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. 2012 Dec 5;304(4):C289–C298. doi: 10.1152/ajpcell.00298.2012

Fig. 7.

Fig. 7.

Myocardin expression in SMC differentiation of hES-MCs. A: TGF-β-induced myocardin (Myocd) mRNA expression as detected by RT-PCR. B: TGF-β-induced Myocd expression as detected by qPCR. *P < 0.05 compared with vehicle-treated cells (0 h). C: TGF-β-induced Myocd expression via p38 MAPK and PI3K signaling pathways. hES-MCs were treated with pathway-specific inhibitors for p38 (SB203580), JNK (SP600125), RhoA (Y27632), PI3K (LY294002), or ERK1/2 (U0126) for 30 min prior to vehicle (Ctrl-β) or TGF-β (1 ng/ml) treatment for 8 h. Myocd expression was detected by qPCR. *P < 0.05 compared with vehicle-treated group; #P < 0.05 compared with TGF-β-treated group without inhibitor (Ctrl+β). D: TGF-β-induced Cnn1 expression was blocked by p38 MAPK and PI3K inhibitors. *P < 0.01 compared with vehicle-treated group (Ctrl-β); #P < 0.01 compared with TGF-β-treated group without inhibitor (Ctrl+β). E and F: both Smad2 and Smad3 were required for TGF-β-induced Myocd expression. Knockdown of Smad2 expression by shRNA (shS2; E) or blockade of Smad3 activity by Smad3 inhibitor (SIS3, Smad3 inhibitor; F) inhibited Myocd expression. *P < 0.01 compared with vehicle-treated group; #P < 0.05 compared with TGF-β-treated group with control shRNA (shCtrl; E) or SIS3 vehicle (Ctrl; F). G: Myocd expression in different developmental stages. Total RNA was extracted from hES cells (H9), mesoderm (J38), hES-MCs, and TGF-β-treated hES-MCs. Myocd expression was detected by RT-PCR. H: quantitative analysis of Myocd expression shown in G by normalization to cyclophilin. *P < 0.05 compared with H9 cells; #P < 0.05 compared with J38 mesoderm; $P < 0.001 compared with all other cells.