Erk1/2 signaling pathway involved in regulating tissue factor expression in trophoblastic and hematopoietic differentiation of human embryonic stem cells. (A) Western blot analysis of phosphorylated Erk1/2 and Akt in various types of cells showing that Erk1/2 signaling pathway is active in granulocyte–macrophage (G-M) cells and trophoblasts. p-Erk1/2, phosphorylated Erk1/2; t-Erk1/2, total Erk1/2; pAkt, phosphorylated Akt; t-Akt, total Akt. (B),(C) Decreased (B) mRNA and (C) protein levels of tissue factor (TF) in G-M cells or trophoblasts treated with Erk1/2-specific inhibitor, U0126. Cells were treated with 10 μM U0126 or dimethylsulfoxide (control) for 4-6 or 7-9 days before harvest for quantitative real-time polymerase chain reaction for CDX2, PU.1, and TF mRNA levels. Data were reported as the mean ± standard error of the percentage of the mRNA levels of CDX2, PU.1, and TF in cells from the control group. *P <0.05. Western blotting analysis was carried out on the days designated in (C). hESC, human embryonic stem cell; HSPC, hematopoietic stem/progenitor cell.