cells. This synergism is specific to FN and the alpha5beta1 integrin. Combinations of VEGF and FN may be useful in promoting differentiation of circulating endothelial progenitors into endothelial cells for tissue engineering.