Summary
We have made use of a novel flow reactor to study the initiation and propagation of
the ex vivo blood coagulation processes at artificial surfaces. The flow reactor consisted
of a primary glass or polymer capillary that is connected to a secondary glass capillary,
which inner wall was coated with a phospholipid bilayer of 25 mol% dioleoylphosphatidylserine/75
mol% dioleoylphosphatidylcholine (DOPS/DOPC). Citrated platelet free plasma and a
CaCl2 solution were delivered by syringe pumps and mixed just before the entrance of the
flow reactor. The outflowing plasma was assayed for factor XIa, factor IXa, factor
Xa and thrombin activity. Perfusion of recalcified plasma through a bare glass capillary
resulted in a transient generation of fluid phase factor XIa. In contrast, factor
IXa production increased slowly to attain a stable steady-state level. We established
that surface-bound factor XIa was responsible for a continuous production of factor
IXa. Factor IXa-induced generation of factor Xa and thrombin was only observed when
contact activated plasma was subsequently perfused through a DOPS/DOPC-coated capillary,
showing that propagation of the factor IXa trigger requires a procoagulant, phosphatidylserine-containing,
phospholipid membrane. The negatively charged inner surface of a heparin-coated polyurethane
capillary, generated like the glass capillary significant amounts of factor XIa and
factor IXa when perfused with recalcified plasma. No differences were found between
unfractionated heparin and heparin devoid of anticoagulant activity. Thus, it is concluded
that contact activation and factor IXa generation in flowing plasma is not inhibited
by immobilised anticoagulant active heparin. Consequently, factor IXa-dependent thrombin
generation at a downstream located phospholipid membrane was similar, regardless the
specific anticoagulant activity of immobilised heparin.
