Inhibition of uPA-induced cell adhesion by kininogen.

(A) uPA-induced U937 cell adhesion to VN was studied in the absence or presence of various concentrations of HK (open circles), HKa (filled circles), isolated domain 3 (filled triangles), domain 5 (open squares), or domain 6 (open triangles). (B) On a VN substrate, seeding of U937 cells together with uPA was performed in the absence (filled circles) and in the presence of 10 μg/mL HK (open circles) or 10 μg/mL HKa (filled squares), and the extent of cell adhesion was analyzed after various times as indicated. (C) After allowing the adhesion of U937 cells to VN in the presence of 50 nmol/L uPA for an incubation period of 90 minutes, 10 μg/mL HK (open circles), 10 μg/mL HKa (filled circles), or 100 nmol/L PAI-1 (open squares), respectively, was added, and the residual extent of cell adhesion was measured. Data are expressed as percentage of control, which is represented by the adhesion of cells in the absence of any stimulus or competitor. Data are mean ± SEM (n = 3) of a typical experiment; similar results were obtained in 3 separate experiments. (D) An adhesion assay with U937 myelomonocytic cells was performed in the absence (−) or presence of HK or HKa. After the incubation period for the adhesion assay (90 minutes), the supernatant was collected, centrifuged, and analyzed in a Western blot with the antibody HKH18, which is directed against domain 1 of kininogen and detects both HK and HKa. As a control, parallel wells without cells were incubated with the adhesion medium and HK or HKa. Molecular weight markers are indicated at the right margin. Similar results were observed in 3 separate experiments.