The Role of Adenine Nucleotide Translocase in the Protection of Airway Epithelial Cells in Chronic Obstructive Pulmonary Disease (COPD)

腺嘌呤核苷酸转位酶在保护慢性阻塞性肺疾病 (COPD) 气道上皮细胞中的作用

• 批准号: 9764469
• 负责人: Corrine R Kliment
• 金额: $ 16.25万
• 依托单位: UNIVERSITY OF PITTSBURGH AT PITTSBURGH
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-08-16 至 2023-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9764469
• 关键词: Adenine Nucleotide Translocase; Animal Model; Apical; Award; Biological; Biological Models; Biology; Cause of Death; Cell Death; Cell Survival; Cell membrane; Cell physiology; Cell surface; Cells; Cellular Metabolic Process; Chronic Bronchitis; Chronic Obstructive Airway Disease; Chronic lung disease; Cilia; Data; Development; Dictyostelium; Dictyostelium discoideum; Disease; Disease Progression; Disease model; Epithelial; Epithelial Cells; Foundations; Frequencies; Functional disorder; Gene Expression; Genetic; Golgi Apparatus; Health Care Costs; Homeostasis; Human; Hydration status; Immunoblotting; Immunofluorescence Immunologic; Impairment; Injury; Inner mitochondrial membrane; Knockout Mice; Lung; Lung diseases; Membrane; Mentors; Metabolism; Mitochondria; Mitochondrial Membrane Protein; Modeling; Modification; Molecular; Morbidity - disease rate; Mucociliary Clearance; Mus; Obstructive Lung Diseases; Pathogenesis; Pathway interactions; Patients; Peptide Signal Sequences; Phenotype; Physicians; Plasma; Play; Process; Production; Protein Isoforms; Proteins; Regulation; Research; Respiration; Risk Factors; Role; SLC25A4 gene; SLC25A5 gene; Scheme; Scientist; Smoke; Solid; Surface; Testing; Therapeutic; Time; Tissue imaging; Tissues; Training; United States National Institutes of Health; Work; airway epithelium; airway surface liquid; body system; career; career development; cellular imaging; cigarette smoke; cigarette smoke-induced; cilium motility; effective therapy; exposure to cigarette smoke; fascinate; gain of function; genetic selection; human model; mitochondrial metabolism; mortality; mouse model; new therapeutic target; novel; overexpression; preservation; prevent; programs; protective effect; reconstitution; skills; therapeutic target; tool; trafficking

PROJECT SUMMARY The overall objectives of this NIH K08 Career Mentored Award are to identify and investigate new therapeutic targets for epithelial injury due to cigarette smoke in chronic obstructive lung disease (COPD), and to facilitate the development of essential skills that will allow the candidate to become an effective and independent physician-scientist. The candidate and her mentors have constructed a rigorous training plan that will provide the foundation for a strong and successful academic career. This proposal explores the core mechanisms of how adenine nucleotide translocase (ANT) is localized to the plasma membrane of ciliated airway epithelium and further how ANT is utilized for more advanced cellular functions, such as ciliary ATP regulation impacting airway hydration and ciliary beating. COPD is the 3rd leading cause of death in the US with cigarette smoke (CS) being the primary insult leading to disease. Despite ongoing mammalian research, no new biologic targets have been identified, and current therapies have limited effect on disease progression or mortality. I have leveraged the power of a genetically tractable model system, the amoebozoan Dictyostelium discoideum, as a genetic selection discovery tool for lung biology. I identified human ANT (ANT1 and ANT2 present in lung) as protective against CS-induced cell death in human bronchial epithelial cells. ANT is a mitochondrial ADP/ATP transporter that plays an active role in mitochondrial metabolism and ATP homeostasis. While exploring the localization of ANTs in human airway tissue, I made a fascinating observation that in addition to mitochondrial localization, human ANT1 and ANT2 localize to the plasma membrane of motile cilia in human airway and isolated primary airway cells (Normal Human Bronchial Epithelial cells, NHBE). GFP-tagged adenoviral ANT1 and ANT2 also go to the ciliary membrane (and mitochondria) in primary NHBEs and ANT1 and ANT2 are present in isolated human ciliary axonemes by western analysis. In addition, ANT2 specifically enhances airway surface hydration and preserves ciliary beat frequency in the context of CS. This early career proposal answers the following questions: How does human ANT1 and ANT2 localize to the plasma membrane of airway epithelium; how do plasma membrane versus mitochondrial ANTs regulate airway metabolism, airway hydration and ciliary function in ciliated airway epithelium; and what is the impact of ANT2 loss and gain of function on airway dysfunction and COPD development through mouse models of disease given its protective phenotype on airway epithelium. This will be accomplished using a combination of molecular tools, ANT isoform genetic modification, cell and tissue imaging, real-time analysis of cellular metabolism, assessment of airway hydration and ciliary function, and correlation with a mouse model of smoke exposure. Ultimately, the role of ANT in lung disease has yet to be described and therapeutic manipulation of its biology may allow us to drive cells to a healthier state. The core biology themes explored here will likely have broad applicability and impact other lung diseases.

项目摘要 NIH K08职业指导奖的总体目标是识别和调查新的治疗性 由于慢性阻塞性肺疾病（COPD）中的香烟烟雾引起的上皮损伤的靶标，并促进 基本技能的发展，这将使候选人成为有效而独立的 医师科学家。候选人和她的导师制定了一项严格的培训计划，将提供 强大而成功的学术生涯的基础。该建议探讨了 腺嘌呤核苷酸易位酶（ANT）如何定位于纤毛气道上皮的质膜 进一步的蚂蚁如何用于更先进的细胞功能，例如纤毛ATP调节影响 气道水合和睫毛跳动。 COPD是美国的第三大死亡原因，香烟烟雾 （CS）是导致疾病的主要侮辱。尽管正在进行哺乳动物研究，但没有新的生物学 已经确定了靶标，并且当前疗法对疾病进展或死亡率的影响有限。我 已经利用了遗传障碍模型系统的功能，即变形虫dictyostelium discoideum， 作为肺部生物学的遗传选择发现工具。我确定了人类蚂蚁（肺中存在的ANT1和ANT2） 作为对人支气管上皮细胞中CS诱导的细胞死亡的保护作用。蚂蚁是线粒体 ADP/ATP转运蛋白在线粒体代谢和ATP稳态中起积极作用。尽管 探索蚂蚁在人类气道组织中的定位，我进行了一个有趣的观察，除了 线粒体定位，人ANT1和ANT2定位于人类纤毛的质膜 气道和孤立的原代气道细胞（正常的人支气管上皮细胞，NHBE）。 GFP标记 腺病毒ANT1和ANT2也转到原代NHBE和ANT1中的纤毛膜（和线粒体） 通过西方分析，孤立的人类睫状轴突中存在和ANT2。另外，ANT2特别 增强气道表面水合，并在CS的背景下保留睫状节频率。这个早期职业 提案回答以下问题：人类ANT1和ANT2如何本地化到等离子体 气道上皮的膜；质膜与线粒体蚂蚁如何调节气道 纤毛气道上皮的代谢，气道水合和睫状功能； ANT2有什么影响 通过小鼠疾病模型，气道功能障碍和COPD开发的功能丧失和功能 鉴于其在气道上皮上的保护性表型。这将使用 分子工具，蚂蚁同工型遗传修饰，细胞和组织成像，细胞的实时分析 代谢，气道水合和睫状功能的评估以及与烟雾模型的相关性 接触。最终，蚂蚁在肺部疾病中的作用尚未描述，并且对 它的生物学可能使我们能够将细胞驱动到更健康的状态。这里探索的核心生物学主题可能很可能 具有广泛的适用性并影响其他肺部疾病。