Udovenko A.V. Regulation of Thrombin and Plasmin activity during clot formation and dissolution in human blood plasma. Qualifying scientific work submitted as a manuscript.
This PhD thesis is devoted to the investigation of the processes underlying the formation and dissolution of fibrin clots. To analyze the relationships between clot formation and dissolution and the activation of prothrombin and plasminogen, methods were developed for determining the activity of enzymes characterizing the coagulation component of thrombin activity and the fibrinolytic activity of plasmin at the moment of complete clot dissolution in human blood plasma.
During the study, it was found that the fibrillar structure of the clot is important for supporting prothrombin activation at the initiation stage of coagulation. Fibrinogen, upon exposure to fibrinogenase from the venom of Agkistrodon halys halys, loses more than an order of magnitude of its ability to support prothrombin activation.
It was established that thrombin accumulated within clots during the initiation stage is required for the amplification stage of thrombin activity and for switching the process of coagulation system activation to its inhibition. Excess thrombin released from the clot during the amplification stage of coagulation binds to FV, FVIII, and FXI, which causes a burst of amidolytic activity.
A novel method for calculating thrombin activity and concentration in the thrombin generation assay (TGA) was proposed, which allows the determination of thrombin concentration at every point of the thrombogram before and after the breakpoint of the curve.
Analysis of the relationship between the process of prothrombin activation and fibrin clot formation and the activation of the fibrinolytic system showed that the onset of fibrinolytic system activation occurs during the period of completion of the fibrillar clot structure formation. This suggests that the mechanism of interaction of fibrin desA polymerization sites functions in such a way that, during clot formation, components of the fibrinolytic system are competitively displaced onto the surface of the formed clot fibrils, where they create Glu-Pg–fibrin–Glu-Pm activation triads that initiate the mechanism of fibrinolysis.
Using a turbidimetric method for the analysis of fibril structure, it was shown that the density of protofibrils within a fibril can vary depending on the initiator of blood plasma coagulation. This indicates that the protofibril density parameter may potentially be used for diagnostic purposes.
Thus, during the course of this dissertation research, new relationships were identified between:
- prothrombin activation and clot formation over time at the initiation stage of blood plasma coagulation induced by an APTT reagent or tissue factor;
- the three-dimensional structure of the clot and the rate of prothrombin activation at the initiation stage of the clot formation process;
- the mechanism of fibril structure formation and the mechanism of formation of the Glu-Pg–fibrin–Glu-Pm activation triads of the fibrinolytic system, which is competitive in nature.
Keywords: fibrin polymerization, fibrinogen, thrombin, plasminogen activation, plasmin, turbidimetry, electron microscopy, inflammation, soluble fibrin, haemostasis, blood coagulation, fibrinolysis, antibodies, diagnostics.
