THIOCYANATE-BASED TOXICITY OF EOSINOPHIL PEROXIDASE

活化的嗜酸性粒细胞（EOS）可破坏寄生虫，但也可损害宿主 组织. 虽然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 是热带非洲和亚洲心脏病死亡的主要原因。 我们有 出乎意料地发现，饮食来源的硫氰酸拟卤化物 (SCN)，而不是如先前所认为的Br，是 生理体液中独特的嗜酸性粒细胞过氧化物酶（EPO）。 这是一 挑衅性的结果，因为这个反应的产物，HOSCN，是一个遥远的 比非特异性漂白氧化剂HOBr的效力弱的氧化剂 和HOCl，嗜中性粒细胞髓过氧化物酶的主要产物。 我们假设 正是由于其有限的氧化剂反应性，HOSCN， 与膜溶解性HOBr和HOCI不同， 氧化特定的细胞内蛋白巯基（SH）。 这个... 特异性可赋予HOSCN杀死寄生虫的能力 优先于宿主细胞。 然而，在血清SCN- 当EPO耗尽时，EPO反而会氧化Br以产生不分青红皂白的毒性， HOBr，其严重损害邻近的宿主组织。 因此，SCN-可能既 介导和调节EO细胞毒性。 我们的第一个目标是展示 生理上被大的调理颗粒激活的EOs产生 霍斯。 我们的第二个具体目标是确定分子机制 试剂HOSCN对EO本身的直接毒性， 哺乳动物细胞，以及寄生虫的经典模型 侵扰 我们假设HOSCN是一种SH特异性代谢产物， 毒素和糖酵解酶甘油醛-3-磷酸 脱氢酶（GAPDH）是其主要靶标。 第三个具体目标 是表征HOSCN毒性的可能间接机制 涉及与H2 O2其它阳离子的二次氧化相互作用 嗜酸性粒细胞颗粒蛋白、蛋白酶和抗蛋白酶。 有毒， HOSCN可用于宿主组织，然而，HOBr至少是宿主组织的十倍。 致命的 因此，我们的第四个具体目标是表征卤化物 嗜酸性粒细胞增多性心脏病患者的血清组成 疾病 我们假设这些病人过氧化消耗他们的 SCN-自身血清，低水平的SCN-是宿主组织的风险因素 通过允许EPO将Br氧化为HOBr而不是将SCN氧化为HOSCN来损害。 我们的第五个具体目标是建立小鼠嗜酸性粒细胞 心内膜炎利用过表达嗜酸性粒细胞的转基因小鼠， 特异性生长因子白细胞介素5（IL-5）。 我们将研究 SCN在调节心脏损伤中的作用 血清SCN耗竭增强。 这些研究如果成功， 为SCN确定了一个重要的、迄今为止未被怀疑的角色--在管理 EO对寄生虫和宿主组织的毒性。 如果是这样，我们将有 发现了一种简单的治疗策略来减弱宿主毒性-- SCN的膳食补充剂-这甚至可以应用于第三个 世界

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.