No abstract

• Vimentin expressed on the platelet surface serves as adhesive receptor for VWF.The interaction between platelet receptor glycoprotein Iba and the A1 domain of von Willebrand factor (VWF) mediates tethering/translocation of platelets to sites of vascular injury. Unexpectedly, we observed platelets translocating over A1A2A3 domains protein slower than on A1 domain at high shear stress. This observation suggests an additional interaction between A domains and an adhesive receptor. We investigated vimentin because we have data showing the interaction of vimentin with the A2 domain of VWF. Moreover, vimentin is expressed on the platelet surface. This novel interaction was analyzed by using purified VWF, recombinant proteins, anti-vimentin antibodies, parallel flow chamber adhesion assays, flow cytometry, and vimentin-deficient murine platelets. The active form of VWF bound to vimentin, and the purified A2 domain blocked that binding. The interaction of a gain-of-function A1A2A3 mutant with platelet was reduced using anti-vimentin antibody. Platelet adhesion to wild-type (WT) A1A2A3 protein, collagen, and fibrin(ogen) was inhibited (32-75%) by anti-vimentin antibody under high shear stress. Compared with WT mice, platelets from vimentin-deficient mice had a reduced flow-dependent adhesion to both collagen and purified murine VWF. Last, the vimentin knockout mice had a prolonged tail bleeding time. The results describe that platelet vimentin engages VWF during platelet adhesion under high shear stress. (Blood. 2014;123(17):2715-2721 Introduction Atherothrombotic events, including acute coronary syndrome and stroke, are the result of platelet adhesion and activation on the ruptured atherosclerotic plaques. This platelet-mediated arterial thrombosis starts with the contact of the rapidly flowing platelets to components of the damaged blood vessel. von Willebrand factor (VWF), a multimeric plasma and subendothelial glycoprotein, is relevant in mediating platelet adhesion and activation at sites of lesions in the coronary arteries, where high shear conditions prevail.1,2 VWF captures the circulating platelets through its interaction with the platelet receptor glycoprotein (GP)Ib/IX/V complex. This interaction is responsible for the tethering, rolling, and activation of platelets that eventually become firmly adhered, leading to thrombus formation within a coronary artery. 3,4 Mature VWF consists of a 2050-residue subunit formed by domains arranged in the order of D9-D3-A1-A2-A3-D4-B-C. 5 The A1 domain contains the binding site for the platelet receptor GPIba 6 ; the cleavage site for the enzyme ADAMTS (a disintegrin and metalloproteinase with thrombospondin motifs)-13 is localized in the A2 domain, 7 and the A3 domain binds to collagen. 8 Unlike the A3 domain, both the A1 and A2 domains do not have access to their ligands until their domain structure is altered.9 This structural modification can be induced by mutations, 10 hydrodynamic forces, 11 immobilization on a surface, or artificially with the modulator ristocetin....

Sickle cell anemia (SCA) is a common hemolytic disorder caused by a gene mutation in the β-globin subunit of hemoglobin (Hb) and affects millions of people. The intravascular hemolysis releases excessive amount of extracellular hemoglobin (ECHb) into plasma that causes many cellular dysfunctions in patients with SCA. ECHb scavenges NO which promotes crisis events such as vasoconstriction, thrombosis and hypercoagulation. ECHb and its degradation product, heme, are known to cause oxidative damage to the vessel wall and stimulate the expression of adhesive protein ligands on vascular endothelium. Our study shows that ECHb binds potently to VWF—largest multimeric glycoprotein in circulation—through the A2-domain, and significantly inhibits its cleavage by the metalloprotease ADAMTS13. Furthermore, a subpopulation of VWF multimers bound to ECHb exists in significant amount, accounting for about 14% of total plasma VWF, in SCD patients. The Hb-bound VWF multimers are resistant to ADAMTS13, and are hyperactive in aggregating platelets. Thus, the data suggest that Hb-bound VWF multimers are ultralarge and hyperactive because they are resistant to the protease. The Hb-bound VWF multimers are elevated parallely with the level of ECHb in patients' plasma, and is associated with the pathogenesis of thrombosis and vascular occlusion in SCA.