Distinct abnormalities in the interaction of purified types IIA and IIB von Willebrand factor with the two platelet binding sites, glycoprotein complexes Ib-IX and IIb-IIIa.

Abstract

IntroductionWe have studied the interaction of the congenitally abnormal type IIA and IIB von Willebrand factor (vWF) molecules, both lacking the larger multimeric forms, with the two vWF binding sites on platelets, the glycoprotein (GP) Ib-IX and GP (8); in the latter, vWF has increased affinity for platelets and, consequently, the larger multimers are removed from the circulation as a result ofplatelet binding (7, 9-1 1). To date, however, there is no detailed information on the modalities of interaction of IIA and IIB vWF with the corresponding binding sites on platelets. The association of vWF with both GP Ib-IX and GP IlbIla appears to be important in thrombus formation (12, 13), and knowledge ofthe molecular mechanisms responsible for it is likely to provide useful information for understanding the processes involved in normal hemostasis as well as pathological vascular occlusion. In this regard, the study of molecular variants of von Willebrand disease is of particular interest if the existence of specific molecular defects can be correlated to relevant abnormalities of interaction with platelets. To address these issues, we have purified the congenitally abnormal vWF molecule from the plasma of patients with type IIA and IIB von Willebrand disease, and evaluated in a quantitative manner the interaction with GP Ib-IX and GP Ib-IhIIa. Our findings demonstrate that IIB vWF can bind to GP Ib-IX in the absence of any mediating or activating molecule, suggesting the existence of a molecular defect resulting in increased affinity for this platelet site. Such increased affinity is also evident in the presence of ristocetin, in spite of the absence of larger multimers. In contrast, IIA vWF interacts with GP Ib-IX with extremely reduced affinity. The severity of this abnormality may reflect the existence of specific structural alterations af-

O-linked carbohydrate of recombinant von Willebrand factor influences ristocetin-induced binding to platelet glycoprotein 1b.

“…A similar mechanism was recently implicated in type IIB vWd, where the decrease in both circulating platelets and the larger vWf multimers from plasma is attributable to spontaneous binding between soluble vWfand platelets ( 18,58). It has been reported that type IIA vWf may have reduced affinity for GP lb as well (45 ). It might be expected that any defect in vWfthat reduced the extent of the interaction with GP lb might also induce a clinically significant bleeding disorder.…”

“…Botrocetin, however, is not known to be a protein-precipitating agent, is reactive with all multimeric forms of vWf, and promotes aggregation with platelets and plasma from many mammalian sources (42)(43)(44). Unlike ristocetin, botrocetin is active with vWf from patients with type IIA vWd (41), which may have a lowered affinity for GP lb compared with native vWf (45). The mechanisms of action by which these agents induce vWf-dependent platelet aggregation may differ as well.…”

von Willebrand factor mutation enhancing interaction with platelets in patients with normal multimeric structure.

“…For example, botrocetin, a snake venom protein, forms a bimolecular complex with vWF that then binds to GP lb with high affinity; botrocetin has been shown to bind to three different segments ofsequence within the above mentioned disulfide loop (46). In type IIB von Willebrand disease, characterized by increased vWF affinity for GP lb (5,6), all the reported mutations considered to be responsible for the functional abnormality are within the same disulfide loop; moreover, six of them are in the first botrocetin-binding segment and two in the second one (3, 1 1-13, 46). Porcine vWF, which spontaneously aggregates human platelets, displays interesting differences as compared to human vWF in the region corresponding to the first botrocetinbinding segment: in fact, it contains three amino acid substitutions (47) corresponding to those found in human type IIB vWF, which also aggregates normal human platelets (43,44).…”

“…In the type IIB variant of von Willebrand disease, vWF in solution displays enhanced affinity for GP Ib, as shown either directly with the purified protein (5) or by the induction of aggregation in platelet-rich plasma at ristocetin concentrations lower than those effective in normal samples (6). The affected patients also exhibit absence of the largest vWF multimers in plasma, probably owing to their spontaneous binding to circulating platelets (7).…”

Heterogeneity of plasma von Willebrand factor multimers resulting from proteolysis of the constituent subunit.

“…6). Thus, a single cleavage occurring in the middle of a long oligomer may have profound effects on its size and, because of the known correlation between size and affinity for platelet receptors (30,31), may alter its function in supporting platelet adhesion and aggregation (1).…”