Fig. 4.

The activation of the Wnt/β-Catenin pathway successfully rescued the repression of mesendoderm differentiation induced by hypoxia. A Schematic diagram of AB2.2 mESC differentiation treated with or without CHIR under hypoxia. B–F Activating the Wnt/β-Catenin pathway with CHIR increased the total and nuclear β-Catenin levels in AB2.2 mESCs undergoing differentiation but had a minor effect on the cytoplasmic β-Catenin expression. G The expression of Wnt/β-Catenin downstream targets (Sp5 and Cdx1) was boosted by CHIR under hypoxia. H CHIR treatment restored T protein expression under hypoxia. T (red) was stained on differentiation day 4. Nuclei were stained with DPAI (blue). I CHIR treatment inhibited the expression of ectoderm markers (Pax6 and Nestin). J Schematic diagram of the parallel comparison of CHIR treatment effects on AB2.2 mESC differentiation under either normoxia or hypoxia. K CHIR treatment similarly improved the expression of mesendoderm markers (T, Eomes, Mesp1, and Gsc) in differentiating AB2.2 mESCs under both normoxia and hypoxia. L The ratio of T+ cells on differentiation day 4 in AB2.2 mESCs was detected by flow cytometry. M The ratio of Sox1+ cells on differentiation day 4 in AB2.2 mESCs was detected by flow cytometry. *, significant (P<0.05); ns, not significant