Fig. 3

Effects of Notch1 activation in arterial specification of ESC-derived ECs. a. ICN1-induced expression of arterial EC markers. Protein levels of ICN1, NRP1 (arterial marker), NRP2 (vein marker), and GAPDH determined by western blotting analysis after Dox stimulation of iICN1-ECs for indicated time. b Effects of ICN1 induction on expression of EC surface markers. iICN1-ECs treated with or without Dox for 3 days, followed by FACS analysis after labeling with PE-conjugated anti-CXCR4, Alexa Fluor® 488-conjugated VE-cadherin, or Alexa Fluor® 647-conjugated CD31 antibodies. Percentages of the endothelial marker-positive populations indicated. c Effects of ICN1 induction on mRNA levels of Notch1 target genes in iICN1-ECs. mRNA levels of Hey1, Hey2, Hes1, Foxc2, Jag1, and Dll4 determined through qRT-PCR at day 1 after Dox treatment, and normalized GAPDH. Data represent mean ± SD (n = 3). *p < 0.05, **p < 0.01. Dox doxycycline, GAPDH glyceraldehyde 3-phosphate dehydrogenase, iICN1 inducible intracellular domain of Notch1, NRP neuropilin, PE phycoerythrin, VE-cad VE-cadherin