Epigenetic regulation of esophageal epithelial barrier function in eosinophilic esophagitis

项目总结 这项拟议的研究探索了嗜酸性食管炎上皮屏障功能障碍的机制 (EoE)。在疾病活动期，EoE的特点是Th2型、富含嗜酸性粒细胞的炎症和高度的 IL-13、IL-4和趋化因子水平。越来越多的人意识到上皮功能障碍 在活动期，EoE可能与炎症成分发挥着同样重要的作用，因为它对变应原致敏 以及暴露在微生物中。此外，数据表明，对于许多患者来说，上皮功能障碍仍然存在 在非活动期EoE期间，当治疗已清除炎性浸润物时。持续性上皮病的发病机制 人们对功能障碍知之甚少。然而，我们和其他人观察到了一种可复制的、调控失调的基因 上皮结构蛋白异常表达的活动期EoE的表达模式。我们证明了 EoE患者的上皮细胞在体外培养时维持这种异常基因表达模式的某些方面 中立的文化环境。这表明表观遗传机制维持了这些基因的表达。 无炎症时EoE上皮细胞的形态。值得注意的是，IL-13对对照上皮细胞的体外处理 细胞复制EoE粘膜转录组的相似方面，提示这种Th2细胞因子具有特定的作用 在EoE上皮屏障功能障碍中的作用。白介素13相关的上皮功能的表观遗传学改变 显示在几种类型的上皮中，但在食道中受影响的部位和功能暗示是 不清楚。这一建议的假设是IL-13诱导食道上皮的表观遗传学变化 EoE患者，导致持续性上皮功能障碍。这项工作将利用与培训团队的联系 凭借在表观遗传学、转录学和生物信息学方面的专业知识，与儿科医生合作具有独特的优势 EoE患者群体。目标1将使用食道鳞状细胞的三维气液界面模型 确定慢性体外暴露于IL-13治疗是否会导致表观遗传学改变和屏障 非EoE对照组患者的初级上皮功能障碍。这个目标的目的是确定IL-13是否 治疗与组蛋白标记的改变和已知影响基因的基因表达变化有关 屏障功能。Aim 2将使用患有和不患有EoE的患者的活检组织来检查组蛋白的变化 标记和基因在食道上皮中的表达。此目标的目标是确定并本地化一组 活动期和非活动期EoE中持续表达异常的上皮基因(与对照组相比 病人)。这些研究的结果将有助于对食道上皮屏障功能障碍的理解 和EoE疾病持续性，并为申请人的第一次R01提交提供基础。最重要的是 本提案中研究策略和培训计划的目标是将候选人发展成为独立的 研究人员领导着一个强大的研究计划，利用表观遗传学的研究和分析技能， 转录学和生物信息学在非IgE介导的食物过敏领域的应用。

PROJECT SUMMARY The proposed research explores the mechanisms of epithelial barrier dysfunction in eosinophilic esophagitis (EoE). During its active disease phase, EoE is characterized by Th2-type, eosinophil-rich inflammation with high levels of IL-13, IL-4 and chemoattractant chemokines. There is a growing appreciation that epithelial dysfunction in active EoE may play an equally important role as the inflammatory component due to sensitization to allergens and exposure to microbes. Additionally, data suggests that for many patients, epithelial dysfunction persists during inactive EoE when therapy has cleared the inflammatory infiltrate. The mechanism of persistent epithelial dysfunction is poorly understood. However, we and others have observed a reproducible, dysregulated gene expression pattern during active EoE with aberrant expression of epithelial structural proteins. We show that epithelial cells from EoE patients maintain aspects of this dysregulated gene expression pattern when grown in a neutral culture environment. This suggests that epigenetic mechanisms maintain these gene expression patterns in EoE epithelium in the absence of inflammation. Notably, in vitro IL-13 treatment of control epithelial cells replicates similar aspects of the EoE mucosal transcriptome, suggesting a specific role for this Th2 cytokine in the epithelial barrier dysfunction of EoE. IL-13-associated epigenetic alteration of epithelial function has been shown in several types of epithelium but the affected loci and functional implications in the esophagus are unclear. The hypothesis of this proposal is that IL-13 induces epigenetic changes in the esophageal epithelium of patients with EoE, leading to persistent epithelial dysfunction. The work will leverage access to a training team with expertise in epigenetics, transcriptomics and bioinformatics is uniquely situated to work with a pediatric population of EoE patients. Aim 1 will use a three dimensional air-liquid interface model of esophageal squamous epithelium to determine if chronic in vitro exposure to IL-13 treatment induces epigenetic changes and barrier dysfunction in primary epithelium from non-EoE control patients. The goal of this aim is to identify if IL-13 treatment is associated with alterations in histone marks and gene expression changes in genes known to affect barrier function. Aim 2 will use biopsy tissue from patients with and without EoE to examine changes in histone marks and gene expression in the esophageal epithelium. The goal of this aim is to identify and localize a set of epithelial genes with persistently dysregulated expression in active and inactive EoE (when compared to control patients). The outcome of these studies will inform the understanding of esophageal epithelial barrier dysfunction and EoE disease persistence, and provide the basis for the applicant’s first R01 submission. The overarching goal of the research strategy and training plan in this proposal is to develop the candidate into an independent investigator leading a robust research program that utilizes research and analysis skills in epigenetics, transcriptomics and bioinformatics in the field of non-IgE mediated food allergies.