Mitochondrial DNA Integrity and Endothelial Free Radical Stress

线粒体 DNA 完整性和内皮自由基应激

• 批准号: 7217672
• 负责人: MARK N GILLESPIE
• 金额: $ 29.99万
• 依托单位: UNIVERSITY OF SOUTH ALABAMA
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-08-01 至 2011-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7217672
• 关键词: DNA damage; DNA repair; apoptosis; enzyme mechanism; free radicals; microcirculation; mitochondrial DNA; oxidative stress; pulmonary artery; pulmonary circulation; vascular endothelium

In several important lung diseases, including emphysema, vascular remodeling after ARDS, and possibly primary pulmonary hypertension, there is a prominent cytotoxic response of pulmonary microvascular endothelial cells (MV ECs) leading to a diminution in capillary density. While there is no doubt that reactive oxygen species play an important role in this response, the specific target(s) of ROS that serve as a sentinel molecule - triggering cell death when the oxidant stress is so severe as to preclude effective recovery or threaten the organism with mutation - is not known. In this regard, an intriguing target of ROS is mitochondrial (mt) DNA. The mitochondrial genome is at least 30-fold more sensitive to oxidative damage than nuclear DNA, and our work during the initial funding period supports the hypothesis that oxidative mtDNA damage is a proximate trigger for lung EC death. If this hypothesis is valid, then mtDNA repair pathways could emerge as a new target for intervention in oxidant-induced MV EC death and capillary rarefaction. However, there is a stark lack of the information about the details of mtDNA repair in MV ECs and other cells. For example, while it is suspected that the base excision repair mechanism is the dominant pathway defending the mitochondrial genome from oxidative damage, the presence of other DNA repair pathway components suggests that a more complicated repair paradigm could be operative. In addition, neither the identities of the enzymes participating in mitochondrial base excision repair nor the rate limiting determinants are known. Against this background, the Aims of this proposal are to: (1) Identify the dominant pathway repairing oxidative damage to the mitochondrial genome in MV ECs; (2) Determine the rate-limiting functional steps in mtDNA repair; and, (3) Establish the critical operational enzymes repairing mtDNA in MV ECs. Collectively, these studies will provide the first detailed understanding of pathways defending the mitochondrial genome in this important lung cell population and determine the suitability of mtDNA repair enzymes to serve as isolated targets for intervention. Importantly, the outcome of these studies also will set the stage for pre-clinical, translational experiments on the ability of augmented mtDNA repair to suppress capillary rarefaction in relevant animal models.

在几种重要的肺部疾病中，包括肺气肿、ARDS 后的血管重塑以及 可能是原发性肺动脉高压，肺动脉有明显的细胞毒性反应 微血管内皮细胞（MV EC）导致毛细血管密度减少。虽然毫无疑问 活性氧在这种反应中发挥着重要作用，ROS 的具体目标 作为哨兵分子——当氧化应激严重到无法有效发挥作用时，就会引发细胞死亡 恢复或威胁生物体的突变 - 尚不清楚。在这方面，ROS 的一个有趣的目标是 线粒体 (mt) DNA。线粒体基因组对氧化损伤的敏感性至少高出 30 倍 比核 DNA 更重要，我们在最初资助期间的工作支持了这样的假设：氧化 mtDNA 损伤是肺 EC 死亡的直接触发因素。如果这个假设成立，那么线粒体DNA修复 途径可能成为干预氧化剂诱导的 MV EC 死亡和毛细血管的新目标 稀疏。然而，关于 MV EC 中 mtDNA 修复细节的信息严重缺乏 和其他细胞。例如，虽然怀疑碱基切除修复机制是主导 保护线粒体基因组免受氧化损伤和其他 DNA 修复的存在的途径 通路成分表明更复杂的修复范例可能是可行的。此外， 既不是参与线粒体碱基切除修复的酶的身份，也不是速率限制 决定因素是已知的。在此背景下，本提案的目的是： (1) 确定主导地位 修复 MV EC 中线粒体基因组氧化损伤的途径； (2) 确定限速 线粒体DNA修复的功能步骤； (3) 建立修复 MV 中 mtDNA 的关键操作酶 EC。总的来说，这些研究将首次详细了解保护病毒的途径 线粒体基因组在这个重要的肺细胞群体中的作用并确定线粒体DNA修复的适用性 酶作为干预的孤立目标。重要的是，这些研究的结果也将确定 增强 mtDNA 修复抑制能力的临床前转化实验阶段 相关动物模型中毛细血管稀疏。