Eosinophil Peroxidase in Allergic Inflammation

描述（由申请方提供）：嗜酸性粒细胞（EO）吞噬细胞可损伤宿主组织，并促进过敏性炎症性疾病（如哮喘和炎症性肠病（IBD））的发病机制。EO特异性颗粒具有丰富的EO过氧化物酶（EPO）和强烈的呼吸爆发，产生H2 O2以促进其他氧化剂的产生。然而，关于EPO介导的氧化损伤对嗜酸性粒细胞炎症状态的病理学的贡献知之甚少。我们发现，三个不寻常的基板-溴（Br-），亚硝酸盐（NO2-），硫氰酸盐（SCN-）-竞争氧化EPO在生理液体中的存在下，过氧化氢，产量，分别，HOBr，NO2，和HOSCN。这些氧化剂对人类细胞的相对毒性为HOBr > NO2 >> HOSCN;而EPO优先氧化SCN- > NO2- > Br-。我们假设SCN-“缓冲剂”对抗EPO产生更具细胞毒性的基于NO2-和Br-的氧化剂，因此血清SCN-水平（由饮食决定）可能调节EPO毒性。我们所提出的工作的总体目标是检验EPO产生的氧化剂以底物决定的方式对宿主组织和EO本身造成损伤的假设。我们的第一个具体目标是在小鼠硫酸葡聚糖模型中验证通过饮食补充硫氰酸盐或其代谢前体氰化物样化合物来调节EPO毒性的底物可改善炎症性肠病的严重程度的假设。我们的第二个具体目的是检验这样的假设，即在一种新的鼠IL- 5/Eotaxin-2（I5/E2）转基因哮喘模型中，1）与EPO-/-小鼠杂交，2）EPO酶活性的药理学抑制，和3）膳食SCN-补充剂都改善了这些动物自发和卵清蛋白（OVA）敏化和激发后发展的严重哮喘表型。第三个具体目的是检验EPO/H2 O2/SCN-系统通过NF-β依赖性机制抑制嗜酸性粒细胞凋亡和有害的继发性坏死脱粒的假设。这些研究如果成功，将证明EPO介导的氧化损伤在哮喘和IBD发病机制中的作用，并提出一种简单的治疗策略-即，膳食补充剂与廉价和无害的SCN/-，可以适用于这些和其他过敏性炎症疾病，即使在贫困社区。 公共卫生相关性：在这项资助中，我们将测试硫氰酸盐，一种在蔬菜中发现的简单，廉价和无毒的化合物，是否可以被摄入治疗哮喘和溃疡性结肠炎，这两种严重的过敏性疾病。我们相信这是可能的，因为硫氰酸盐可以钝化由嗜酸性粒细胞引起的组织损伤，嗜酸性粒细胞是一种白色细胞，会引起过敏症状和疾病。如果这些研究成功，我们将发现一种简单，廉价和安全的治疗过敏的方法。

DESCRIPTION (provided by applicant): Eosinophil (EO) phagocytes can damage host tissue and contribute to the pathogenesis of allergic inflammatory diseases such as asthma and inflammatory bowel diseases (IBD). EO specific granules are endowed with abundant amounts of EO peroxidase (EPO) and a vigorous respiratory burst that generates H2O2 to fuel the generation of other oxidants. However, little is known about the contribution of EPO-mediated oxidative damage to the pathology of eosinophilic inflammatory states. We find that three unusual substrates -- bromide (Br-), nitrite (NO2-), and thiocyanate (SCN-) -- compete for oxidation by EPO in physiologic fluids in the presence of H2O2, yielding, respectively, HOBr, NO2, and HOSCN. The relative toxicity of these oxidants for human cells is HOBr > NO2 >> HOSCN; yet EPO preferentially oxidizes SCN- > NO2- > Br-. We hypothesize that SCN- "buffers" against generation by EPO of the more cytotoxic NO2-- and Br-- based oxidants and consequently serum SCN- levels, which are dietarily determined, may modulate EPO toxicity. The overall goal of our proposed work is to examine the hypothesis that that EPO-generated oxidants impose damage in a substrate-determined manner to host tissue and EOs themselves. Our first specific aim s to test the hypothesis that substrate modulation of EPO toxicity by dietary supplementation with either thiocyanate or its metabolic precursors, cyanide-like compounds, ameliorates the severity of inflammatory bowel disease in a murine dextran sulfate model. Our second specific aim is to test the hypothesis that in a novel murine IL- 5/Eotaxin-2 (I5/E2) transgenic model of asthma, 1) crossbreeding with EPO-/- mice, 2) pharmacologic inhibition of EPO enzymatic activity, and 3) dietary SCN- supplementation all ameliorate the severe asthmatic phenotype which develops in these animals both spontaneously and after ovalbumin (OVA) sentitization and challenge. The third specific aim is to test the hypothesis that the EPO/H2O2/SCN- system functions through an NF-(B-dependent mechanism to inhibit eosinophil apoptosis and deleterious secondary necrotic degranulation. These studies, if successful, will prove a role for EPO-mediated oxidant damage in the pathogenesis of asthma and IBD and suggest a simple strategy for its treatment-- i.e., dietary supplementation with inexpensive and innocuous SCN/-- that could be applied to these and other allergic inflammatory diseases even in impoverished communities. PUBLIC HEALTH RELEVANCE: In this grant we will test whether thiocyanate, a simple, inexpensive and non-toxic chemical compound found in vegetables, can be ingested to treat asthma and ulcerative colitis, two serious allergic diseases. We believe this is possible because thiocyanate blunts tissue damage caused by eosinophils, a type of white cell that causes allergic symptoms and diseases. If these studies succeed, we will have discovered a simple, cheap and safe way to treat allergies.