Molecular Interrogation of the Host-Pathogen Interface in Idiopathic Subglottic Stensosis

项目摘要/摘要 特发性声门下狭窄(ISGS)是一种原因不明的声门下黏膜炎症性疾病。 几乎只发生在成年高加索女性的上呼吸道。病人需要紧急服务 防止因呼吸道阻塞而死亡的手术。新的治疗方法旨在阻止 进行性呼吸道纤维化是减轻这种疾病负担的关键。考虑到它的稀有性， 受影响患者的地理分布以前限制了定义基本的 疾病的生物学。我们最近发表的文章揭示了ISGS呼吸道疤痕中的微生物失调， 伴随着炎症途径的激活。利用拥有1000名患者的国际ISGS队列 在这项提案的PI(ISGS1000)的领导下，该项目旨在了解 通过回答以下问题：ISGS与活动性感染有关，还是与细菌有关 触发自我反应性免疫反应。我们的提案利用独立但又相互关联的 解决这一问题的方法。在目标1中，我们将应用尖端的分子免疫学 以及生物信息学技术从呼吸道瘢痕中筛选浸润性CD8+T细胞，直接测序 通过RNAseq的单个T细胞受体(TCR)，然后将高频TCR克隆到体外 用于绘制抗原特异性图谱的模型系统。这将使我们能够研究CD8+ ISGS呼吸道瘢痕粘膜中的T细胞表现出针对细菌的克隆性反应 抗原。然后在目标2中，我们将利用ISGS中细菌的全基因组测序(WGS 呼吸道疤痕以确认独特的细菌与ISGS的关联。精确分子 利用变转录分析鉴定细菌将提供新的信息 影响致病性的遗传特征。在目标3中，我们将调查目前的治疗方法是如何 ISGS既影响当地的微生物群，也影响宿主的免疫。把我们的独立人士 但相互关联的方法将有助于确定宿主和病原体如何碰撞以产生致病 ISGS中的组织纤维化。

Project Summary/Abstract Idiopathic subglottic stenosis (iSGS) is an unexplained mucosal fibroinflammatory disease of the upper airway that occurs almost exclusively in adult, Caucasian women. Patients require urgent surgery to prevent death from airway obstruction. Novel therapies aimed at halting the progressive airway fibrosis are critical to reduce the burden of this disease. Given its rarity, the geographical dispersal of affected patients has previously limited investigations to define the basic biology of the disease. Our recent publications reveal microbial dysbiosis in iSGS airway scar, along with inflammatory pathway activation. Leveraging a 1000-patient international iSGS cohort led by this proposal’s PI (iSGS1000), this project is designed to understand the pathogenesis of iSGS by answering the question: Is iSGS related to an active infection, or does bacteria trigger a self-reactive immune response. Our proposal utilizes independent but interrelated approaches to address this question. In Aim 1 we will apply cutting edge molecular immunology and bioinformatic techniques to sort infiltrating CD8+ T cells from airway scar, directly sequence individual T cell receptors (TCR) via RNAseq, then clone high frequency TCRs into an in vitro model system to map antigen specificity. This will allow us to investigate the hypothesis that CD8+ T cells in the mucosa of iSGS airway scar demonstrate a clonal response directed at a bacterial antigen. Then in Aim 2 we will utilize whole genome sequencing (WGS) of the bacteria in iSGS airway scar to confirm a unique bacterial association with iSGS. Precise molecular characterization of the bacteria with metatranscriptomic analysis will provide new information on genetic features impacting pathogenicity. In Aim 3 we will investigate how current treatments for iSGS impact both the local microbiome as well as host immunity. Taken together our independent but interrelated approaches will help define how host and pathogen collide to produce pathogenic tissue fibrosis in iSGS.