Fig. 2

Analysis and modulation of the A_2bR pathway, which is dysregulated in the endothelium of cecr1b-LoF embryos.

a Schematic representation of the A_2bR-dependent eAdo signaling pathway. A_2bR activation in the endothelium leads to the expression of HSPC-specific genes (runx1 and cmyb) through activation of the cAMP/Pka. Expression levels of (b) cxcl8 and (c) cmyb measured by RT-qPCR in endothelial cells sorted (ECs) from 30–32 hpf Tg(fli1a:GFP)^y1. Data are presented as mean ± SEM; n = 3 biologically independent experiments. Statistical significance was assessed by Ordinary one-way ANOVA test with Tukey’s correction; ***p < 0.001; **p < 0.002; ns, not significant. d Representative images of WISH staining for the HE markers runx1 and scl in the trunk region of 24–26 hpf embryos and for the specifying HSPCs marker cmyb in 30-32 hpf embryos. DA dorsal aorta, PCV posterior cardinal vein, s-HSPCs specifying HSPCs, HE hemogenic endothelium. Numbers indicate the embryos belonging to the representative phenotype of each category, shown in the image. Scale bar = 100 microns. e WISH staining of the arterial (efnb2) and venous (efnb4) markers in the trunk region of 28–30 hpf embryos. DA dorsal aorta, PCV posterior cardinal vein. Red arrowheads indicate the region of the DA, blue arrowheads indicate the region of the PCV. Scale bar = 100 microns.