Eosinophil Peroxidase in Allergic Inflammation

描述(申请人提供)：嗜酸性粒细胞(EO)吞噬细胞可损害宿主组织，并有助于过敏性炎症性疾病的发病机制，如哮喘和炎症性肠病(IBD)。EO特定颗粒具有丰富的EO过氧化物酶(EPO)和旺盛的呼吸爆发，产生过氧化氢以促进其他氧化剂的产生。然而，关于EPO介导的氧化损伤在嗜酸性炎症状态病理中的作用还知之甚少。我们发现，在过氧化氢存在的生理液体中，三种不寻常的底物--溴(Br2-)、亚硝酸盐(NO2-)和硫氰酸盐(SCN-)--竞争EPO的氧化，分别产生HOBr2、NO2和HOSCN。这些氧化剂对人体细胞的相对毒性为HOBR&GT；NO2&GT；&GT；HOSCN；而EPO优先氧化SCN-&GT；NO2-&GT；BR-。我们假设，SCN-“缓冲”了EPO产生的更具细胞毒性的NO2-和BR-基氧化剂，因此血清SCN-水平由饮食决定，可能调节EPO的毒性。我们拟议工作的总体目标是检验这一假设，即EPO产生的氧化剂以底物决定的方式对宿主组织和Eos本身造成损害。我们的第一个特定目标是在小鼠硫酸葡聚糖模型中测试通过饮食补充硫氰酸盐或其代谢前体氰化物类氰化物来调节EPO毒性的底物可以改善炎症性肠病的严重程度的假说。我们的第二个特定目的是验证这样的假设：在一种新的小鼠IL-5/eoaxin-2(i5/e2)转基因哮喘模型中，1)与EPO-/-小鼠杂交，2)药物抑制EPO酶活性，3)饮食补充SCN都能改善这些动物自发和卵清蛋白(OVA)敏感和激发后出现的严重哮喘表型。第三个具体目的是验证EPO/H_2O_2/SCN-系统通过依赖于核因子(B)的机制来抑制嗜酸性粒细胞凋亡和有害的继发性坏死性脱颗粒的假设。这些研究如果成功，将证明EPO介导的氧化剂损伤在哮喘和IBD的发病机制中发挥了作用，并提出了一种简单的治疗策略--即通过廉价和无害的SCN/饮食补充--即使在贫困社区也可以应用于这些和其他过敏性炎症性疾病。 与公众健康相关：在这笔赠款中，我们将测试硫氰酸盐是否可以用于治疗哮喘和溃疡性结肠炎这两种严重的过敏性疾病。硫氰酸盐是一种在蔬菜中发现的简单、廉价和无毒的化合物。我们相信这是可能的，因为硫氰酸盐可以钝化嗜酸性粒细胞造成的组织损害，嗜酸性粒细胞是一种会导致过敏症状和疾病的白细胞。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.