The function and regulation of IL-33 in the airway epithelium in asthma

描述(由申请人提供)：在这份题为“哮喘患者呼吸道上皮细胞中IL-33的功能和调节”的修订申请中，我提出了加州大学旧金山分校肺部医学部的一项全面的研究和职业发展计划。这项建议不仅将使我能够发展建立独立的学术研究生涯所需的技能、技术和合作，而且它还将推动哮喘生物学领域的发展。我是这个指导职业发展奖的理想人选，因为我坚定地致力于建立一个独立的研究生涯。在加州大学伯克利分校读本科时，我开始对研究科学感兴趣，在那里我在一个病毒学实验室工作了三年。我以优异的成绩毕业，获得了全额学费奖学金，在南加州大学学习医学。从医学院班级第一名毕业后，我选择在加州大学圣地亚哥分校完成内科住院医师培训，在那里我在帕特里夏·芬恩博士的实验室进行基础科学研究。在那里，我对哮喘生物学产生了浓厚的兴趣，并参加了美国内科委员会的研究培训计划。这使我能够很早就开始在加州大学旧金山分校担任肺危重护理研究员，并花更多的时间进行研究。在加州大学旧金山分校时，我继续在约翰·法希的实验室里研究哮喘。在Fahy博士、Erle博士、Woodruff博士和Seppard博士的研究和职业指导下，我完成了一个专注于Periostin在哮喘中的作用的项目。我在2012年1月发表了这些结果。自那以后，我一直专注于IL-33在人类哮喘中的作用，这也是我目前提案的主题。我对研究事业的奉献，加上坚持不懈和成功的记录，将使我能够最大限度地受益于这个奖项。我已经概述了这项应用的研究计划，重点是了解IL-33在人类哮喘中的功能和调节。哮喘是一种慢性肺部疾病，影响着近3000万美国人和全球约3亿人。虽然存在有效的治疗方法，但许多哮喘患者对可用的治疗方案没有最佳反应；因此，需要新的治疗方法。长期以来被认为是一种免疫系统疾病，越来越多的人认识到呼吸道上皮在协调哮喘的免疫反应中的重要性。新近发现的上皮细胞细胞因子IL-33及其受体ST2在多项大规模全基因组相关性研究中与哮喘的发病机制密切相关。小鼠模型表明，IL-33通过ST2受体发挥作用，促进Th2炎症。然而，对IL-33在人类哮喘中的研究是有限的。在这项资助中，我将集中讨论IL-33生物学中的三个重要问题：1)IL-33是如何从呼吸道上皮细胞释放出来的；2)I-33与人类哮喘中Th2炎症的关系是什么；以及3)ST2基因上的SNPs在可溶性抑制物Sst2调节IL-33方面有什么作用？IL-33在呼吸道上皮细胞胞核有结构性表达，但缺乏信号序列，其释放机制尚不清楚 是未知的。我已经开发了一种新的模型系统，通过在呼吸道上皮细胞系中表达GFP标记的IL-33，这将使我能够剖析导致IL-33释放的途径。 据推测，IL-33一旦释放，就会在呼吸道内诱导Th2炎症。利用加州大学旧金山分校呼吸道临床研究中心庞大的人类生物谱库，我将确定IL-33与急性和慢性哮喘患者Th2炎症和粘膜重塑的标记物之间的关系。最后，细胞外IL-33可受其可溶性抑制物Sst2的调节。这就做 对大量哮喘患者的ST2基因座进行测序，并确定与ST2上皮细胞和痰小球基因表达改变相关的SNPs。我将使用启动子报告试验和功能生物测定来测试Candidat SNPs在人类原代上皮细胞培养中的功能。这项研究计划不仅将推动哮喘生物学领域的发展，还将使我能够发展新的研究技能和合作，这对我作为一名独立研究者的成功至关重要。最后，我概述了一个利用加州大学旧金山分校众多机会的职业发展计划。我的主要导师约翰·法希非常适合担任我的顾问。他带来了哮喘的呼吸道上皮细胞生物学和分子表型方面的专业知识，并是加州大学旧金山分校大型人类哮喘组织库的主任。我已经组建了一个由其他顾问和合作者组成的团队，其中包括谢泼德博士、厄尔、伍德拉夫和塞博尔德，他们拥有互补的技能。他们将是我在细胞生物学、原代呼吸道上皮细胞转基因、分子克隆、生物统计学和高级人类遗传学方面的理想顾问。我将在加州大学旧金山分校开设遗传学、细胞生物学、免疫学和生物统计学的正式课程，以及每周的免疫学和普通肺部医学研讨会，以补充这个团队的指导。医学部和肺危重护理司在支持和培训早期内科科学家方面有着良好的记录，并拥有资源充足的实验室和核心设施。这些资源对执行我的研究计划将是无价的，并将确保我的成功。医学系对我的职业发展道路和这份申请给予了大力支持。2012年7月，我被任命为肺部重症监护科助理教授，有80%的保护研究时间。有了这些资源和支持，我将很好地建立一个成功的独立研究生涯。

DESCRIPTION (provided by applicant): In this revised application entitled "Function and Regulation of IL-33 in Airway Epithelial Cells in Asthma" for the Mentored Clinical Scientist Research Career Development Award, I propose a comprehensive plan of research and career development at the University of California, San Francisco in the Division of Pulmonary Medicine. This proposal will not only enable me to develop the skills, techniques and collaborations necessary to establish an independent academic research career, but it will also advance the field of asthma biology. I am an ideal candidate for this mentored career development award, as I am strongly committed to establishing an independent research career. I became interested in research science as an undergraduate at the University of California, Berkeley where I worked in a virology laboratory for three years. I graduated with honors earning a full tuition scholarship to study medicine at the University of Southern California. Graduating first in my medical school class, I chose to complete my Internal Medicine residency training at UC San Diego where I performed basic science research in the laboratory of Dr. Patricia Finn. There I developed a strong interest in asthma biology and enrolled in the American Board of Internal Medicine Research Training Program. This allowed me to begin fellowship in Pulmonary Critical Care at UCSF early and spend additional years performing research. While at UCSF, I continued studying asthma in the laboratory of John Fahy. Under the research and career guidance of Dr. Fahy, Erle, Woodruff, and Sheppard, I completed a project focused on the role of periostin in asthma. I published these results in January of 2012. I have since focused on the role of IL- 33 in human asthma, and this is the topic of my current proposal. My dedication to a research career along with persistence and a track record of success will enable me to maximally benefit from this award. I have outlined a research plan in this application, which focuses on understanding the function and regulation of IL-33 in human asthma. Asthma is a chronic lung disease, which affects nearly 30 million Americans and an estimated 300 million people worldwide. Although effective treatments exist, many asthmatics do not respond optimally to available regimens; thus, novel treatments are needed. Long considered a disease of the immune system, there is increasing recognition of the importance of the airway epithelium in orchestrating the immune responses in asthma. The newly described epithelial cell cytokine IL-33 and its receptor ST2 have been strongly implicated in asthma pathogenesis in multiple large-scale genome wide association studies. Mouse models suggest that IL-33, acting through the ST2 receptor, promotes Th2 inflammation. Studies of IL-33 in human asthma are limited however. In this grant, I will focus on three important questions in IL-33 biology: 1) how is IL-33 released from airway epithelial cells, 2) what is the relationship of I-33 to Th2 inflammation in human asthma, and 3) what is the function of SNPs in the ST2 locus on the regulation of IL-33 by the soluble inhibitor sST2? IL-33 is constitutively expressed in the nuclei of airway epithelial cells, but it lacks a signal sequence, and the mechanism of its release is unknown. I have developed a novel model system by expressing GFP-tagged IL-33 in an airway epithelial cell line, which will enable me to dissect the pathways leading to IL-33 release. Once released, IL-33 is postulated to induce Th2 inflammation in the airway. Using the vast human biospecimen bank at the UCSF Airway Clinical Research Center, I will determine the relationship of IL-33 to markers of Th2 inflammation and mucous remodeling in both acute and chronic asthma. Finally, extracellular IL-33 can be regulated by its soluble inhibitor sST2. I will sequence the ST2 locus from a large number of asthmatics and identify SNPs associated with altered ST2 epithelial cell and sputum pellet gene expression. I will test the function of candidat SNPs in human primary epithelial cell culture using promoter reporter assays and a functional bioassay. This research plan will not only advance the field of asthma biology, but will enable me to develop new research skills as well as collaborations that will be essential for my success as an independent investigator. Finally, I have outlined a career development plan that takes advantage of the many opportunities here at UCSF. My primary mentor, John Fahy, is ideally suited to serve as my advisor. He brings expertise in airway epithelial cell biology and molecular phenotyping in asthma and is the director of the large UCSF human asthma tissue bank. I have assembled a team of additional advisors and collaborators, including Dr. Sheppard, Erle, Woodruff and Seibold, who have complementary skills. They will be ideal in advising me on cell biology, primary airway epithelial cell transfection, molecular cloning, biostatistics, and advanced human genetics. I will supplement the guidance of this team with formal coursework offered at UCSF in Genetics, Cell Biology, Immunology, and Biostatistics as well as weekly seminars in Immunology and general pulmonary medicine. The Department of Medicine and the Division of Pulmonary Critical Care has a strong track record of support and training for early-stage physician scientists and boasts well-resourced laboratories and core facilities. These resources will be invaluable in carrying out my research plan and will ensure my success. The Department of Medicine strongly supports my career path and this application. In July 2012, I was appointed to Assistant Professor within the Division of Pulmonary Critical Care with 80% protected research time. With these resources and support, I will be well positioned to establish a successful independent research career.