Research Article

Fine Tuning of Hepatocyte Differentiation from Human Embryonic Stem Cells: Growth Factor vs. Small Molecule-Based Approaches

Figure 3

Small molecule, CHIR-mediated differentiation of hESCs into definitive endoderm (DE). (a) H9 cells were treated with 3 μM or 4 μM CHIR for in DE media followed by 1 day in CHIR-withdrawn DE media (3 μM CHIR (2 d) and 4 μM CHIR (2 d), respectively) and 3 μM or 4 μM CHIR for in DE media followed by 4 days in CHIR-withdrawn DE media (3 μM CHIR (6 d) and 4 μM CHIR (6 d), respectively). The cells were fixed and photographed for phase images. The cells were then stained and imaged by a fluorescence microscope using antibodies against FOXA2 and SOX17 as DE markers. DAPI represents nuclear staining. . (b) The cells as explained above were collected on D0 (undifferentiated H9 cells), D2 (3 μM CHIR (2 d) and 4 μM CHIR (2 d)), or D6 (3 μM CHIR (6 d) and 4 μM CHIR (6 d)) of DE differentiation and analyzed for mRNA expression by RT-qPCR for the indicated genes using gene-specific primers. The bars represent normalized (18S rRNA) fold mRNA expression with values of undifferentiated cells (D0) set as 1. The data are represented as .
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