Migration and differentiation of CEPs.

(A) Freshly isolated CD34⁺ cells at 105 per well containing 2 × 103CD34⁺VEGFR-2⁺ cells were placed in the upper chamber of 5-μ Costar transwell plates. Immediately, SDF-1 (200 ng/mL) or VEGF (100 ng/mL) was added in the lower chamber and, after 3 hours, migrated cells were quantified for the presence of CD34⁺VEGFR-2⁺ endothelial colonies. As a control, serum and cytokine-free media were used in the lower chamber in a separate experiment. (B) Incubation of migrated CD34⁺ cells with VEGF and FGF-2 on collagen-coated plastic dishes for 2 weeks resulted in differentiation of CD34⁺VEGFR-2⁺ cells into adherent endothelial cell monolayers. Adherent endothelial colonies expressed endothelial-specific markers E-selectin, VE-cadherin, and vWF but did not express AC133 (magnification 200x). E-selectin expression was induced by IL-1β stimulation.