Figure 7
Changes in cell adhesion between U937 FPR (ΔST) and VCAM-1-transfected B78H1 cells at resting state and in response to receptor stimulation. A, U937 FPR (ΔST) cells were preincubated for 10 min at 37°C with forskolin (20 μM), or DMSO (vehicle). Cells were stimulated (arrow) with fMLFF (100 nM), or DMSO (vehicle). Data are plotted as % of U937 FPR (ΔST) singlets versus time. Five different experimental conditions are shown: Block/DMSO (preincubation with blocking 2 μM LDV small molecule (addition of DMSO, 1 μl), DMSO/DMSO, resting state (preincubation with DMSO/addition of DMSO), Forsk/DMSO (preincubation with forskolin/addition of DMSO), Forsk/fMLFF (preincubation with 20 μM forskolin/stimulation with 100 nM fMLFF), DMSO/fMLFF (preincubation with DMSO, stimulation with 100 nM fMLFF). The non-desensitizing FPR mutant is used to maintain VLA-4 in a state of constant high affinity. Mean and standard error of mean of three independent experiments are shown (n = 3). Bars on the right side of the panel indicate a relative contribution of affinity and extension in the overall cell adhesion. B, Effect of forskolin addition on adhesion in real-time. Cells were stimulated with fMLFF, or DMSO (1st addition). Next, forskolin (20 μM) was added. Representative experiment out of two experiments is shown.