Fig. 2.

Identification of receptors and ligands required for flow-induced Smad2/3 activation. (A) ECs were subject to KD of the indicated receptors and exposed to flow (12 dynes/cm²); activation of Smad2/3 was assayed by Western blotting. (B) Alk5 KDs were rescued by reexpression of siRNA-resistant Alk5 and flow activation of Smad2/3 assayed; n = 3. (C) Nrp1 KD was rescued by reexpression of siRNA-resistant Nrp1 and flow activation of Smad2/3 assayed. For quantification, n = 3. (D) ECs were stimulated by the addition of BMP9 in the presence or absence of FSS at 12 dynes/cm² for 30 min, and p-Smad2/3 was assayed by Western blotting; n = 3. (E) HUVECs in 2% serum were exposed to laminar FSS for 2 h without (2 h FSS) or with (2 h FSS+) blocking antibodies to BMP9 (n = 4) or BMP10 (n = 3), as indicated. Samples were then assayed for p-Smad. Quantified values are means ± SEM *P < 0.05, **P < 0.01, and NS: not significant, calculated by one-way ANOVA with Tukey’s multiple comparison tests.