High Molecular Weight Kininogen Regulates Platelet-Leukocyte Interactions by Bridging Mac-1 and Glycoprotein Ib

Leukocyte-platelet interaction is important in mediating leukocyte adhesion to a thrombus and leukocyte recruitment to a site of vascular injury. This interaction is mediated at least in part by the β2-integrin Mac-1 (CD11b/CD18) and its counter-receptor on platelets, glycoprotein Ibα (GPIbα). High molecular weight kininogen (HK) was previously shown to interact with both GPIbα and Mac-1 through its domains 3 and 5, respectively. In this study we investigated the ability of HK to interfere with the leukocyte-platelet interaction. In a purified system, HK binding to GPIbα was inhibited by HK domain 3 and the monoclonal antibody (mAb) SZ2, directed against the epitope 269–282 of GPIbα, whereas mAb AP1, directed to the region 201–268 of GPIbα had no effect. In contrast, mAb AP1 inhibited the Mac-1-GPIbα interaction. Binding of GPIbα to Mac-1 was enhanced 2-fold by HK. This effect of HK was abrogated in the presence of HK domains 3 or 5 or peptides from the 475–497 region of the carboxyl terminus of domain 5 as well as in the presence of mAb SZ2 but not mAb AP1. Whereas no difference in the affinity of the Mac-1-GPIbα interaction was observed in the absence or presence of HK, maximal binding of GPIbα to Mac-1 doubled in the presence of HK. Moreover, HK/HKa increased the Mac-1-dependent adhesion of myelomonocytic U937 cells and K562 cells transfected with Mac-1 to immobilized GPIbα or to GPIbα-transfected Chinese hamster ovary cells. Finally, Mac-1-dependent adhesion of neutrophils to surface-adherent platelets was enhanced by HK. Thus, HK can bridge leukocytes with platelets by interacting via its domain 3 with GPIbα and via its domain 5 with Mac-1 thereby augmenting the Mac-1-GPIbα interaction. These distinct molecular interactions of HK with leukocytes and platelets contribute to the regulation of the adhesive behavior of vascular cells and provide novel molecular targets for reducing atherothrombotic pathologies.