ACTIVATION OF PROTHROMBIN

这个研究项目旨在了解凝血酶是如何产生的以及凝血酶是如何被调节的。我们解决这些问题的方法来自于四个独立方向的融合：1)凝血酶复合物的物理性质，它们的成分以及这些复合物如何组装成有效的酶催化剂。2)混合多种凝血催化剂/抑制剂，试图复制与凝血酶表达的组织因子途径相关的联合催化剂体系的性能的研究。3)在最低限度修饰的生物系统（全血）中研究这一过程，以评估从纯化系统中得出的假设的正确性。4)建立数学模型，在定量的基础上定义凝血过程及其调控，以帮助实验设计1、2、3，也有助于药物制剂的评估以及止血和血栓性疾病的诊断和治疗。本研究的目的是了解促凝剂和抗凝剂维生素k依赖性复合物的性质及其在凝血酶生成过程中的调节作用。研究将采用物理化学技术，包括流体动力学和荧光光谱（后者在溶液和表面），在封闭系统和流动下研究合成膜和细胞上的复合物。反应将使用合成和天然底物来监测前稳态和稳态动力学事件。天然和重组抑制剂将用于研究与凝血酶生成相关的促凝和抗凝过程的调节。我们将把通过单个反应研究获得的详细信息与从多反应中心系统获得的信息结合起来。相反，在全血系统中发生的过程将在纯化系统分析中引起适当的注意。我们期望在凝血事件中同时发生的复杂反应的生物学相关化学的定量评估。这些数据在解释正常生理和发展纠正与血栓和血友病相关的凝血病理的方法方面具有重要意义。我们开发的技术将为评估止血和血栓性疾病的潜在药物干预提供工具。

This research program is aimed at understanding how thrombin is generated and how thrombin generation is regulated. Our approach to these questions comes via the convergence of four separate directions associated with 1) the physical properties of coagulation enzyme complexes, their constituents and how these complexes can assemble into efficient enzyme catalysts. 2) Studies in which multiple coagulation catalysts/inhibitors are mixed to attempt to duplicate the performance of the combined catalyst system associated with the tissue factor pathway of thrombin expression. 3) To study this process in minimally modified biological systems (whole blood) to evaluate the correctness of hypothesis derived from purified systems. 4) To create mathematical models which can be used to define, on a quantitative basis, the process of blood clotting and its regulation both to aid in experimental designs 1,2,3, and also to aid in the evaluation of the pharmacologic agents and the diagnosis and treatment of hemostatic and thrombotic diseases. The aim of the present investigation is to understand the nature of procoagulant and anticoagulant vitamin-K dependent complexes and their regulation during the process of thrombin generation. Studies will employ physical chemistry techniques including hydrodynamics and fluorescence spectroscopy, (the latter both in solution and on surfaces) in closed systems and under flow to study complexes on synthetic membranes and cells. Reactions will be followed using both synthetic and natural substrates to monitor both presteady state and steady state kinetic events. Natural and recombinant inhibitors will be used to study the regulation of procoagulant and anticoagulant processes associated with thrombin generation. We will integrate the detailed information available through studies of individual reactions with that obtained from multi-reaction center systems. Conversely, the processes noted to occur in the whole blood system will direct appropriate attention in the purified system analyses. We anticipate developing a quantitative evaluation of the biologically relevant chemistry associated with the complex reactions which occur simultaneously during a blood clotting event. These data have significance in interpreting normal physiology and in developing approaches to correct the coagulation pathology associated with thrombosis and hemophilia. The techniques we develop will provide tools for the evaluation of potential pharmacological intervention in hemostatic and thrombotic disease.