Effects of clustering versus ligation of VLA-4 on HL60 cell adhesion to HUVECs.
(A) Myelomonocytic HL60 cells were preincubated with medium alone (control, not shown), isolated Fab fragments of mAb K20 (5 μg/mL), or mAb K20 (20 μg/mL) for 30 minutes at 37°C. Cells were washed, and cells preincubated with K20 Fab were further incubated with increasing concentrations of a secondary goat (Fab)2 antimouse Fab for 30 minutes at 37°C. After washing, adhesion to endothelial cell monolayers was performed. (B) HL60 cells were preincubated (20 minutes, 37°C) with medium (open bars) or human IgG (20 μg/mL, filled bars) to saturate Fc receptors. Medium alone (control, not shown), soluble VCAM-1 (sVCAM-1; 40 μg/mL), or soluble VCAM-1–Fc (sVCAM-1–Fc; 40 μg/mL) was added for 30 minutes at 37°C. Cells were washed, and cells preincubated with soluble VCAM-1 were further incubated with increasing concentrations of mAb anti–VCAM-1 for 30 minutes at 37°C. Following washing, adhesion to endothelial cell monolayers was performed. Values are displayed as percentage of control and represent the mean ± SEM of at least 3 independent experiments. *Indicates P < .01 compared with the respective control.
