Fig. 7

Hypoxia enhanced the hematopoietic differentiation by promoting the EHT progression. A Flow cytometry analysis for HEPs (hESC-derived CD144⁺CD34⁺ cells) at day 11 after the hematopoietic differentiation under the hypoxia or normoxia conditions. B CLSM images of the immunostaining for VE-Cadherin and RUNX1 at day 15 cells during the hematopoietic specification under the normoxia or hypoxia conditions. The colors of green, red and blue represent the staining for RUNX1, VE-Cadherin and nuclei. (Original magnification, 40×, scale bar 50 μm). C Relative endothelial gene expression of differentiated cells at different time points under the normoxia or hypoxia conditions. D Flow cytometry analysis for HPCs (hESC-derived CD34⁺CD45⁺ cells) during the hematopoiesis under the hypoxia or normoxia conditions. E T lineage potential of HPCs at day 15 after the hematopoietic differentiation following 4 weeks of culture for induction of T cells. F and G Cell cycle distribution was determined by flow cytometry after the hematopoietic differentiation under the normoxia or hypoxia conditions. The percentage of S phase was significantly increased in hESC-derived CD34⁺CD45⁺ cells under the normoxia conditions. H CLSM images of the immunostaining for ROS1 in hESC-derived CD34⁺CD45⁺ cells under the normoxia or hypoxia conditions. The colors of red and blue represent the staining for ROS1 and nuclei. (Original magnification, 40×, scale bar 50 μm). I Gene expression analysis of cell cycle gene (P27) and apoptosis gene (BAX) by RT-qPCR at different time points during the hematopoietic differentiation under the normoxia or hypoxia conditions (n = 3, mean ± SEM). J Gene set enrichment analysis for the identified genes involved in HIF-1a induced hypoxia during the hematopoietic specification. K RT-qPCR analysis for the expressions of HIF-1α, VEGFA, NOTCH1 and RUNX1. Data represent the mean ± SEM. *p < 0.05, **p < 0.01, and ns (no significance)