Hh and VEGF are required for arterial identity in mice. (A) Whole-mount in situ hybridization for Dll4 mRNA in E8.5 (approximately 8-somite stage) Smo^+/− and Smo^−/− embryos. Brackets indicate the occipital somite (OS) and presomitic/primitive streak (PS) regions of the embryo microdissected for Q-RT-PCR in panel B. Scale bar equals 500 μm. (B) Q-RT-PCR for Dll4 mRNA in Smo control (Smo^+/+, n = 2 and Smo^+/−, n = 2) and Smo^−/− (n = 4) littermates. Q-RT-PCR was performed on distinct regions of 8-somite stage embryos corresponding to the occipital somites region (labeled OS in panel A) and presomitic region (labeled PS in panel A) in control or mutant embryos. Data are presented as Dll4 expression relative to the loading control, Polr2a. Asterisk indicates statistical significance between control and mutant sample groups, P < .05. (C) Whole-mount in situ hybridization for Dll4 mRNA in E8.5 (7- to 8-somite stage) VEGF hypomorphic embryos. Representative embryos from VEGF^+/+ (n = 4), VEGF^hypo/+ (n = 3), and VEGF^hypo/hypo (n = 2) littermates are shown. EC indicates endocardium; SV, sinus venosus; DA, dorsal aorta; YS, yolk sac plexus; and AL, allantoic artery.