Costimulation induces neutrophil homotypic adhesion under shear. Neutrophils (2 × 106/mL) were preincubated with or without inhibiting reagents at room temperature for 3 minutes. Neutrophils were then stimulated with rPF4 (1 μM) or protamine sulfate (PS; 25 μg/mL) in the presence or absence of 10 μg/mL 197.2 or IgG1 isotype control at 37°C for 5 minutes prior to shear application at 1077/s. Mac-1 is up-regulated in this 5-minute interval. t = 0 designates time of shear application in this figure. (A) Kinetics of neutrophil homotypic aggregation. (B) Neutrophil homotypic aggregation inhibited by 1 U/mL heparin and 10 μg/mL of function blocking mAbs directed against CD32a (IV.3), L-selectin/CD62L (DREG56), or β2-integrin/CD18 (IB4). Error bars represent SEM (P ≥ 3).
