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）： 石棉会导致石棉（由于石棉引起的肺纤维化）和恶性肿瘤（肺癌和间皮瘤），这是通过未完全阐明的机制。肺泡上皮细胞（AEC）损伤和修复的程度是毒素（例如石棉）的纤维化潜力的关键决定因素。 SIRTUIN 3（SIRT3）是主要线粒体脱纤酶，调节线粒体代谢，从而将线粒体活性氧（ROS）排毒。我们已经表明，线粒体ROS介导石棉诱导的AEC DNA损伤和凋亡通过p53-和线粒体调节的死亡途径，以及一种新的机制，通过这种机制，线粒体人类8-氧气糖苷-DNA-DNA糖基化合酶1（MT-HOGGG1）通过玫瑰花质量刺激了ACOIRASER，可以保存玫瑰花孔。 （ACO-2），进而防止线粒体DNA（mtDNA）损伤。与野生型（WT）小鼠相比，我们表明OGG1 - / - 小鼠的石棉诱导的肺纤维化增加了，部分原因是肺泡II型（AT2）细胞凋亡因ACO-2水平降低和MTDNA损伤增加而增加。 SIRT3控制了包括OGG1，ACO-2等线粒体蛋白的功能，可以在神经细胞中氧化应激的情况下增强mtDNA修复。我们的初步研究表明，石棉降低了AEC SIRT3表达。 SIRT3缺乏症会促进石棉诱导的AEC MTDNA损伤，凋亡和肺纤维化。 SIRT3强制表达（EE）减弱了氧化物诱导的AEC ACO-2部署，mtDNA损伤和凋亡。我们认为AEC mtDNA是通过SIRT3调节机制暴露于石棉后整合细胞存活 /死亡信号的关键目标。假设：SIRT3通过保留MT-OGG1/ ACO-2功能来减轻石棉诱导的AEC MTDNA损伤，并降低线粒体功能障碍对于限制凋亡和肺纤维化很重要。我们未来4年将检查的具体目标包括：（1）确定SIRT3缺乏症是否促进了由于OGG1和/或ACO-2乙酰化的改变，导致指导蛋白质功能的AEC MTDNA损伤和内在凋亡。我们将评估SIRT3缺乏对OGG1和ACO-2，mtDNA损伤和凋亡的影响。 SIRT3 - / - 和肺上皮细胞特异性SIRT3 - / - 小鼠将用于评估SIRT3缺乏症是否增加了石棉和白霉素诱导的AEC MNSOD，OGG1和ACO-2乙烯酯，mtDNA损伤，凋亡，凋亡和肺纤维化。 （2）确定SIRT3强制表达（EE）是否可以防止石棉诱导的AEC线粒体蛋白乙酰化（总，OGG1，ACO-2，MNSOD），MTDNA损伤和凋亡因OGG1 / ACO-2的改变而产生的凋亡。我们将在体外使用药物（白藜芦醇 /葡萄蛋白）和遗传方法的组合。我们将使用我们可用的SIRT3-EE小鼠和我们将开发的肺上皮细胞特异性SIRT3小鼠 确定SIRT3是否通过防止AEC线粒体蛋白乙酰化（OGG1和ACO-2），ACO-2部署，mtDNA损伤和凋亡来减轻石棉和博来霉素诱导的肺纤维化。 （3）确定SIRT3是否删除了SIRT3缺乏症的有害影响，如石棉暴露后发生的那样。使用过表达MT-OGG1和SIRT3耗尽的MLE-12细胞的体外研究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损伤和内在凋亡。我们还将在不存在和存在SIRT3沉默的情况下使用线粒体HOGG1-EE小鼠，以确定MT-HOGG1-EE是否通过保持AEC ACO-2水平和MTDNA水平来减少肺纤维化（石棉 /博氏菌素）是否会减轻肺纤维化（石棉 /博氏菌素）。创新：这些研究将阐明SIRT3在维持AT2细胞MTDNA完整性方面对防止AT2细胞凋亡和肺纤维化至关重要的重要性。提出的研究将提高我们对肺纤维化的理解，这可能对需要有效治疗的资深人群中存在的更常见的肺部疾病（即IPF和肺癌）具有更广泛的影响。