THIOCYANATE-BASED TOXICITY OF EOSINOPHIL PEROXIDASE

嗜酸性粒细胞过氧化物酶的硫氰酸盐毒性

• 批准号: 2225015
• 负责人: ARNE SLUNGAARD
• 金额: $ 19.69万
• 依托单位: UNIVERSITY OF MINNESOTA
• 依托单位国家: 美国
• 财政年份: 1994
• 资助国家: 美国
• 起止时间: 1994-02-01 至 1998-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2225015
• 关键词: SDS polyacrylamide gel electrophoresis; Schistosoma mansoni; cellular pathology; collagenase; cytotoxicity; endocarditis; enzyme activity; eosinophil; exocytosis; genetically modified animals; glyceraldehyde 3 phosphate dehydrogenase; halogens; human subject; hydrogen peroxide; laboratory mouse; model design /development; oxidizing agents; peroxidases; phagocytosis; photomicrography; protease inhibitor; thiocyanates; thiols; tissue /cell culture; toxicant interaction

Activated eosinophils (EOS) destroy parasites but can also damage host tissue. Although the most common human disease caused by EOS is allergies, the most catastrophic is eosinophilic endocarditis, the second leading cause of cardiac deaths in tropical Africa and Asia. We have unexpectedly found that the dietarily derived pseudohalide thiocyanate (SCN), not Br as previously thought, is the primary substrate for the unique eosinophil peroxidase (EPO) in physiologic fluids. This is a provocative results because the product of this reaction, HOSCN, is a far weaker oxidant than are the potent by nonspecific bleaching oxidants HOBr and HOCl, the main product of neutrophil myeloperoxidase. We hypothesize that precisely because of its restricted oxidant reactivity, HOSCN, unlike membrane lytic HOBr and HOCI, penetrates intact membranes to oxidize specific intracellular protein sulfhydryl (SHs). This SH- specificity may endow HOSCN with the capacity to kill parasites preferentially over host cells. However, in conditions of serum SCN- depletion, EPO will instead oxidize Br to produce indiscriminantly toxic HOBr, which severely damages adjacent host tissues. Thus, SCN-may both mediated and modulate EO cytotoxicity. Our first specific aim is to show that EOs physiologically activated by large opsonized particles produce HOSCN. Our second specific aim is to define molecular mechanisms underlying the direct toxicity of reagent HOSCN for EOs themselves, for mammalian cells, and for schistosomules, a classical model of parasite infestation. We hypothesize that HOSCN is a SH-specific metabolic poison, and the glycolytic enzyme glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is its principal target. Our third specific aim is to characterize possible indirect mechanisms of HOSCN toxicity involving secondary oxidative interactions with H2O2 other cationic eosinophil granule proteins, proteases, and anti-proteases. Toxic as HOSCN may be for host tissue, however, HOBr is at least tenfold more lethal. Therefore our fourth specific aim is to characterize the halide composition of serum from individuals with hypereosinophilia heart disease. We hypothesize that such patients peroxidatively deplete their own serum of SCN-, low levels of which are a risk factor for host tissue damage by permitting EPO to oxidize Br to HOBr instead of SCN to HOSCN. Our fifth specific aim is to establish a mouse model of eosinophilic endocarditis utilizing transgenic mice that overexpress the eosinophil- specific growth factor interleukin 5 (IL-5). We will examine the role of SCN in regulating cardiac damage in this model by experimentally augmenting of depleting serum SCN. These studies, if successful, will define an important, hitherto unsuspected role for SCN- in governing the toxicity of EOs for parasites and host tissues. If so, we will have identified a simple therapeutic strategy for blunting host toxicity -- dietary supplementation of SCN -- that could be applied even in a Third World.

激活的嗜酸性粒细胞（EOS）破坏寄生虫，但也可能损坏宿主 组织。 尽管由EOS引起的最常见的人类疾病是 过敏，最灾难性的是嗜酸性心内膜炎，第二个 热带非洲和亚洲心脏死亡的主要原因。 我们有 出乎意料地发现，饮食中衍生的硫烷硫氰酸酯 （SCN），而不是以前认为的BR，是 生理液中独特的嗜酸性粒细胞过氧化物酶（EPO）。 这是一个 挑衅性的结果是因为该反应的产物HoSCN是一个很远的 氧化剂比非特异性漂白氧化剂HOBR弱氧化剂 和HOCL，嗜中性粒细胞髓过氧化物酶的主要产物。 我们假设 正是由于其受限的氧化反应性hoscn， 与膜裂解HOBR和HOCI不同，将完整的膜穿透到 氧化特异性细胞内蛋白硫酰基（SHS）。 这个sh- 特异性可能会赋予杀死寄生虫的能力 优先于宿主细胞。 但是，在血清scn-的条件下 耗尽，EPO将氧化BR氧化以产生不加选择的毒性 HOBR，严重损害相邻的宿主组织。 因此，scn-may两者兼而有之 介导和调节EO细胞毒性。 我们的第一个具体目的是显示 由大型重质颗粒在生理上激活的EOS会产生 霍斯特。 我们的第二个特定目的是定义分子机制 基础试剂HOSCN对EOS本身的直接毒性，因为 哺乳动物细胞，对于Scistosomules，是寄生虫的经典模型 侵扰。 我们假设HOSCN是SH特异性的代谢 毒药和糖酵解酶甘油醛-3-磷酸盐 脱氢酶（GAPDH）是其主要目标。 我们的第三个特定目标 是为了表征HOSCN毒性可能的间接机制 涉及与H2O2其他阳离子的继发氧化相互作用 嗜酸性粒细胞颗粒蛋白，蛋白酶和抗蛋白酶。 有毒为 HOSCN可能是用于宿主组织的，但是，HOBR至少要多十倍 致命。 因此，我们的第四个具体目的是表征卤化物 血清的组成来自血清粒细胞性心脏的患者 疾病。 我们假设此类患者过氧化耗尽了 SCN-的血清，低水平是宿主组织的危险因素 通过允许EPO将BR氧化为HOBR而不是SCN来损坏HOSCN。 我们的第五个具体目的是建立嗜酸性粒细胞的小鼠模型 使用过表达嗜酸性粒细胞的转基因小鼠的心内膜炎 特定生长因子白介素5（IL-5）。 我们将检查角色 通过实验来调节此模型中心脏损伤的SCN 增加耗尽的血清SCN。 这些研究，如果成功的话， 定义了迄今为止在scn的重要角色 EOS对寄生虫和宿主组织的毒性。 如果是这样，我们将有 确定了一种简单的治疗策略，用于钝化宿主的毒性 - 饮食补充SCN-即使在第三次中也可以应用 世界。