Serum proteins modulate CD13 dependent endothelial invasion in vitro. (A) Primary endothelial cells (HUVECs) were plated in the top chamber of Matrigel-coated transwell plates in the presence (▩) or absence (□) of added growth factors or 10% FBS (■) and the number of cells invading through the Matrigel barrier assessed after 24 hours. The CD13 functional antagonists bestatin (100 μg/mL) and MY7 antibody (1:100 dilution; CD13 mAb) were added to medium containing 10% FBS. (B) Invasion was assessed in the presence of increasing doses of the CD13 antagonists or irrelevant inhibitors or noninhibitory CD13 antibodies. The highest concentration of each agent used is set at 100%. For bestatin (○) and soybean trypsin inhibitor (□), 100% equals 100 μg/mL, while for antibodies MY7 (●; neutralizing mAb) and WM4.7 (■; noninhibitory mAb) 100% equals 1:40 dilution. (C) FBS was fractionated by gel filtration, and fractions were assayed for their ability to induce endothelial invasion. (D) Invasion was assessed in the presence of increasing concentrations (“Materials and methods”) of the plasma components fibronectin (♦), vitronectin (▾), plasminogen (▴), fibrinogen (□), high-molecular-weight kininogen (HK; ●), and low-molecular-weight kininogen (○), LDL (*), and HDL (▵). (E) HK (100 μg/mL; ▩)–induced invasion was assessed in the presence of the CD13 inhibitors bestatin or MY7 antibody (CD13 mAb, 1:100 dilution) or the bradykinin B₂ receptor antagonist HOE140 (10 μM). (F) Effect of increasing concentrations of B₂ receptor antagonist HOE140 on invasion induced by 10% FBS. Relative invasion in treated cells is expressed as percentage of controls. Data are shown as means plus or minus a standard deviation (SD), n = 3.