Mitochondrial Genes as New Targets in the Protection of Airway Epithelial Cells Against Cigarette Smoke

项目总结 NRSA个人奖学金的总体目标是确定和研究新的治疗靶点。 治疗慢性阻塞性肺疾病(COPD)中因吸烟引起的上皮损伤，并促进 培养基本技能，使应聘者成为一名有效的内科科学家。这个 候选人和她的导师已经制定了一份严格的培训计划，这将为 扎实成功的学术生涯。该提案涉及的基本生物学过程是 线粒体和代谢调节与吸烟相关的肺部疾病有关。慢性阻塞性 肺部疾病(COPD)是美国第三大死亡原因，吸烟(CS)是最主要的死亡原因 最初的侮辱导致疾病。尽管哺乳动物研究仍在进行，但还没有新的生物靶点 目前的治疗方法对疾病进展或死亡率的影响有限。我们正在利用 盘状网囊阿米巴原虫与人呼吸道上皮细胞的比较研究 寻找预防CS的新途径和治疗策略。作为一种氧化侮辱，CS导致 形态和代谢变化。我们已经发现CS导致了相关的生长表型 网柄网柄菌中的抑制作用。此外，使用cDNA文库从约35,000个网柄蕨类植物中进行遗传选择 在香烟烟雾提取物(CSE)作用下，我们确定腺嘌呤核苷酸转位酶 (ANT)作为对CS诱导的生长缺陷的保护。Ant是线粒体ADP/ATP转运蛋白 这对线粒体新陈代谢和能量动态平衡起着积极的作用。目前，蚂蚁的角色是 线粒体系统在保护肺上皮细胞免受吸烟损伤中的作用尚不清楚。 该提案的长期目标是确定蚂蚁通过哪些机制来保护自己免受 CS损伤，并确定新的潜在治疗靶点。在目标1中，候选人将破译 人蚂蚁对吸烟(CS)后支气管上皮细胞(HBE)细胞活性和形态的影响 这将通过分子表达工具、细胞成像和遗传学的组合来完成 对这些蛋白质进行修饰以评估细胞的活性和结构。在目标2中，候选人将确定 人类蚂蚁保护内皮细胞免受CS损伤的代谢和炎症机制。这 将通过评估蚂蚁引起的实时线粒体新陈代谢和细胞因子变化来完成 HBE中的调制。这些实验的目标是利用哺乳动物和网柄蕨类动物模型 剖析导致慢性阻塞性肺疾病上皮细胞失调的线粒体机制。完成这些目标 将阐明线粒体蚂蚁在肺上皮细胞对CS的反应中所起的作用，作为一种新的保护 慢性阻塞性肺疾病的通路。此外，这项工作将为研究新的治疗策略奠定基础。 使用体内翻译模型和FDA批准的药库用于COPD。

PROJECT SUMMARY The overall objectives of this NRSA individual fellowship 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 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. The proposal addresses the fundamental biological processes of mitochondrial and metabolic regulation involved in cigarette smoking-related lung disease. Chronic obstructive pulmonary disease (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. We are leveraging the amoebozoan Dictyostelium discoideum to develop a comparative approach with human airway epithelial cells to identify new pathways and therapeutic strategies protective against CS. As an oxidative insult, CS results in morphologic and metabolic changes. We already found that CS leads to a relevant phenotype of growth suppression in Dictyostelium. Furthermore, using a cDNA library genetic selection from ~35,000 Dictyostelium cDNA transformants exposed to cigarette smoke extract (CSE), we identified adenine nucleotide translocase (ANT) as protective against CS-induced growth defects. ANT is a mitochondrial ADP/ATP transport protein that plays an active role in mitochondrial metabolism and energy homeostasis. Currently, the role of the ANT mitochondrial system in protecting the lung epithelium against cigarette-smoke induced injury remains unclear. The long-term goal of the proposal is to determine the mechanisms through which ANT is protective against CS injury and identify new potential therapeutic targets. In Aim 1, the candidate will decipher the effects of human ANT on cell viability and morphology in bronchial epithelial cells (HBEs) after cigarette smoke (CS). This will be accomplished by a combination of molecular expression tools, cell imaging, and genetic modification of these proteins to assess cell viability and structure. In Aim 2, the candidate will identify the metabolic and inflammatory mechanisms through which human ANT protects against CS injury in HBEs. This will be accomplished by evaluating real-time mitochondrial metabolism and cytokine changes due to ANT modulation in HBEs. The goal of these experiments is to leverage mammalian and Dictyostelium models to dissect the mitochondrial mechanisms leading to epithelial dysregulation in COPD. Completion of these aims will elucidate the role of the mitochondrial ANT in lung epithelium in response to CS, as a novel protective pathway in COPD. Furthermore, this work will lay groundwork for examination of new therapeutic strategies for COPD using translational in vivo models and an FDA-approved drug library.