Fig. 5

HPMEC were stimulated with 20 ng/ml of VEGF₁₆₅a, VEGF₁₆₅b or without stimulation (Ctl/control). Representative western blots showing VEGF stimulation at 5, 10 and 60 min plus loading control (α-tubulin) with densitometry (n = 3). a Western blot of phosphorylated p-38MAPK in HPMEC. VEGF₁₆₅a significantly increased phosphorylation of p-38MAPK at 5 and 10 min where VEGF₁₆₅b only induced increased phosphorylation at 5 min. b & c: Measurement of HPMEC resistance by Endohm system. Dotted lines correspond to cell pre-incubated with p38 inhibitor SB203580. SB203580 inhibited the effect of VEGF₁₆₅a (b) and the effect of VEGF₁₆₅b (c). d Graphs represent the mean of the scratched area for HPMEC treated with SB203580 inhibitor at 24 h relative to control. VEGF₁₆₅a but not VEGF₁₆₅b stimulation significantly diminished scratched area (p = 0.002) and was inhibited by SB203580 (p = 0.02). e Western blot of phosphorylated AKT in HPMEC. VEGF₁₆₅a and VEGF₁₆₅b significantly increased phosphorylation of p-Akt at 5 and 10 min. f & g Measurement of HPMEC resistance by Endohm system. Dotted lines correspond to cell pre-incubated with AKT inhibitor LY294002. LY294002 inhibited the effect of VEGF₁₆₅a (f) and the effect of VEGF₁₆₅b (g). Densitometry and scratch assay analysed by Kruskal-Wallis test with post hoc Dunn’s analysis. TEER measurement analysed using two-way ANOVA and Bonferroni post-test. All data were plotted as mean ± SEM (n = 3-6)