Figure 5.

LDN193189 reduces the BMP9- and BMP10-induced up-regulation of E-selectin, VCAM-1, and ICAM-1 and monocyte recruitment in TNFα-stimulated HAECs. HAECs were pre-treated with LDN193189 (250 nm resuspended in DMSO), DMSO, then stimulated with BMP9 or BMP10 (5 ng/ml; 16 h) prior to TNFα treatment (0.05 ng/ml for 4 h). Surface expression of E-selectin (FITC-conjugated anti-human E-selectin) (A), VCAM-1 (PE-Cy5-conjugated anti-human VCAM-1) (B), and ICAM-1 (APC-conjugated anti-human ICAM-1) (C) was assessed using flow cytometry. Forward scatter and side scatter gating was applied to the HAEC population. Data are shown as median fluorescence intensity (MFI) expressed as fold-change relative to HAECs transfected with siRNA control pool (siCP). D, HAECs were treated as described above and then monocytes were perfused in a flow adhesion assay. E, monocyte behavior (rolling, clear bar; adherence, gray bar; and transmigration, black bar) was expressed as a percentage of total recruitment. Experiments were performed in triplicate and the data are representative of n = 3 biological repeats. Error bars represent mean ± S.E. *, p ≤ 0.05; **, p ≤ 0.01; ***, p ≤ 0.001.