Fig. 7. Synergistic interaction of Activin and canonical Wnt/β-catenin signaling pathways in the anterior PS/endoderm specification during hES cell differentiation. (A) qPCR analysis on RNA isolated from HES-3 cells, with or without Activin A (100 ng/ml) or DKK1 (100 ng/ml) for 3 days, was performed (v, vehicle control; A, Activin A treated). (B,C) The Nβ-cateninER cells were cultured with Activin A (100 ng/ml) in the presence or absence of 4OHT for 3 days, and then analyzed by qPCR (B), or cell lysates were subjected to SDS-PAGE and probed with specific antibodies as indicated (C) (OHT, 4OHT treated). (D) Proposed model for early lineage determination of hES cell differentiation by the canonical Wnt/β-catenin, Activin and BMP signaling pathways (see Discussion).