Inhibitory effect of RAP on NDSK-II–induced transendothelial migration of neutrophils in vitro. HUVECs were grown to confluence on gelatin-coated cell culture inserts. Calcein AM–labeled HL-60 cells differentiated into neutrophil-like cells were added to the upper chambers on top of the HUVEC monolayer in the presence of PBS used as a control or 1.5μM NDSK-II without or with 0.5μM RAP or 10μM (β15-66)2; RAP and (β15-66)2 at the same concentrations without NDSK-II were also added. The cells were allowed to migrate into the lower chambers containing chemoattractant fMLP for 4 hours at 37°C, collected, and measured by fluorescence at 530 nm. The results are expressed as percentage of HL-60 cells migrated in the presence of PBS (control). The graph shows combined data obtained from 2 independent experiments; bars denote means ± SEs (n = 6-9 wells for each condition).
