title: Prothrombin activation in blood coagulation: the erythrocyte contribution to thrombin generation.
abstract: 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 α-thrombin antithrombin (αTAT) complexes. We determined that Tf-activated blood produced both αTAT and mTAT. The presence of mTAT suggested that nonplatelet surfaces 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 αTAT formation. Only the addition of RBCs to PRP was capable of normalizing αTAT 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.
authors.affiliation: Department of Biochemistry, University of Vermont, Burlington, VT, USA.
authors.forename: Matthew F
authors.initials: MF
authors.lastname: Whelihan
authors.forename: Vicentios
authors.initials: V
authors.lastname: Zachary
authors.forename: Thomas
authors.initials: T
authors.lastname: Orfeo
authors.forename: Kenneth G
authors.initials: KG
authors.lastname: Mann
mesh_terms: D001777:Blood Coagulation / Q000502:physiology*; D002469:Cell Separation; D004792:Enzyme Precursors / Q000378:metabolism*; D004797:Enzyme-Linked Immunosorbent Assay; D004912:Erythrocytes / Q000378:metabolism*; D005434:Flow Cytometry; D006801:Humans; D011516:Prothrombin / Q000378:metabolism*; D015398:Signal Transduction / Q000502:physiology; D013917:Thrombin / Q000096:biosynthesis* / Q000378:metabolism
chemical_list: D004792:Enzyme Precursors; D011516:Prothrombin; D013917:Thrombin; C026812:meizothrombin