AMD3465 inhibits migration and intracellular signaling in AML cell lines. (A) Surface expression of CXCR4 was measured by flow cytometry, and the results are expressed as percent change in the mean fluorescent intensity (MFI) compared with control (untreated) cells. (B) U937 and MOLM13 cells (0.5 × 106) were plated onto the upper chamber of transwell plates and exposed to 50 ng/mL SDF-1α in the lower chamber or to 0.1 × 106 MS-5 cells preplated in the lower chamber with or without 1 μM AMD3465 for 24 hours. Migrating cells were counted after 24 hours of incubation. The results are expressed as a percentage of the migrating cells relative to the numbers of input cells. (C) Cells were pretreated with or without SDF-1α for 30 minutes, followed by exposure to 1 μM AMD3465 for 4 hours. Phosphorylation of Akt (pAkt) and Erk (pErk) was detected by Western blot analysis, and the intensity of the bands was quantified by densitometry and displayed as ratios of either phospho-proteins to total proteins. Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) was used as a loading control.
