Prothrombin activation in blood coagulation: the erythrocyte contribution to thrombin generation

Prothrombin activation can proceed through the intermediates meizothrombin or prethrombin-2. To assess the contributions that these 2 intermediates make to prothrombin activation in tissue factor (Tf)-activated blood, immunoassays were developed that measure the meizothrombin antithrombin (mTAT) and alpha-thrombin antithrombin (alpha TAT) complexes. We determined that Tf-activated blood produced both alpha TAT and mTAT. The presence of mTAT suggested that nonplatelet sur-faces were contributing to approximately 35% of prothrombin activation. Corn trypsin inhibitor-treated blood was fractionated to yield red blood cells (RBCs), platelet-rich plasma (PRP), platelet-poor plasma (PPP), and buffy coat. Compared with blood, PRP reconstituted with PPP to a physiologic platelet concentration showed a 2-fold prolongation in the initiation phase and a marked decrease in the rate and extent of alpha TAT formation. Only the addition of RBCs to PRP was capable of normalizing alpha TAT generation. FACS on glycophorin A-positive cells showed that approximately 0.6% of the RBC population expresses phosphatidylserine and binds prothrombinase (FITC Xa.factor Va). These data indicate that RBCs participate in thrombin generation in Tf-activated blood, producing a membrane that supports prothrombin activation through the meizothrombin pathway. (Blood. 2012;120(18):3837-3845)