Protective role of intestinal microbiota in food allergy

描述(申请人提供)：在美国和其他发达国家，食物过敏的流行率正在以惊人的速度上升。改变有益共生细菌种群的环境刺激与这一增长有关。我们实验室早期的工作表明，无法通过TLR4发出信号的小鼠对食物的过敏反应增强。我们假设共生细菌是TLR4配体的来源，并证明了新生儿服用广谱抗生素(ABX)的鸡尾酒在TLR4充足的小鼠中诱导了与TLR4突变小鼠相同的过敏反应。在这份修订后的申请中提供的初步数据中，我们建立了一个新的食物过敏的灵知生物模型，并表明直接来自健康小鼠肠道微生物区系的确定的细菌联合体可以防止系统对食物过敏原的高反应性。我们证明，梭状芽胞杆菌类细菌选择性地诱导屏障保护反应，包括激活IL-23/IL-22轴，诱导抗菌肽Reg3b和Reg3g的表达，以及扩张肠道Tregs和IgA分泌B细胞；这种反应的一部分是TLR4依赖的。我们假设，含有梭状芽胞杆菌的微生物群足以引发屏障保护反应，防止对食物的过敏反应。在提出的实验中，我们将研究过敏保护性细菌群体如何在细胞和分子水平上向宿主传递信号。提供了七个新的初步数据数字，以支持本订正申请中概述的两个目标。目标1将确定哪些细胞与共生细菌的相互作用是必要的，并且足以诱导屏障保护性反应。我们使用Cre-Lox技术在CD11c+树突状细胞(DC)和肠上皮细胞(IEC)中产生了MyD88信号的靶向突变小鼠。我们还将检查TLR4信令是否是Tregs本身所需的。肠上皮细胞的微阵列分析强调了两个新的基因/通路在梭状芽胞杆菌定植的小鼠上皮中选择性上调；抗菌肽Reg3b和芳香烃受体(AhR)的靶基因。这两条途径都与肠道免疫调节密切相关。Aim 2a将研究AhR介导的信号和IL22如何促进梭状芽胞杆菌诱导的屏障保护性反应，以防止对食物的过敏反应。最后，Aim 2b将研究梭状芽胞杆菌介导的先天性和获得性免疫系统的激活如何影响上皮紧密连接蛋白的表达和功能。所提出的目标的成功完成为开发基于肠道微生物区系组成的预防或治疗食物过敏的新方法带来了希望。

DESCRIPTION (provided by applicant): The prevalence of food allergy is rising at an alarming rate in the United States and in other parts of the developed world. Environmental stimuli that alter populations of beneficial commensal bacteria have been implicated in this increase. Earlier work from our laboratory showed that mice unable to signal via TLR4 exhibit enhanced allergic responses to food. We hypothesized that commensal bacteria were the source of the TLR4 ligand and demonstrated that neonatal administration of a cocktail of broad-spectrum antibiotics (Abx) induced an allergic response in TLR4 sufficient mice equivalent to that seen in TLR4 mutant mice. In the preliminary data presented in this revised application we have established a novel gnotobiotic model of food allergy and show that a defined bacterial consortium, derived directly from the intestinal microbiota of healthy mice, protects against systemic hyperreactivity to a food allergen. We demonstrate that bacteria in the Clostridia class selectively induce a barrier protective response that includes activation of the IL-23/IL-22 axis, induction of the expression of the anti-microbial peptides Reg3b and Reg3g and the expansion of intestinal Tregs and IgA secreting B cells; part of this response is TLR4-dependent. We hypothesize that a Clostridia-containing microbiota is sufficient to elicit a barrier protective response that protets against allergic responses to food. In the experiments proposed we will examine how allergy-protective bacterial populations deliver signals to their hosts at both the cellular and molecular level. Seven new figures of preliminary data are provided in support of the two Aims outlined in this revised application. Aim 1 will determine which cellular interactions with commensal bacteria are necessary and sufficient to induce a barrier protective response. We have used Cre- Lox technology to generate mice with targeted mutations in MyD88 signaling in CD11c+ dendritic cells (DC) and in intestinal epithelial cells (IEC). We will also examine whether TLR4 signaling is required by the Tregs themselves. Microarray analysis of intestinal epithelial cells highlighted two novel genes/pathways selectively upregulated in the epithelium of Clostridia colonized mice; the anti-microbial peptide Reg3b and a target gene for the aryl hydrocarbon receptor (Ahr). Both pathways have been intimately linked to the regulation of intestinal immunity. Aim 2a will examine how Ahr-mediated signals and IL22 contribute to a Clostridia induced barrier protective response that prevents against an allergic response to food. Finally, Aim 2b will examine how Clostridia mediated activation of the innate and adaptive immune system impacts epithelial tight junction protein expression and function. The successful completion of the Aims proposed holds promise for the development of novel approaches to prevent or treat food allergy based on modulation of the composition of intestinal microbiota.