Interfering with ALK-5 and TGF-βRII abolishes the inhibitory effect of GDF-15 on myeloid cell adhesion. PMNs (A,C) or THP-1 cells (B,D-G) were pre-incubated with the inhibitors SB-431542 (1 μM) (A-B) or GW-788388 (200 nM) (C-D), or 50 μg/mL anti–hTGF-βRII blocking antibody or control antibody (F), or GW-788388 (200 nM) in combination with 50 μg/mL anti–hTGF-βRII blocking antibody, or control antibody (G) for 10 minutes followed by 20 minutes preincubation with hGDF-15 (100 ng/mL) at 37°C. Where indicated, CXCL1 or CCL2 was added for 1 minute. Subsequently, cells were allowed to bind to immobilized mouse ICAM-1-Fc or mouse EM-Fc (A,C), or to hVCAM-1-Fc or control hIgG1 (B,D,F-G). (E) THP-1 cells were treated with ctrl siRNA or ALK-5 siRNA (as indicated) for 3 days. Resulting expression levels of ALK-5 were analyzed by immunoblotting, and quantification is shown beside. Adhesion of siRNA-treated THP-1 cells to immobilized VCAM-1-Fc or hIgG1 was analyzed as for (B,D,F-G). Experiments were done 4- to 9-times and for each adhesion assay 6 wells were analyzed per condition. Data are shown as mean ± standard error of the mean (SEM). *P < .05; **P < .01; ***P < .001. ab, antibody; ctrl, control; h, human; m, mouse; n.s., not significant.