VEGF₁₆₅ induces arterial specification of hMAPCs but not hAC133⁺ cells. (A) Q-RT-PCR for arterial (EphrinB1, Dll-4, Hey-2, EphrinB2) and venous markers (EphB4) on hAC133⁺ cell–derived ECs (▪) or hMAPC-derived ECs (⊡) at different time points (0, 7, 14, and 21 days) after the start of the differentiation process. While hMAPCs up-regulated arterial and venous markers during the differentiation process, hAC133⁺ cell–derived ECs showed reduced arterial marker expression. Expression levels are presented as fold increase (in logarithmic scale) in comparison with baseline levels and were normalized by using GAPDH as housekeeping gene. The mRNA levels in undifferentiated hMAPCs were considered as 1. Expression between baseline levels (day 0) and day 7, 14, and 21 for each cell population was compared (*P < .05; **P < .01). (B-D) Immunofluorescent staining of hMAPC-derived ECs. After 14 days, hMAPCs were positive for arterial markers EphrinB1 (B), Hey-2 (C), and venous marker EphB4 (D) (see text for percentage of positive cells). A representative example from 3 different clones is shown. (E) Comparative expression, plotted as percentage of total number of cells, based on FACS analysis, of the microvascular-specific marker CD36 in hMAPC (⊡) and hAC133⁺ cell–derived ECs (▪) (**P < .01 versus hMAPC-ECs). The mean (± SEM) of 3 (A) or 5 (E) different experiments in triplicate is shown. Magnification ×40.