EphrinB2/EphB4 signaling modulates endothelial proliferation

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EphrinB2/EphB4 signaling modulates endothelial proliferation EphrinB2/EphB4 signaling modulates endothelial proliferation A–DMuscles were harvested 3, 4, and 7 days after implantation of V‐low or V‐low sEphB4 clones, or V‐high cells while treating animals systemically with ephrinB2‐Fc or control Fc proteins. Endothelial proliferation was assessed by quantifying the percentage of endothelial cells positive for Ki67, which marks all cycling cells (A and C), or phosphorylated histone H3, which marks only cells in the G2/M phase (pHH3, B, and D), by immunofluorescence staining on frozen muscle sections. EphB4 inhibition specifically increased the rate of endothelial proliferation (pHH3+ cells) and its stimulation by ephrinB2‐Fc conversely decreased it. Mean ± SEM; n = 4 independent samples/group; *P < 0.05, **P < 0.01, and ***P < 0.001 (one‐way ANOVA with Bonferroni multiple comparisons test).E–GHuman dermal microvascular endothelial cells (HDMEC) were treated in vitro with recombinant VEGF or FGF2, while EphB4 was stimulated with ephrinB2‐Fc (50 or 2,000 ng/ml). Cell cycle analysis was performed by FACS after staining for Ki67 and pHH3 (E), and the proportion of cells withdrawn from cycle (G0) or in mitosis (M) were quantified (F, G). EphB4 stimulation dose‐dependently increased quiescence and decreased mitosis by both mitogens. Mean ± SEM; n = 3 independent samples/group; ***P < 0.001 vs. ephrinB2‐Fc control, #P < 0.05, ##P < 0.01 and ###P < 0.001 (one‐way ANOVA with Bonferroni multiple comparisons test). Elena Groppa et al. EMBO Rep. 2018;embr.201745054 © as stated in the article, figure or figure legend