Molecular diagosis of von Willebrand disease

von Willebrand disease (vWD) is one of the most common inherited human bleeding disorders, which is caused by quantitative or qualitative defects of von Willebrand factor (vWF). vWF is a highly multimerized glycoprotein that promotes platelet adhesion and aggregation at a high shear rate, while also acting as a carrier of coagulation factor VIII. vWD has been subdivided into three categories, which reflect their pathophysiology. Type 1 and type 3 vWD reflect partial or complete deficiency of vWF, whereas type 2 vWD reflects qualitative defect of vWF. The ability of vWF to interact with its platelet receptor and factor VIII, and the analysis of the multimeric composition of vWF are essential to identify patients with different vWD subtypes. The prevalence of different vWD subtypes was reported in the literature. In the past years, ninety-one patients with vWD were consulted in our institution. Of all the vWD patients, 56 (61.5%) belong to type 1, 26 (28.6%) type 2 and 9 (9.89%) type 3. The analysis of vWF gene was performed in some type 2 and type 3 vWD by denature gradient gel electrophoresis and sequencing. We have found six cases of point mutations of vWF gene, Ala737-->Glu, Gly 22-->Glu, Met37 Val and Ser71-->stop codon. Substitutions, are first reported in international database. We constructed an expression plasmid pSVA737EvWF containing full length of cDNA of vWF which included the Ala737 Glu substitution by site-direct mutagenesis. The structure of recombinant vWF within transfected COS-7 cells and the secretion of high-molecular-weight (HMW) multimers were similar to wild-type vWF. HMW forms of vWF multimers were absent in plasma but present in platelets. The mutation corresponds to the group II type 2A vWD characterized by normal secretion of all vWF multimers.