Mechanisms of Asbestos-Induced Alveolar Epithelial Cell Injury

石棉引起的肺泡上皮细胞损伤的机制

• 批准号: 9273259
• 负责人: DAVID W KAMP
• 金额: --
• 依托单位: JESSE BROWN VA MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-04-01 至 2020-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9273259
• 关键词: 8-Oxoguanine DNA Glycosylase; Acetylation; Aconitate Hydratase; Alveolar; Amphiboles; Apoptosis; Asbestos; Asbestosis; Attenuated; Bleomycin; Bronchogenic Carcinoma; Cell Death; Cell Survival; Cells; Cessation of life; Crocidolite Asbestos; DNA; DNA Damage; DNA Repair; DNA Repair Enzymes; Disease; Electron Transport; Epithelial Cells; Event; Exposure to; Fiber; Fibrosis; Hamman-Rich syndrome; Health; Human; In Vitro; International Union Against Cancer; Lung; Lung diseases; Malignant Neoplasms; Malignant neoplasm of lung; Mediating; Mesothelioma; Mitochondria; Mitochondrial DNA; Mitochondrial Proteins; Molecular; Mus; Neurons; OGG1 gene; Oxidants; Oxidative Stress; Pathogenesis; Pathway interactions; Pharmacology; Population; Preparation; Protein Acetylation; Pulmonary Fibrosis; Reactive Oxygen Species; Resveratrol; Role; SOD2 gene; Signal Transduction; Sirtuins; TP53 gene; Toxin; Veterans; Wild Type Mouse; alveolar type II cell; biological adaptation to stress; carcinogenesis; cell injury; effective therapy; genetic approach; in vivo; injury and repair; innovation; insight; mitochondrial dysfunction; mitochondrial metabolism; mouse model; novel; overexpression; prevent; protein function; public health relevance

 DESCRIPTION (provided by applicant): Asbestos causes asbestosis (pulmonary fibrosis due to asbestos) and malignancies (lung cancer and mesothelioma) by mechanisms that are not fully elucidated. The extent of alveolar epithelial cell (AEC) injury and repair are critical determinants of the fibrogenic potential of toxins, such as asbestos. Sirtuin 3 (SIRT3), the major mitochondrial deactylase, regulates mitochondrial metabolism that detoxifies mitochondrial reactive oxygen species (ROS). We have shown that mitochondrial ROS mediate asbestos-induced AEC DNA damage and apoptosis by a p53- and mitochondria-regulated death pathway as well as a novel mechanism by which mitochondrial human 8-oxoguanine-DNA glycosylase 1 (mt-hOGG1) prevents ROS-induced AEC apoptosis by preserving mitochondrial aconitase (ACO-2), which in turn prevents mitochondrial DNA (mtDNA) damage. Compared to wild type (WT) mice, we showed that Ogg1-/- mice have increased asbestos-induced lung fibrosis due in part to alveolar type II (AT2) cell apoptosis from reduced ACO-2 levels and increased mtDNA damage. SIRT3, which controls the function of mitochondrial proteins including OGG1, ACO-2, and others, can augment mtDNA repair in the setting of oxidative stress in nerve cells. Our preliminary studies show that asbestos reduces AEC SIRT3 expression; that SIRT3 deficiency promotes asbestos-induced AEC mtDNA damage, apoptosis, and pulmonary fibrosis; and that SIRT3 enforced expression (EE) attenuates oxidant-induced AEC ACO-2 depletion, mtDNA damage, and apoptosis. We reason that AEC mtDNA is a key target that integrates cell survival / death signals following exposure to asbestos by a SIRT3-regulated mechanism. HYPOTHESIS: SIRT3 attenuates asbestos-induced AEC mtDNA damage by preserving mt-OGG1/ ACO-2 function and reducing mitochondrial dysfunction important for limiting apoptosis and lung fibrosis. Our SPECIFIC AIMS that will be examined over the next 4 years include: (1) To determine if SIRT3 deficiency promotes AEC mtDNA damage and intrinsic apoptosis due to alterations in OGG1 and/or ACO-2 acetylation that directs protein function. We will assess the effects of SIRT3 deficiency on acetylation of OGG1 and ACO-2, mtDNA damage, and apoptosis. Sirt3-/- and lung epithelial cell specific Sirt3-/- mice will be used to assess whether SIRT3 deficiency augments asbestos- and bleomycin-induced AEC MnSOD, OGG1 and ACO-2 acetrylation, mtDNA damage, apoptosis, and pulmonary fibrosis. (2) To determine whether SIRT3 enforced expression (EE) prevents asbestos-induced AEC mitochondrial protein acetylation (total, OGG1, ACO-2, MnSOD), mtDNA damage, and apoptosis resulting from altered OGG1 / ACO-2. We will use a combination of pharmacologic (resveratrol / viniferin) and genetic approaches in vitro. We will use Sirt3-EE mice available to us and lung epithelial cell specific Sirt3 mice that we will develop to assess whether SIRT3 attenuates asbestos- and bleomycin-induced lung fibrosis by preventing AEC mitochondrial protein acetylation (OGG1 and ACO-2), ACO-2 depletion, mtDNA damage, and apoptosis. (3) To determine if mitochondrial hOGG1-EE attenuates the deleterious effects of SIRT3 deficiency as occurs following asbestos exposure. In vitro studies using MLE-12 cells that overexpress mt-Ogg1 and are SIRT3 depleted will be used to assess asbestos-induced mitochondrial protein acetylation (total, OGG1, ACO-2, and MnSOD), ACO-2 levels, mtDNA damage, and intrinsic apoptosis. We will also use Mitochondrial hOgg1-EE mice in the absence and presence of SIRT3 silencing to ascertain if mt-hOGG1-EE attenuates lung fibrosis (asbestos / bleomycin) by preserving AEC ACO-2 levels and mtDNA while diminishing apoptosis. Innovation: These studies will elucidate the importance of SIRT3 in maintaining AT2 cell mtDNA integrity crucial for preventing AT2 cell apoptosis and pulmonary fibrosis. The studies proposed will advance our understanding of lung fibrosis that may have broader implications for more common lung disorders (i.e. IPF and lung cancer) present in the veteran population for which effective treatments are needed.

 描述（由申请人提供）： 石棉导致石棉肺（石棉引起的肺纤维化）和恶性肿瘤（肺癌和间皮瘤）的机制尚未完全阐明。肺泡上皮细胞 (AEC) 损伤和修复的程度是毒素纤维化潜力的关键决定因素，例如。 Sirtuin 3 (SIRT3) 是一种主要的线粒体脱乙酰酶，可调节线粒体代谢，从而解毒线粒体反应性。我们已经证明，线粒体 ROS 通过 p53 和线粒体调节的死亡途径以及线粒体人 8-氧鸟嘌呤-DNA 糖基化酶 1 (mt) 介导石棉诱导的 AEC DNA 损伤和细胞凋亡。 -hOGG1) 通过保留线粒体乌头酸酶 (ACO-2) 来防止 ROS 诱导的 AEC 细胞凋亡，从而与野生型相比，防止线粒体 DNA (mtDNA) 损伤。在 WT 型 (WT) 小鼠中，我们发现 Ogg1-/- 小鼠石棉诱导的肺纤维化增加，部分原因是 ACO-2 水平降低和控制功能的 mtDNA 损伤增加导致肺泡 II 型 (AT2) 细胞凋亡。我们的初步研究表明，石棉 AEC SIRT3 表达； SIRT3 缺乏会促进石棉诱导的 AEC mtDNA 损伤、细胞凋亡和肺纤维化；SIRT3 强制表达 (EE) 会减弱氧化剂诱导的 AEC ACO-2 耗竭、mtDNA 损伤和细胞凋亡。通过 SIRT3 调节机制整合暴露于石棉后的细胞存活/死亡信号。石棉通过保留 mt-OGG1/ACO-2 功能和减少线粒体功能障碍来诱导 AEC mtDNA 损伤，这对限制细胞凋亡和肺纤维化至关重要。我们将在未来 4 年内进行检查的具体目标包括：(1) 确定 SIRT3 是否缺乏。由于指导蛋白质功能的 OGG1 和/或 ACO-2 乙酰化的改变，促进 AEC mtDNA 损伤和内在细胞凋亡。我们将评估 SIRT3 缺陷对的影响。 OGG1 和 ACO-2 的乙酰化、mtDNA 损伤和细胞凋亡将用于评估 SIRT3 缺陷是否会增强石棉和博来霉素诱导的 AEC MnSOD、OGG1 和 ACO-。 2乙酰化、mtDNA损伤、细胞凋亡和肺纤维化(2)确定SIRT3是否强制表达(EE)。防止石棉诱导的 AEC 线粒体蛋白乙酰化（总、OGG1、ACO-2、MnSOD）、mtDNA 损伤以及 OGG1/ACO-2 改变导致的细胞凋亡。我们将结合药理学（白藜芦醇/葡萄素）和遗传方法来治疗。我们将使用我们现有的 Sirt3-EE 小鼠和我们将开发的肺上皮细胞特异性 Sirt3 小鼠。 评估 SIRT3 是否通过阻止 AEC 线粒体蛋白乙酰化（OGG1 和 ACO-2）、ACO-2 耗竭、mtDNA 损伤和细胞凋亡来减轻石棉和博莱霉素诱导的肺纤维化 (3) 确定线粒体 hOGG1-EE 是否会减轻肺纤维化。使用过度表达的 MLE-12 细胞进行的体外研究显示，SIRT3 缺乏会产生有害影响。 mt-Ogg1 和 SIRT3 耗尽将用于评估石棉诱导的线粒体蛋白乙酰化（总、OGG1、ACO-2 和 MnSOD）、ACO-2 水平、mtDNA 损伤和内在细胞凋亡。我们还将使用线粒体 hOgg1-。 SIRT3 沉默缺失和存在的 EE 小鼠以确定 mt-hOGG1-EE 是否减轻肺纤维化（石棉/博莱霉素）通过保留 AEC ACO-2 水平和 mtDNA，同时减少细胞凋亡 创新：这些研究将阐明 SIRT3 在维持 AT2 细胞 mtDNA 完整性方面的重要性，这对于防止 AT2 细胞凋亡和肺纤维化至关重要。对肺纤维化的了解可能对退伍军人群体中存在的更常见的肺部疾病（即 IPF 和肺癌）产生更广泛的影响，需要有效的治疗。