Summary
Three small animal models of bleeding are described and used to evaluate the effects
of preparations intended for therapy of human bleeding disorders. We modified techniques
for the assessment of bleeding to be able to reproducibly quantify blood loss and
rate of blood flow in addition to the measurement of bleeding time.
Temporary hemophilia was induced in a rabbit model by injection of high titer human
inhibitor plasma [>1000 Bethesda units (BU)/ml]. A decrease in rabbit FVIII from normal
values to below the limit of detection was observed within 30 min, cuticle bleeding
time changed from normal (approx. 10 min) to steady state bleeding (>30 min), and
the rate of blood flow increased from 4 to >30 µl blood/min. Infusion of an activated
prothrombin complex concentrate (FEIBA STIM4, Immuno) at doses between 75 and 150
U/kg normalized the rate of blood flow, while infusion of FVIII/vWF concentrate resulted
in partial correction. Administration of FVIIa, both recombinant and plasma-derived,
failed to correct bleeding, however. In an analogous murine model, FVIII/ vWF inhibitor
plasma was obtained by immunizing goats with a purified human FVIII/vWF complex. This
plasma cross-reacted with mouse vWF in vitro. Injection of the anti-FVIII/vWF inhibitor
plasma into mice caused a decrease in vWF antigen, in some animals with a complete
loss of vWF multimers comparable to severe von Willebrand disease. A specific anti-vWF
inhibitor plasma obtained by immunization of goats with recombinant vWF was used in
a further murine model, resulting in a gradual but substantial decrease in FVIII as
well as in intensive bleeding. The infusion of a FVIII/vWF concentrate (IMMUNATE,
IMMUNO) normalized the rate of blood flow in both murine models. The same assessment
methods were used to characterize bleeding in a natural mouse model of von Willebrand
disease (strain RIIIS/J). The use of quantitative techniques of assessment of blood
loss and rate of blood flow appears to be a helpful tool for characterizing hemorrhagic
situations and evaluating the capacity of therapeutic preparations to correct hemostatic
defects.
