Fig. 8. Effect of Smad4 knockdown on TGFβ-induced EndMT of MESECs. MESECs were sorted from the vascular cells derived from ESCs, transfected with Smad4 siRNA or with scrambled sequence as a negative control (NTC), and cultured in the absence (–) or presence of TGFβ2 (Tβ). (A) Levels of endogenous expression of Smad4 in the MESECs were analyzed by immunoblotting. (B,C,E,F) Levels of expression of PAI1 (B), Snail (C), claudin 5 (E) and SMA (F) in the MESECs were analyzed by quantitative real-time RT-PCR. Error bars indicate s.d. (D) The MESECs were subject to immunofluorescence staining for PECAM1 (red) and SMA (green).