CHARACTERIZATION OF HEMATIN DERIVED ANTICOAGULANT

本建议书中描述的项目旨在定义 血红素衍生蛋白的生化、生物学和药理学意义 抗凝剂(HDA)。HDA在体外可延长凝血时间 在活体内。HDA是一种衍生品或降解产物 当血红素被允许站在溶液中或当它被注入到 老鼠。HDA将从体外和体内得到纯化和鉴定 来源通过层析和光谱技术。这将考验 假设HDA是血红素(血红素)的单一衍生物，这是 能够与凝血蛋白相互作用，产生深刻的 功能效应。HDA与特异体相互作用的机理 纤维蛋白原、凝血酶和因子VIII：C的定义如下 测定结合亲和力及对酶功能的影响 天然和合成底物。紫外可见光谱的具体技术 光谱学、平衡透析和铁结合拮抗 化合物将检验是否涉及铁或微量金属的假设 在HDA与凝血蛋白的相互作用中。制作和制作 HDA在体内的作用机制将通过分离的肝血红素来确定 代谢系统和整个动物。这将检验这一假设 HDA是血红素的代谢产物，其作用机制与 这是在体外观察到的。具体技术涉及放射性同位素和 药理探头。最后，HDA的药理潜力将是 由利用放射性标记进行的药代动力学和疗效研究确定 HDA和各种给药途径。完成这些研究后， 明确HDA的生化、生理和药理学意义。

The project described in this proposal is designed to define the biochemical, biologic and pharmacologic significance of hematin-derived anticoagulant (HDA). HDA produces prolongation of clotting times in vitro and in vivo. HDA is a derivative or degradation product which is produced when hematin is allowed to stand in solution or when it is infused into rats. HDA will be purified and identified from in vitro and in vivo sources by chromatographic and spectroscopic techniques. This will test the hypothesis that HDA is a single derivative of hematin (heme) which is capable of interacting with coagulation proteins to produce profound functional effects. The mechanism of interaction of HDA with the specific proteins fibrinogen, thrombin and factor VIII:C will be defined by determining binding affinities and effects on enzymatic function with natural and synthetic substrates. The specific techniques of UV-Vis spectroscopy, equilibrium dialysis and antagonism with iron binding compounds will test the hypothesis that iron or a trace metal is involved in the interaction of HDA with clotting proteins. The production and mechanism of HDA in vivo will be defined with isolated hepatic heme metabolic systems and whole animals. This will test the hypothesis that HDA is a metabolic product of heme and that the mechanism is identical to that seen in vitro. Specific techniques involve radioisotope and pharmacologic probes. Finally, the pharmacologic potential of HDA will be determined by pharmacokinetic and efficacy studies utilizing radiolabelled HDA and various routes of administration. Completion of these studies will define the biochemical, physiologic and pharmacologic significance of HDA.