Figure 1. Knockdown of Gα_s increases endothelial inflammation induced by disturbed flow in BAECs.

(A–D) Confluent BAECs were transfected with control siRNA or siRNA directed against Gα_s and were then exposed to oscillatory (osc.) flow for the indicated time period. p65 and/or CREB phosphorylation was analyzed by immunoblotting (A and B, n = 3 independent experiments), cellular p65 localization was determined by staining of cells with an anti-p65 antibody (C, n = 3 independent experiments, at least 3 view fields were analyzed per experiment), and inflammatory gene expression was analyzed by qPCR (D, n = 4 independent experiments). The diagrams show the densitometric evaluation of p65 phosphorylation (A and B) or of nuclear p65 staining (C). Scale bar: 20 μm. (E) Confluent BAECs were incubated without or with the PKA-inhibitor PKI (1 μM). Inflammatory gene expression after osc. flow induction was analyzed (n = 4 independent experiments). Data represent mean values ± SD; *P ≤ 0.05, **P ≤ 0.01, ***P ≤ 0.001 (2-way ANOVA and Bonferroni’s post hoc test [A, B, D, and E] and 2-tailed Student’s t test [C]).