Figure 5 | Scientific Reports

Figure 5

From: Spatiotemporal patterning of EpCAM is important for murine embryonic endo- and mesodermal differentiation

Figure 5

EpCAM impact on endodermal differentiation. (a) E14TG2α ESC were subjected to endodermal differentiation upon treatment with retinoic acid (RA) and basic fibroblast growth factor (bFGF) as described in the Methods section. Oct3/4, EpCAM, Foxa2, Gata4, Eomes, and Afp mRNA levels were assessed by quantitative PCR in pluripotent (WT) and differentiated cells. (n = 2 independent experiments with duplicates). (b) EpCAM, Foxa2 and Gata4 expression in EB (d5). Shown are representative immunohistochemistry staining of EB from two independent experiments with multiple EB each. (c) Representative diagram of cell surface expression of EpCAM in pCAG and EpCAM transfectants measured by FACS (n = 3 independent experiments). (d) Shown are mean cell surface expression of EpCAM and SSEA-1 in EpCAM transfectants of pluripotent ESC and differentiated EB at day 21 (n = 3 independent experiments). (e) Oct3/4, SOX2 and NANOG mRNA expression was measured by quantitative PCR in pluripotent (ES cells) and differentiated transfectants at the indicated time points (n = 3 independent experiments). (f) Afp and Fn1 mRNA expression was measured by quantitative PCR in pluripotent and differentiated pCAG and EpCAM-expressing E14TG2α ESC at the indicated time points (n = 3 independent experiments). (g) Cell surface EpCAM expression was measured by FACS and in cell lysates of wild-type and EPCAM knockout clones under pluripotency conditions (n = 3 independent experiments). (h) Oct3/4, Sox2 and Nanog mRNA expression was measured by quantitative PCR in wild-type and EpCAM knockout clones under pluripotency conditions (n = 3 independent experiments). (i) Foxa2, Afp and Eomes mRNA expression was measured by quantitative PCR in wild-type and EpCAM knockout clones after endodermal differentiation upon treatment with RA and bFGF at day 5 as described in Methods section (n = 3 independent experiments). Mean ± SEM; Student’s T-test (n = 2 groups) or One-Way ANOVA (n ≥ 3 groups); p < 0.05, **p < 0.01, ***p < 0.001.

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