ADAM10 promotes migration and fibronectin binding of human neutrophils and murine BMDMs. (A-C) Human neutrophils were pretreated with 10 µM GI254023X or 0.1% DMSO or left untreated for 15 minutes and were subsequently assayed for (A) cell migration, (B) adhesion to fibronectin, and (C) α5 integrin surface expression, induced by CXCL8 (10 nM). (D) Murine BMDMs were generated from wild-type mice treated with GI254023X (10 µM) or DMSO (0.1%) and investigated for chemotaxis induced by various concentrations of CCL2. (E-F) Murine BMDM from Vav-Adam10−/− and Vav-Adam17−/− mice and litter controls were pretreated with 10 µM GI254023X or vehicle (DMSO) and investigated for (E) CCL2-induced cell migration and (F) adhesion to fibronectin. (G) ADAM10-deficient BMDMs were transfected with bovine ADAM10, treated with 10 µM GI254023X or 0.1% DMSO, and investigated for CCL2-induced adhesion to fibronectin after 15 minutes. (H) BMDMs from Vav-Adam10−/− and litter control mice were stimulated with murine CCL2 (3 nM) for the indicated time periods and investigated for phosphorylation of p38 by western blotting. Samples were run on the same gel. (I) BMDMs from Vav-Adam10−/− and litter controls were treated for 15 minutes with CCL2 and investigated for upregulation of α5 integrin surface expression. (J) BMDM from Vav-Adam10−/− and litter controls were treated with 3 nM CCL2 for the indicated time periods and examined for polymerization of F-actin by flow cytometry. Quantitative data represent means ± SEM of 3 independent experiments and cell preparations. Crosses indicate significance among treated cells calculated using 1-way ANOVA and the Bonferroni post-test. Asterisks without a line indicate significant differences to the nontreated control analyzed by 1-sample t test.
