In vitro correction of the abnormal multimeric structure of von Willebrand factor in type IIa von Willebrand's disease.

Abstract

Type Ha von Willebrand's disease (vWd) has been characterized by the absence of the largest and a reduction in the intermediate-sized multimers of the plasma and platelet von Willebrand factor (vWf) and by the diminished response of the platelet-rich plasma of these patients to ristocetin. Other recently demonstrated abnormalities include the presence of an abnormal triplet structure of vWf. We have studied the plasma and platelets from three patients with this form of vWd and have found that both their plasma and platelets manifest the previously described abnormalities. Because of the heterogeneity of the multimeric structure of the vWf in these patients, we considered the possibility that postsynthetic events may have modified the vWf. When blood was collected in 5 mM EDTA or 5 mM EDTA/leupeptin/N-ethylmaleimide, the abnormal multimeric structure of the plasma and platelet vWf was partially normalized in that the intermediate and the largest vWf multimers were increased, the abnormal multimer structure was no longer as apparent, and the fastest migrating band (an abnormality seen only in the type Ila vWd plasma and platelets) disappeared. The enzymatic activity responsible for this degradation can be classified as a calcium-dependent protease. Studies of normal radiolabeled vWf incubated with platelet lysates from normal subjects and these patients revealed that the patients' platelets did not contain increased amounts of calcium-dependent protease activity as assessed by degradation of normal vWf. These data suggest that patients with type Ha vWd synthesize an abnormal vWf protein that is susceptible to in vitro proteolytic degradation and that proteolytic degradation can play a significant role in the phenotypic expression of vWd by modifying the plasma and platelet vWf multimeric structure. von Willebrand's disease (vWd) has been classified by the pattern of the plasma von Willebrand factor (vWf) multimers on agarose gel electrophoresis and by the response of the patient's platelet-rich plasma (PRP) to ristocetin (1, 2). In particular, the group of type II patients show varied and abnormal electrophoresis patterns of their plasma vWf. These include absence of the largest and/or intermediatesized plasma vWf multimers and alterations in the triplet structure comprising each multimer (1-7). The presence of a triplet structure in normal subjects has suggested that the series of multimers is based on more than a single protomer, which polymerizes to form the various multimers (3, 4). The mechanism responsible for the abnormalities of the multimeric structure in vWd is generally thought to involve decreased or abnormal synthesis or release of vWf from the endothelial cell. The effects of DDAVP infusion on multimeric structure have supported this concept in the type Ila variants (8).We have studied the multimeric organization of the vWf multimers in these patients who fit the category of type Iha vWd. We report our findings, indicating that the abnormalities of the multimeric structure in these patients a...

Impact of mutations in the von Willebrand factor A2 domain on ADAMTS13-dependent proteolysis

“…Targeting ADAMTS13-dependent proteolysis of VWF might be a potential therapeutic approach in VWD. 17,31 …”

Epitope mapping of the von Willebrand factor subunit distinguishes fragments present in normal and type IIA von Willebrand disease from those generated by plasmin.

“…In type IIA von Willebrand disease (vWD) the relative proportion of the 176 and 140 kD fragments is increased, supporting the hypothesis that proteolysis is responsible, at least in part, for the absence of large multimers in that disorder (1,2). Plasmin, the platelet calcium-activated protease (calpain), and one or more enzymes released from polymorphonuclear leukocytes proteolyze vWF in vitro with resultant loss of large multimers (2)(3)(4)(5)(6). Thus, all are candidates for producing the loss of large multimers seen in type IIA vWD.…”

Genetic linkage of two intragenic restriction fragment length polymorphisms with von Willebrand's disease type IIA. Evidence for a defect in the von Willebrand factor gene.

“…Recently, new insight has been gained on the structure of vWF in the plasma of vWD type-IIA patients by Gralnick and co-workers (33). These authors show that the multimeric structure of vWF in the plasma of these patients is similar to that of healthy individuals, provided that blood is collected in a mixture of protease inhibitors.…”

“…Small multimers exhibit less GPIB binding activity (2). Alternatively, since Gralnick and co-workers showed that the vWFRco activity did not change significantly upon correction of the abnormal vWF type-IIA multimeric structure (33), the defect might be due to an alteration in the GPIB binding domain. The domain on the mature vWF subunit that is responsible for the interaction with GPIB has been localized on a tryptic fragment composed of aminoacid residues 449-728 (38).…”