TLR Ligand and Pesticide Exposure in Farms: Role in Atopic Immune Responses

描述（由申请人提供）：最近由NIEHS， NIOSH和NCI的研究人员对25,814名农场妇女进行的农业健康研究报告了282名受试者在农场使用农药与特应性哮喘之间的高度显著关联。在农场长大的女性中，这种联系最为强烈。与农场环境的保护作用相反，在农场长大和使用杀虫剂与特应性哮喘的总体风险较高共同相关（基于卫生假说的预期）；同样的研究也证明了在农场长大的女性从未使用过杀虫剂。最近的另一项研究表明，农药处理小鼠内毒素诱导的几种细胞因子的产生受到抑制。农场环境中暴露于toll样受体（TLR）配体，包括来自微生物和微生物产物的MAMPs（微生物相关分子模式），对过敏反应的抑制有重要影响。最近卫生学假说的修正表明，这种抑制反应与调节性T细胞（Treg）的诱导有关。因此，我们建议探索和确定TLR配体和农药联合气道暴露如何影响农场环境中与特应性哮喘相关的免疫反应。在第一个具体目标中，我们将描述俄亥俄州10个农场在使用农药前后环境TLR配体（内毒素、肽聚糖和细菌DNA）和两种与特应性哮喘相关的传统农药的暴露情况。将使用最先进的生物气溶胶取样方法。它包括一个可吸入采样器和niosh开发的尺寸选择采样系统，该系统具有两级旋风（三个粒径部分：<1.0 m, 1 - 1.8 m和>1.8 m），用于收集空气中的TLR配体和农药，以及用于收集土壤表面可雾化TLR配体和农药的新型微生物源强度测试仪。我们将使用LUDEP ICRP呼吸沉积模型来确定男性、女性和儿童受试者呼吸沉积TLR配体和农药的潜力。因此，第一个具体目标的结果将使我们能够了解现实世界中农场中可吸入TLR配体和农药的同时暴露水平。在第二个具体目标中，我们将确定来自上述农场的未纯化但临床相关的空气和表面样品提取物如何影响宿主对卵清蛋白诱导的超敏反应和诱导Tregs的敏感性。这些影响的TLR4依赖性将通过卵清蛋白致敏野生型和TLR4 -/- C57BL/6小鼠来确定。这些发现将为农药暴露的不良影响与TLR配体暴露在农业工人特应性哮喘发展中的保护作用之间的关系提供重要的初步信息。

DESCRIPTION (provided by applicant): The recent Agricultural Health Study among 25,814 farm women conducted by the researchers from NIEHS, NIOSH, and NCI reported a highly significant association between any use of pesticides in farms and atopic asthma in 282 subjects. This association was strongest among women who grew up on a farm. Growing up on farms and application of pesticides were jointly associated with higher overall risk for atopic asthma in contrary to the protective effect of farm environment (as expected based on the Hygiene Hypothesis); which was in the same study demonstrated for women who grew up on farms and never applied pesticides. Another recent study showed that the production of several cytokines induced by endotoxin was suppressed in pesticide treated mice. Exposures to toll like receptor (TLR) ligands in farm environment including MAMPs (microbe-associated molecular patterns) from microorganisms and microbial products have significant influence on the suppression of allergic response. This suppressive response was implicated to the induction of regulatory T cells (Treg) by recent revisions of the Hygiene Hypothesis. Therefore we propose to explore and determine how combined airway exposures to TLR ligands and pesticides influence immune responses associated with atopic asthma in farm environments. In the first specific aim we will characterize exposures of environmental TLR ligands (endotoxin, peptidoglycan, and bacterial DNA) and two conventional pesticides relevant to atopic asthma in 10 farms of Ohio before and after pesticide application. A state-of-the-art bioaerosol sampling methodology will be used. It includes an inhalable sampler and a NIOSH-developed size- selective sampling system with two-stage cyclone (three particle size fractions: <1.0 5m, 1 - 1.8 5m, and >1.8 5m), for the collection of airborne TLR ligands and pesticides and a novel microbial source strength tester for the collection of aerosolizable TLR ligands and pesticides from soil surface. We will determine the respiratory deposition potential of particulate TLR ligands and pesticides for male, female, and children subjects using the LUDEP ICRP respiratory deposition model. Results from the first specific aim will thus allow us to understand the real world simultaneous exposure levels to inhalable TLR ligands and pesticides in farms. In second specific aim we will determine how unpurified but clinically relevant air and surface sample extracts from above-mentioned farms analogous to real world exposures influence host susceptibility towards ovalbumin induced hypersensitivity and induction of Tregs. The TLR4 dependence of these influences will be determined by using ovalbumin-sensitized wild type and TLR4 -/- C57BL/6 mice. The findings will provide vital preliminary information on the relationship between adverse effects of pesticide exposure versus protective effects of TLR ligand exposure in the development of atopic asthma among agricultural workers. PUBLIC HEALTH RELEVANCE: Pesticide usage can be significantly associated with atopic asthma among the farmers and therefore farming environment may not provide the protective effect predicted based on the traditional Hygiene Hypothesis. Wide range of immunostimulatory materials and pesticides are present in farming environments and traditional investigations with purified materials and pesticides may not provide precise information on relationship between adverse effect of pesticide exposures versus protective effect of TLR ligands in developing atopic asthma among farmers. In this R21 project we will explore immunological activities of unpurified but clinically relevant environmental samples collected in farms (before and after pesticide application) in ovalbumin allergen sensitized mice.