Different mechanisms define the antiadhesive function of high molecular weight kininogen in integrin- and urokinase receptor-dependent interactions

Proteolytic cleavage of single-chain high molecular weight kininogen (HK) b y kallikrein releases the short-lived vasodilator bradykinin and leaves beh ind a-chain high molecular weight kininogen (HKa) that has been previously reported to exert antiadhesive properties as well as to bind to the urokina se receptor (uPAR) on endothelial cells. In this study we defined the molec ular mechanisms for the antiadhesive effects of HKa related to disruption o f integrin- and uPAR-mediated cellular interactions. Vitronectin (VN) but n ot fibrinogen or fibronectin-dependent alpha v beta(3), integrin-mediated a dhesion of endothelial cells was blocked by HKa or its isolated domain 5, I n a purified system, HKa but not HK competed for the interaction of VN with alpha v beta(3) integrin, because HKa and the isolated domain 5 but not HK bound to both multimeric and native VN in a Zn2+-dependent manner. The int eraction between HKa or domain 5 with VN was prevented by heparin, plasmino gen activator inhibitor-1, and a recombinant glutathione-S-transferase (GST )-fusion peptide GST-VN (1-77) consisting of the amino terminal portion of VN (amino acids 1-77), but not by a cyclic arginyl-glycyl-aspartyl peptide, indicating that HKa interacts with the amino terminal portion of VN ("soma tomedin B region"). Furthermore, we have confirmed that HKa but not HK boun d to uPAR and to the truncated 2-domain form of uPAR lacking domain 1 in a Zn2+-dependent manner. Through these interactions, HKa or its recombinant H is-Gly-Lys-rich domain 5 completely inhibited the uPAR-dependent adhesion o f myelomonocytic U937 cells and uPAR-transfected BAF-3 cells to VN and ther eby promoted cell detachment. By immunogold electron microscopy, both VN an d HK/HKa were found to be colocalized in sections from human atheroscleroti c coronary artery indicating that the described interactions are likely to take place in vivo. Taken together, HK and HKa inhibit different VN-respons ive adhesion receptor systems and may thereby influence endothelial cell- o r leukocyte-related interactions in the vasculature, particularly under Inf lammatory conditions. (C) 2000 by The American Society of Hematology.