PB0581 - In vivo activation of the contact pathway by thrombin in sterile models of DIC in baboons

Background: While data from in vitro and ex vivo systems support the conclusion that thrombin activates components of the contact system, particularly factor (F) XI, there is limited evidence for this process occurring in vivo.

Aims: We used baboon models of sterile disseminated intravascular coagulation (DIC) to test the hypothesis that thrombin promotes activation of contact phase proteases in vivo.

Methods: We used two models of thrombin-mediated DIC: (i) intravenous bolus of a mixture of 36.6 pmol/kg FXa and 56.3 nmol/kg phosphatidylcholine/phosphatidylserine (PC/PS), followed by blood collection at select intervals; (ii) continuous intravenous infusion of thrombin (27pmole/kg/min) for 60 min with periodic blood collection over 6 hours post-challenge. Plasma levels of antithrombin (AT) complexes with FXIIa, FXIa, FIXa, thrombi and kallikrein as well as total and cleaved kininogen were measured by sandwich ELISAs.

Results: FXa:PC/PS infusion induced a rapid burst of thrombin generation that peaked 10 min post-challenge, as shown by a rise in thrombin-AT (TAT) complexes. This was paralleled by rapid generation of AT complexes with FXIIa, FXIa, FIXa and kallikrein, all reaching maximums within 10-30 min post-challenge, followed by a decrease at 60 min. Thrombin infusion led to a gradual increase in AT complexes, reaching maximums at 60 min. Total AT levels transiently decreased by 10-20% after both challenges, then rapidly normalized. Changes in cleaved kininogen followed the pattern of kallikrein generation, with rapid increase after FXa-PC/PS bolus and steady increase during thrombin infusion. Unlike AT, there was a slight transient increase in total kininogen, suggesting rapid release from tissue stores post-thrombin generation.

Conclusion(s): Traditional models of coagulation are based on contact activation initiating thrombin production. Our data support the hypothesis that serine proteases including FXa and thrombin also contributes to activation of contact pathway proteases, indicating a more complex relationship between coagulation and contact activation.