Fig. 6. Effect of Snail knockdown on MESECs. MESECs were sorted from the vascular cells derived from ESCs, transfected with Snail siRNA or with scrambled sequence as a negative control (NTC), and cultured in the absence (–) or presence of TGFβ2 (Tβ). (A) The levels of endogenous expression of Snail in the MESECs were analyzed by quantitative real-time RT-PCR. Error bars indicate s.d. Black and gray bars, represent +TGFβ2 and –TGFβ2, respectively. (B) The MESECs were subjected to immunofluorescence staining for PECAM1 (red) and SMA (green). (C) Quantitative analysis of the effects of Snail on colony formation from single MESECs, performed as described in Fig. 1F. Briefly, MESECs transfected with Snail siRNA or scrambled sequence were cultured at low density with 10% FCS in the absence (–) or presence of TGFβ2 (Tβ) for 4 days, followed by staining of colonies for PECAM1 and SMA. E, pure endothelial colony; M, mural-containing colony. Scale bars: 100 µm.