Requirement of a plasma fraction for the loss of large Von Willebrand factor multimers induced by high wall shear rate

RESUMO

The perfusion of serum, citrated whole blood and citrated plasma, through a simple tube system resulted in a significant loss of large von Willebrand factor (vWf) multimers, without decrease in antigen levels. Maximum loss of large multimers was observed at a shear rate of 15,000 s(-1) for 15 min. Heparin, aprotinin, soybean trypsin inhibitor, phenylmethylsulphonylfluoride, N-ethylmaleimide, leupeptin or calpain inhibitor peptide could not prevent the loss of large vWf multimers in citrated plasma. The addition of EDTA calcium salt partially prevented it, and it was totally prevented by EDTA without calcium. Perfusion of purified vWf did not induce the loss of large multimers, but this did happen after the addition of either whole serum or a plasma fraction. The activity of this plasma fraction disappeared at pH < 6.8. Besides, we have found that the binding to subendothelium of purified vWf diluted in dialized serum was lower at pH 7.2 than at pH 6.0. Chromatographic studies demonstrated that the loss of large vWf multimers, induced by high shear rates, involves a plasma substance(s) of molecular weight larger than 200 kD; calpain and granulocyte or cysteine proteases do not seem to be this plasma substance(s).