Asthma susceptibility due to environmental programing of innate immunity in utero

描述（由申请人提供）：产前环境暴露越来越被认为是哮喘的重要风险因素。关键暴露包括城市空气污染及其成分-柴油废气。这些机制的特征不明确。该项目旨在弥补这一知识差距。为了建立机制研究的框架，我们开发了小鼠模型。在该模型中，妊娠小鼠反复暴露于柴油机排气颗粒（DEP）使其后代对出生后过敏原-卵清蛋白（OVA）过敏。这些后代在受到次优的OVA致敏和激发时发生哮喘。相比之下，未暴露母鼠所生的幼崽在次优的OVA暴露后不会发生哮喘。我们的初步数据表明，该模型中的哮喘是由产生IL-5，IL-13和IL-17的NK细胞介导的。使用特异性抗体消除这些细胞可以预防哮喘。我们还表明，IL-5/IL-13/IL-17+ NK细胞在人类哮喘中增加。除了IL-5、IL-13和IL-17的产生之外，致病性小鼠NK细胞的特征在于芳香烃受体（AhR）特征转录物（AhRR、Cyp 1a 1和Cyp 1b 1）的表达增加。AhR是已知响应DEP的有机化合物，即多环芳烃（PAH）的转录因子。AhR还已知调节IL-17基因转录。一些报道表明AhR在IL-5和IL-13产生中的积极作用。因此，AhR提供了母体暴露于DEP与后代NK细胞中细胞因子产生之间的分子联系。基于这些数据，我们的总体假设是，在子宫内暴露于柴油机废气促进哮喘易感性通过AhR依赖性的NK细胞启动。我们提出了以下具体目标：1）检测NK细胞在产前暴露于DEP诱导的哮喘易感性中的重要性; 2）研究芳烃受体在NK细胞活化和哮喘风险的母胎传递中的作用; 3）检测致病性DEP效应在第一代和后代中的持续性; 4）检测IL- 5/IL-13/IL-17+ NK细胞和AhR/AhRR/Cyp 1b 1 + NK细胞在人哮喘中的相关性。为了实现这些目标，我们将使用NK细胞缺陷型小鼠、无适应性免疫的小鼠和具有NK细胞特异性AhR缺失的小鼠。我们还将从产前暴露的后代向未暴露的小鼠转移NK细胞。为了研究在整个后代生活中保持哮喘易感性，我们将使用交叉培养和延迟过敏原暴露的方法。为了研究DEP暴露的跨代效应，我们将检查F2和F3后代的哮喘易感性。为了确定人类相关性，我们将测量患有哮喘的幼儿和成人中的IL-5/IL-13/IL-17+ NK细胞和AhR/AhRR/Cyp 1b 1 + NK细胞。我们将NK细胞结果与PAH-DNA加合物的量（环境暴露的测量）相关联。使用体外培养系统，我们将确定DEP是否可以将来自非过敏性非哮喘供体的非极化NK细胞偏斜为产生IL-5/IL- 13/IL-17的细胞。然后，我们将研究AhR在这一过程中的作用，使用击倒的方法。

DESCRIPTION (provided by applicant): Prenatal environmental exposures are increasingly recognized as important risk factors for asthma. Critical exposures include urban air pollution and its component - diesel exhaust. The mechanisms are poorly characterized. This project seeks to address this gap in knowledge. To build a framework for mechanistic studies we have developed a mouse model. In this model, repeated exposure of pregnant mice to diesel exhaust particles (DEP) renders their offspring hypersensitive to the postnatal allergen - ovalbumin (OVA). These offspring develop asthma when subjected to suboptimal OVA sensitization and challenge. In contrast, pups born to unexposed dams do not develop asthma upon suboptimal OVA exposure. Our preliminary data indicate that asthma in this model is mediated by NK cells that produce IL-5, IL-13 and IL-17. Depletion of these cells using specific antibodies prevents asthma. We also show that IL-5/IL-13/IL-17+ NK cells are increased in human asthma. In addition to IL-5, IL-13 and IL-17 production, the pathogenic mouse NK cells are characterized by increased expression of the aryl hydrocarbon receptor (AhR) signature transcripts (AhRR, Cyp1a1 and Cyp1b1). AhR is a transcriptional factor that is known to respond to organic compounds of DEP, namely, polycyclic aromatic hydrocarbons (PAH). AhR is also known to regulate the IL-17 gene transcription. Some reports suggest the positive role of AhR in IL-5 and IL-13 production. Thus, AhR provides a molecular link between maternal exposure to DEP and cytokine production in offspring NK cells. Based on these data, our overarching hypothesis is that in utero exposure to diesel exhaust promotes asthma susceptibility through AhR-dependent priming of NK cells. We propose following specific aims: 1) Examine the importance of NK cells in asthma susceptibility induced via prenatal exposure to DEP; 2) Study the role of the aryl hydrocarbon receptor in NK cell activation and maternal-fetal transmission of asthma risk; 3) Examine the persistence of the pathogenic DEP effect through the life of first and subsequent generations; 4) Examine the relevance of IL- 5/IL-13/IL-17+ NK cells and AhR/AhRR/Cyp1b1+ NK cells in human asthma. To meet these goals we will use NK cell-deficient mice, mice without adaptive immunity and mice with NK cell-specific deletion of AhR. We will also perform transfers of NK cells from prenatally exposed offspring to unexposed mice. To study preservation of asthma susceptibility throughout the offspring life we will use cross-fostering and delayed allergen exposure approaches. To study transgenerational effects of DEP exposure, we will examine asthma susceptibility in F2 and F3 offspring. To determine human relevance we will measure IL-5/IL-13/IL-17+ NK cells and AhR/AhRR/Cyp1b1+ NK cells in young children and adults with asthma. We will correlate NK cell results with amounts of PAH-DNA adducts (a measure of environmental exposure). Using an in vitro culture system we will determine whether DEP can skew non-polarized NK cells from non-allergic non-asthmatic donors into IL-5/IL- 13/IL-17-producing cells. We will then study the role of AhR in this process using a knockdown approach.