Fig. 3

ENDOA2 controls endothelial cell migration and polarity. a VEGF-induced cell proliferation (6 nM, 48 h stimulation) assessed by XCelligence system in siControl (Ctrl) and siENDOA2 silenced HUVECs (N = 4 independent experiments; two-way ANOVA: ns P > 0.05, **P < 0.01). b Cleaved caspase-3 staining of HUVECs cultured in 0.5% FBS for 24 h (N = 3 independent experiments; Mann–Whitney U test: ns P > 0.05). c HUVEC scratch wound migration in response to VEGF (3 nM). d Quantification of wound closure shown in c (N = 5 independent experiments; Mann–Whitney U test: **P < 0.01). e Phalloidin, Dapi, and GM130 Golgi labeling at the scratch wound edge to assess Golgi polarization in front of the nucleus (arrows indicate direction of migration) in response to VEGF (3 nM, 2 h). f Quantification of Golgi orientation (N = 5 independent experiments; Mann–Whitney U test: *P < 0.05). g Western-blot analysis of phosphorylation of the indicated proteins in response to VEGF (1.5 nM) in HUVECs. h Quantification of phosphorylation normalized to total protein levels (N = 4–5 independent experiments; two-way ANOVA: *P < 0.05, **P < 0.01, ***P < 0.001, ns P > 0.05). i Western-blot analysis of VEGF-induced (1.5 nM) PAK and ERK phosphorylation in mLEC from EndoA2^+/+ and EndoA2^−/− mice. Each lane represents one mouse. j Quantification of phosphorylation normalized to total proteins (N = 3–5 mice; two-way ANOVA: ns P > 0.05, **P < 0.01). Error bars represent mean ± s.e.m. Scale bars: c 200 μm, e 25 μm