FIGURE 3.

hCD177-induced suppression in neutrophil is mediated at least partially by the binding of CD177 to CD31. PEC and PEC/hCD177 were plated at a concentration of 6 × 10⁴ cells/well as target cells in a 24 well gelatin-coated plate. After culturing for 24 h, 3 × 10⁵ neutrophils were added to each well. Neutrophils were treated for 1 h with vehicle (PEC, PEC/hCD177) or 200 μg/mL of anti-human CD31 (α CD31) or isotype mouse IgG (Iso). Cells were cocultured for 2 h in the presence of 200 nmol/L PMA, followed by triple-color staining with Annexin V-FITC, 7-AAD, and APC-conjugated anti CD11b antibody. The % cytotoxicity was calculated as (% Annexin V⁺ 7-AAD⁺ cells) + (% Annexin V⁺ 7-AAD^– cells) + (% Annexin V^– 7-AAD⁺ cells). The bars indicate the SEM. *P < 0.05, **P < 0.005. APC, allophycocyanin; CD177, cluster of differentiation 177; CD31, cluster of differentiation 31; FITC, fluorescein isothiocyanate; N.S., no significant difference, n = 8; PEC, porcine endothelial cells; PMA, phorbol 12-myristate 13-acetate.