Supra-normal closure time caused by recombinant von Willebrand factor in PFA

T. Kragh, M. Spannagl (Munich, Germany)

Platelets - Physiology and Disorders of platelet number and function
Date: 17.02.2017,
Time: 17:15 - 18:15

Objective: High molecular weight (HMW) von Willebrand factor (VWF) multimers play a crucial role in primary hemostasis. It was shown that ultralarge multimers (UL) are even more important for VWF-platelet interaction. Especially von Willebrand disease (VWD) patients lacking HMW multimers (type 2+3) show specific bleeding symptoms and have to be treated with factor supplements. In vitro we simulated basic, shear dependent processes of primary hemostasis and showed the influence of VWF supplements on VWD patient’s whole blood.

Methods: Citrate anticoagulated whole blood from healthy donors and severe VWD patients was obtained under standardized conditions and tested. The platelet functional analyzer (PFA-200) measures functional primary hemostasis mimicking the vascular lesion including flow dynamic effects. In the capillary a collagen coated membrane is perfused with increasing shear rates beginning at 5k s^-1. Several platelet agonists (ADP and epinephrine) start the adhesion and aggregation reaction on the surface. Then recombinant VWF containing UL (rVWF/+UL; Baxalta, Vienna, Austria) or plasma derived VWF products were added (1 IU/ml) and the influence on the blood closure time determined. Additionally the effects of high shear (30k s^-1) on VWF spiked whole blood over collagen surface were video documented with flow chamber experiments.

Results: The healthy control group showed PFA results in normal range (ADP 68-120s; EPI 84-170s). Aggregate formation in the flow chamber was 5-25%. Patients lacking large VWF multimers had no closure in the PFA and no aggregation in the flow chamber. Adding rVWF/+UL to patient’s blood supra-normalized the blood closure time and promoted normal to increased aggregate formation on collagen surfaces (ADP 50-70s; EPI 60-90s). On the contrary plasma derived VWF products hardly changed results when added to the blood samples lacking VWF (VWD patients type 2+3).

Conclusion: We demonstrated and visualized that UL are important for primary hemostasis and can establish VWF-platelet interaction in VWD patient blood leading to normal aggregate formation. Independent of the sample’s VWF:Ag value UL decreases blood closure time to supra normal levels. Equimolar doses of plasma derived VWF do not show these effects. Nonetheless all VWF products show good results in patient application.