Fig. 3

AMG9810 suppresses CXCR4 expression and activity in MM cells. Effect of AMG9810 treatment (10 µM, 1 h, 37 °C) on cell surface CXCR4 expression in RPMI8226, RPMI8226-CXCR4, OPM-1 and primary CD138 + MM cells, measured by flow cytometry. a Representative histograms demonstrating CXCR4 levels in non-treated and AMG9810-treated cells. b CXCR4 expression level is shown as mean fluorescent intensity (MFI), data are presented as mean of triplicates ± SD (**p < 0.01). c RMI8226-CXCR4 cells were pre-treated with indicated doses of AMG9810 for 30 min, and their migratory response to CXCL12 (200 ng/ml) was measured using the trans-well migration assay. d Effect of pre-treatment with AMG9810 (10 µM for 1 h at 37 °C) on CXCL12-induced (200 ng/ml, 30-min activation) intra-cellular signaling in RPMI8226-CXCR4 and OPM-1 cells. Levels of phosphorylated Erk1/2 and AKT proteins, measured by Western blot, β-actin was used as internal control. Representative data from at least two independent experiments are shown. e, f Effect of capsaicin treatment (10 µM, 1 h, 37 °C) on cell surface CXCR4 expression in RPMI8226, RPMI8226-CXCR4 and OPM-1 cells, measured by flow cytometry. g RMI8226-CXCR4 cells were pre-treated with indicated doses of capsaicin (10 µM) for 30 min, and their migratory response to CXCL12 (200 ng/ml) was measured using the trans-well migration assay. h CFSE-prelabeled RPMI8226 and RPMI8226-CXCR4 cells were pre-treated with AMG9810 (10 µM) or capsaicin (10 µM) for 30 min and then co-incubated with BMSC monolayer during 30 min; non-adherent cells were removed, and adherent CFSE-positive cells were enumerated using flow cytometry and expressed as percent of total input cells. Data are presented as mean of triplicates ± SD (**p < 0.01)