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. 2020 Feb 21;16(8):1450–1462. doi: 10.7150/ijbs.44414

Figure 1.

Figure 1

Early epidermal cell fate determination under the influence of multiple signaling pathways in hESCs. See also Figure S1. A. Stagewise method for keratinocyte differentiation. B. The time course of epidermal differentiation under TGFβ inhibition. hESCs were treated with SB431542 (SB) in differentiation medium (See Materials and Methods), and the cells were harvested at specific time points for analysis of gene expression by RT-qPCR. The results were normalized to GAPDH. C. The emergence of TP63 under pulse treatment of TGFβ inhibitor. hESCs were treated with SB431542 (SB) for specified periods in differentiation medium followed by differentiation medium only until day 6. Cells were collected on day 6 for analysis of gene expression by RT-qPCR. The results were normalized to GAPDH. D. BMP4 promotes TP63 expression under TGF-β inhibition. hESCs were treated with SB431542 (SB) in differentiation medium, and BMP4 was added from day 1 till day 6 when cells were harvested for gene expression analysis by RT-qPCR. The results were normalized to hESC; Data are presented as the mean±SD of three independent experiments. *, p < 0.05. E. The impact of WNT pathway modulation on epidermal cell fate determination. hESCs were treated with SB431542 (D0-6) and BMP4 (D1-6) in differentiation medium, and CHIR99021(CHIR, 5 µM) or IWR-1 (1 μM) was added from day 1 to day 6. Cells were then harvested for gene expression analysis by RT-qPCR. The results were normalized to hESC; Data are presented as the mean±SD of three independent experiments. *, p < 0.05. ns, not significant.