Micro RNA Regulation of Human Airway Epithelial Phenotype

描述(由申请人提供)： 这项申请解决了广泛的挑战领域“(15)翻译科学”和具体的挑战主题“15-HL-102：开发基于microRNA技术的心、肺和血液疾病的新治疗策略”。该项目的题目是《人呼吸道上皮细胞表型的microRNA调控》。MicroRNAs(MiRNAs)在调节基因组和基因方面发挥着重要作用，并被认为参与了与肺部疾病相关的多种生物学过程，如肺发育、肺部炎症、对吸烟和肺癌的反应等。然而，大多数miRNAs在关键的肺细胞类型中的表达和功能仍然知之甚少，例如人支气管上皮(HBe)细胞。当HBe细胞在体外的气液界面(ALI)中生长时，它们概括了体内天然上皮结构和功能的关键方面。本项目的目标是全面了解ALI HBe细胞中的miRNA谱系及其在相关损伤/修复条件下的表达。然后，我们将测试选择的，高度调控的miRNAs在这一重要细胞类型中的功能。这一建议的中心假设是miRNAs是HBe细胞分化和损伤反应的关键调节因子，miRNAs在病理状态下会发生变化，调节miRNAs可以改变HBe细胞的表型，从而达到治疗的目的。为了验证这一假设，我们提出了以下具体目标：目的1)建立HBe细胞在正常分化过程中和相关损伤/修复条件下miRNA表达的综合图谱；以及目的2)确定操纵候选miRNAs在HBe细胞中表达的功能后果。我们将在ALI HBe细胞正常生长和分化以及一系列高度相关的损伤/修复条件下进行最先进的miRNA发现研究和相关的miRNA和mRNA表达阵列研究。我们将操作miRNA的表达作为概念证明，即调节miRNAs可以改变HBe细胞的表型，从而达到治疗的目的。这些实验是通过资源、专业知识和规模经济的独特组合得以实现的，而且雄心勃勃，但可以在RFA-OD-09-003的两年范围内实现。它们通过减轻细胞类型的异质性和抽样的担忧，扩展和补充了对人类患者样本的研究。该模型系统将创建一个有价值的数据库，并将能够评估健康和疾病中HBe细胞的miRNA功能，作为基于miRNA的肺部疾病改进治疗的关键里程碑测试的平台。 与公共卫生相关：在这项研究中，我们将全面确定一种关键的肺细胞类型-人支气管上皮细胞的miRNA谱系，并将测试新的和/或高度调控的miRNAs的功能，以改变它们的结构和功能。我们将创建一个有价值的数据库和一个平台，用于对患有3000多万美国人的肺部疾病的改进治疗进行关键里程碑测试。

DESCRIPTION (provided by applicant): This application addresses the broad challenge area "(15) Translational Science" and specific challenge topic, "15-HL-102: Develop new therapeutic strategies for heart, lung, and blood diseases based on microRNA technology". The project title is "microRNA Regulation of Human Airway Epithelial Cell Phenotype". microRNAs (miRNAs) play critical roles regulating genomes and genes and are known to be involved in multiple biological processes relevant to lung diseases, such as lung development, lung inflammation, the response to tobacco smoking and lung cancer. However, the expression and function of the majority of miRNAs in critical lung cell types, such as human bronchial epithelial (hBE) cells, remains poorly understood. When hBE cells are grown in vitro at an air-liquid interface (ALI), they recapitulate key aspects of the structure and function of the native epithelium in vivo. The goals of this project are to gain a comprehensive understanding of the miRNA repertoire in ALI hBE cells and its expression under relevant injury/repair conditions. We will then test the function of select, highly regulated miRNAs in this important cell type. The central hypothesis of this proposal is that miRNAs are critical regulators of hBE cell differentiation and the response to injury, that miRNAs become altered in pathologic states, and that modulating miRNAs can change hBE cell phenotype for therapeutic benefit. To test this hypothesis we pose the following specific aims: Aim 1) Develop a comprehensive portrait of miRNA expression in hBE cells during normal differentiation and in a spectrum of relevant injury/repair conditions; and Aim 2) Determine functional consequences of manipulating expression of candidate miRNAs in hBE cells. We will conduct state-of-the art miRNA discovery studies and correlative miRNA and mRNA expression array studies in ALI hBE cells during normal growth and differentiation and under a series of highly relevant injury/repair conditions. We will manipulate miRNA expression as proof-of-concept that modulating miRNAs can alter hBE cell phenotype for therapeutic benefit. These experiments are enabled by a unique combination of resources, expertise and economies of scale and are ambitious, but achievable within the 2-year scope of RFA-OD-09-003. They extend and complement studies in human patient samples by mitigating cell type heterogeneity and sampling concerns. The model system will create a valuable database and will enable evaluation of miRNA function in hBE cells in health and disease, serving as a platform for critical milestone testing of improved treatments for lung diseases based on miRNA. PUBLIC HEALTH RELEVANCE: In this study we will comprehensively determine the miRNA repertoire of a key lung cell type, human bronchial epithelial cells, and will test the function of novel and/or highly regulated miRNAs to change their structure and function. We will create a valuable database and a platform for critical milestone testing of improved treatments for lung diseases that afflict greater than 30 million Americans.