Effect of SSL5 on rolling adhesion of neutrophils under shear conditions. (A) Neutrophils were treated with SSL5 (1-30 μg/mL) for 15 minutes at 37°C. Subsequently, the neutrophils were perfused over glass coverslips coated with 10 μg/mL P-selectin/Fc at a shear stress of 1.6 dyn/cm2 during 5 minutes at 37°C. Rolling on the P-selectin/Fc surface was observed in 95% of untreated cells. After washing for 1 minute, accumulated neutrophils were quantified. (B) Effect of 10 μg/mL SSL5 on neutrophil rolling on P-selectin/Fc compared with the effect of 10 μg/mL anti–PSGL-1 KPL1 and the isotype control mAb W17/1 directed against the C5aR. (C) Effect of SSL5 on neutrophil rolling on activated endothelial cells. Freshly isolated HUVECs were cultured on glass coverslips. After stimulation of HUVECs with 100 μM histamine for 3 minutes, neutrophils treated with 10 μg/mL SSL5, SSL7, KPL1, or W17/1 were perfused at 0.8 dyn/cm2 for 5 minutes. After washing for 1 minute, accumulated neutrophils were quantified. The data represent relative accumulation of neutrophils compared with control-treated cells and are mean values ± SEMs of at least 3 independent experiments.
