Figure 5

Effect of Adducin on cAMP-mediated endothelial barrier response. (A) Confluent WT and Add-KO cell monolayers were treated with Vehicle or F/R. The TER was measured up to 3 h after the treatment. The red-dashed line indicates the time of application. Data are normalized to the initial point of each corresponding group. “*” denotes significant difference between Vehicle and F/R in WT cells (N = 3 per group). Immunostainings for (B) VE-cadherin, β-catenin and (C) claudin-5, ZO-1 in WT and Add-KO cells treated for 1 h with either Vehicle or F/R. In WT cells arrows notice areas on the membrane where thicker signal is observed, arrowheads highlight finger-like projections. In KO cells arrows indicate fragmented junctional staining (N = 3–4 per group). (D) Representative Western blots for key AJ and TJ components from confluent WT and Add-KO cell monolayers exposed to Vehicle or F/R for 1 h. The graphs represent the relative pixel intensity of the bands, normalized to the WT, vehicle controls (N = 4 per group). (E) The graph shows intracellular cAMP concentration in pmol/ml from confluent endothelial cells monolayers analyzed by cAMP-specific ELISA (N = 8 per group). Data are represented as mean ± SEM; *p < 0.05; **p < 0.01.