Fig. 2. Characterization of β-catenin-activated hES cells. (A) The Nβ-cateninER cells cultured with or without 4OHT for the indicated time periods were subjected to immunostaining with anti-KDR and anti-CXCR4 antibodies, and analyzed by FACS. The gated region denoted the cells scored as antigen-positive (the percent cells scored positive is shown in each panel). ES, unstimulated Nβ-cateninER cells. (B) Developmental potentials of β-catenin-activated cells towards mesoderm lineage. RT-PCR analysis of endothelial cell-specific markers at different time points: the Nβ-cateninER cells were cultured with 4OHT for 3 days (day 3), and further cultured with vascular endothelial growth factor (VEGF) for additional 3 days (day 6). The transgenic cell lines derived from different ES cell lines are shown (K1βcatER derived from KhES-1 cells, K3βcatER derived from KhES-3 cells). (C) Morphology of endothelial-like cells derived from β-catenin-activated cells (day 6). (D) Capillary tube-like networks formed on matrigel for 24 hours after culturing endothelial-like cells derived from β-catenin-activated cells. Scale bar: 100 µm.