Innovative methodology: The development of a method to measure flow-based thrombin generation

Occlusion of a vessel is a dynamic process in which several different systems
collaborate in an organized fashion. A too big change in any of these systems
(vessel, platelets and coagulation factors) can lead to either a bleeding or
thrombotic phenotype. To quantify such changes and thereby detect pathogenic
phenotypes several assays can be used. Thrombin generation is a widely used
assay to detect a prothrombotic phenotype due to a hyper-active coagulation
system. It has repeatedly been shown that increased thrombin generation is
associated with myocardial infarction and has prognostic value. A higher
potential to generate thrombin is related with thrombosis and a lower one with
bleeding. With respect to arterial thrombosis, where platelets and other blood
cells play a much larger role than in venous thrombosis, it is an advantage of
this assay that it can also measure the interaction between coagulation and
platelets as it occurs in platelet-rich-plasma. Recent developments in our
laboratory even allow measurement in whole blood.
Furthermore, as soon as platelets are involved flow also starts to play an
important role. We have found that in flow platelets need VWF to exert their
full prothrombotic character. VWF seems to be a mediator between fibrin and
platelets.
Because of the importance of platelets, other blood cells and flow, we would
like to expand the current thormbin generation method by measuring hemostasis
in whole blood under flow to explore whether we are able to better detect and
predict a prothrombotic phenotype thereby its risk on clinical symptoms such as
a myocardial infarction.

The objective of this study is to investigate whether thrombin generation is
effected by different shear rates and different tissue factor concentrations in
plasma, platelet rich plasma and whole blood. Thrombin generation in plasma
with/without platelets is a well established method for the detection of a
prothrombotic or bleeding phenotype. Recently a method was developed to measure
thrombin generation in whole blood, thereby taking into account the total
cellular compartment of the blood. We would like to expand this assay be
including the flow (=shear) that also normally occurs in a blood vessel.
Therefore we will test thrombin generation under flow in whole blood, platelet
rich plasma and plasma alone. This will be done for 4 different shear rates
with 4 different tissue factor concentrations.

This study is of invasive design. However, the impact on the subjects will be
minimal. Blood will be drawn from healthy volunteers (male and female) between
the ages of 18 and 65 years. They will be recruited from Maastricht University
and Maastricht UMC+. Before blood collection an informed consent will be
signed. For all study objectives described above blood from healthy volunteers
is needed. For every study objective an estimated 10 blood donations are needed
(in total 120 blood donations over a period of 4 years).

The venipunctures will be made by experienced coworkers. Nevertheless, blood
sampling causes local bruising, and incidentally a hematoma can be formed.
There will be no direct benefit of the subjects.