The role of VEGFR3 in the mouse aorta. (A) VEGFR3 expression in arterial endothelium. Total RNA isolated from the endothelial layer was analyzed by qPCR for the indicated genes. VEcad and VEGFR3 expression are represented as mean fold enrichment of the endothelial preparation over the remaining media ± SEM (error bars) from four aortas. The relative abundance of the medial layer markers SMA and SM22 indicate the purity of endothelial preparations. (B) VEGFR3 reporter. Aortas from adult VEGFR3-driven YFP gene reporter mice were sectioned longitudinally and stained for the YFP reporter and for nuclei using Hoechst staining. IC, inner curvature. Images are representative of five mice from several litters. (C) VEGFR2 iΔEC. Endothelial-specific, inducible VEGFR3 knockout (iΔEC) and WT control mice were treated with tamoxifen, and aortas were removed after 1 wk. Tissue lysates were collected and immunoblotted with the indicated antibodies. IB, immunoblotting. (D and E) Inflammatory markers. VEGFR3 iΔEC and WT control mice were treated with tamoxifen and examined at 3 wk. Aortas were sectioned longitudinally and stained for fibronectin (D) or VCAM-1 (E). Images are representative of 6 mice from two independent experiments. Bars, 100 µm. The ratio of mean fluorescence intensity between the inner and outer curvature was then quantified. Values are means ± SEM (error bars). *, P < 0.05. Open circles denote outliers excluded from analysis by Grubbs’ test (α = 0.05).