Direct binding of rhAPC to neutrophil integrins. (A) Binding of rhAPC to human neutrophils was assayed using chromogenic substrate S-2366 in the presence of 1 mM MnCl₂ ± cyclic RGD peptide (20 μg/mL), β₁ blocking Ab, β₃ blocking Ab, or β₂ blocking Ab (10 μg/mL each). *P < .05 versus MnCl₂-treated cells. (B) Binding of control or 10 nM fMLP-treated neutrophils to immobilized FN in the presence of rhAPC (10 μg/mL) or Gla-less APC (10 μg/mL), or an equivalent amount of PBS. (C) Binding of control or 10 nM fMLP-treated neutrophils to immobilized FN in the presence of rhPC (10 μg/mL) or an equivalent amount of PBS. (D) Induction of LIBS epitopes by rhAPC. Control (PBS) or rhAPC-treated neutrophils were incubated with the indicated concentrations of MgCl₂ and CaCl₂. The LIBS of β₁ and β₃ integrins were detected by B44 and D3 mAb, respectively. *P < .05 versus PBS-treated cells. (E) Solid-phase binding of immobilized α₃β₁, α₅β₁, and α_Vβ₃ to FN or rhAPC. (F) Solid-phase binding of immobilized α₃β₁, α₅β₁, and α_Vβ₃ to wild-type or mutant rhAPC (RGE-APC). (G) Migration of human neutrophils on FN-coated cover glass in the presence of fMLP ± RGE-APC, rhPC, Gla-less APC, or S360A-APC. Experiments were repeated on neutrophil preparations from 3 independent donors. Results in panels A and C through F are expressed as mean (± SEM) of 3 independent experiments.