SERPIN STRUCTURE AND THE DEVELOPMENT OF NEUTROPHIL RESISTANT SERPINS

颗粒、蛋白酶和氧化剂与激活的中性粒细胞有关 在病理过程中炎症反应对组织有贡献 脓毒症器官衰竭前的破坏和循环衰竭 还有ARDS。血管壁细胞外基质与止血途径 成分对中性粒细胞的破坏特别敏感。 弹性蛋白酶和组织蛋白酶G(CATG)及其降解相关 伴有低血压和弥散性血管内凝血(DIC)。在……里面 项目3我们将把我们的蛇形结构/功能研究扩展到两个方面 开发缓蚀剂的实际目标将有助于 败血症和ARDS。抗凝血酶III(ATIII)的研究进展 研究表明，可能存在重要的功能差异 自然产生的α-和β-ATIII亚型的相互作用 血管壁硫酸乙酰肝素蛋白多糖(HSPGs)。这将是 在目标1中通过测定异构体结合亲和力来研究 内皮细胞，并比较它们与FX激活的可及性 在内皮细胞(含有HSPG)上组装的复合体 血小板和磷脂小泡(不含)。目标2是让 血管壁导向、弹性蛋白酶和CATG耐药的凝血酶抑制物 和FXA。多项动物和人体研究表明，高剂量 静脉输注ATIII可逆转感染性DIC及相关低血压 和器官衰竭。我们假设大剂量的ATIII是 所需的原因是：(I)在 功能上重要的ATIII反应环，以及(Ii)耗尽 商品抗凝血酶的高肝素亲和力β-ATIII亚型 浓缩液。这些考虑表明，减少的 重组ATIII被设计成具有增强肝素亲和力和 弹性蛋白酶和CATG耐药可能对治疗有效 炎症性DIC。目标3是制造抗氧化性抗弹力酶 和与肝素结合的抗CATG蛇毒。肝素的投标性 应增加SEE抑制剂与弹性蛋白酶和 CAT G，并将其靶向阻断细胞外的血管壁 基质破坏与维持凝血的正常调节 通路组装很重要。最后，Airm 4将调查 高亲和力中性粒细胞耐药的治疗潜力 ATIII和血管壁导向的抗弹性蛋白酶和抗CATG蛇 脓毒症的内毒素血症大鼠模型。

Granule, proteinases and oxidants relesd from activatedneutrophils during pathological inflammatory reactions contribute to the tissue destruction and circulatory collapse preceding organ failure in sepsis and ARDS. Vessel wall extracellular matrix and hemostatic pathway components are particularly sensitive to destruction by neutrophil elastase and cathepsin G (catG), and their degradation is associated with hypotension and disseminated intravascular coagulation (DIC). In Project 3 we will extend our serpin structure/funciton research twoards the practical goal of developing inhibitors which would be useful in sepsis and ARDS. Recent developments in antithrombin III (ATIII) research suggest potentially important funcitonal differences in the interactions of the naturally occurring alpha- and beta-ATIII isoforms with vessel wall heparan sulfate proteoglycans (HSPGs). This will be investigated in Aim 1 by determining isoform binding affinities for endothelial cells, and comparing their accessibility to fX-activating complex assembled on endothelial cells (which contain HSPGs) and on platelets and phospholipid vesicles (which do not). Aim 2 is to make vessel wall directed, elastase- and catG-resistant inhibitors of thrombin and fXa. Several animal and human studies have shown that high dose infusion of ATIII can reverse septic DIC and associated hypotension and organ failure. We hypothesize that high doses of ATIII are required due to (I) the preseence of an elastase cleavage site in the funcitonally important reactive loop of ATIII, and (ii) depletion of the high-heparin-affinity beta-ATIII isoform from commercial antithrombin concentrates. These considerations suggest that reduced amounts of a recombinant ATIII engineered to have enhanced heparin affinity and elastase- and catG- resistance may be effective for treating inflammatory DIC. Aim 3 is to make oxidation-resistant antielastase and anti-catG serpins that bind heparin. The heparin bidning property should increase association rates ofthesee inhibitors with elastase and cat G, and target them to the vessel wall where blocking extracellular matrix destruction and maintaining normal regulation of coagulation pathway assemblies is important. Finally, Airm 4 is to investigate the therapeutic potenital of the high-heparin-affinity neutrophil-resistant ATIII and the vessel wall directed anti-elastase and anti-catG serpins in an endotoxemic rat model of sepsis.