The fibrinolytic cascade functions to remove blood clots through the enzyme plasmin, which is generated as needed in a controlled fashion by a complex series of reactions. The proposed work is designed to provide a fuller, more quantitative understanding of the properties of the cascade and how its activity is regulated. The project is intimately related to the mission of the NIH because it is directed at obtaining a better understanding of the mechanisms which, when improperly regulated, underlie a good deal of the morbidity and mortality that afflict western society and manifest as heart attacks, stokes and other thrombotic pathologies.
SPECIFIC AIMS :
The specific aims of the project are:(1) To determine the dynamics of the interactions between fibrin, tPA and plasminogen .involved in plasminogen activation;(2) To investigate the enzymology of key reactions in the fibrinolytic cascade including: a) the fibrin dependent conversion of glu plasminogen to lys plasminogen by plasmin with and without TAFIa, b) the down regulation by fibrin of the inhibition of one and two chain tPA by PAI-1 with and without TAFIa, c) the activation of lys plasminogen by tPA on plasmin modified fibrin with and without TAFIa, and d) the plasmin catalyzed digestion of fibrin;(3) To determine the effects of TAFIa on components and reactions of the coagulation cascade;(4) To determine the effect of chronic TAFI activation on clot lysis;(5) To investigate the inhibition of fibrinolysis by platelets and study fibrinolysis in whole blood;(6) To determine the time courses and extents of TAFI activation in normal and factor VIII deficient whole blood and plasmas;and (7) To continue the development of the Lys Speed computer model of the fibrinolytic cascade. RESEARCH DESIGN AND METHODS:
The first aim will be accomplished by measuring the dynamics of the interactions by which the tPA-fibrin-plasminogen complex is assembled;the second will be accomplished by standard measurements of steady-state kinetics;the third, in collaboration with Dr. Mann, by exposing the clotting factors to TAFIa and assessing functional consequences;the forth by chronically generating TAFIa and monitoring clot lysis; the fifth, in collaboration with Drs. Tracy and Samis, by measuring lysis in the presence of platelets, whole blood, and other blood cells;the sixth, in collaboration with Dr. Brummel-Ziedins by measuring TAFIa activation and clot lysis in normal and factor VIIl-deficient whole blood or plasma;and the seventh by refining and validating the computer model of the fibrinolytic cascade.
The relevance of this work to public health is that it will lead to a better understanding of the means by which blood clots are removed and therefore to a better understanding of pathologies associated with improperly balanced fibrin deposition and removal such as venous thrombosis, pulmonary embolism, heart attack and stroke. In so doing, this work may contribute to better means to diagnose, treat and prevent these maladies.
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