MtDNA damage as a biomarker for environmental mitochondrial toxicity

线粒体 DNA 损伤作为环境线粒体毒性的生物标志物

• 批准号: 8501468
• 负责人: J Timothy Greenamyre
• 金额: $ 33.41万
• 依托单位: UNIVERSITY OF PITTSBURGH AT PITTSBURGH
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-09-19 至 2016-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8501468
• 关键词: Acute; Affect; Bacterial Toxins; Biological; Biological Assay; Biological Markers; Blood; Brain; Brain region; Cell Death; Cessation of life; Clinical; Collection; Complex; DNA; DNA Damage; Data; Defect; Degenerative Disorder; Development; Disease; Dose; Drug Kinetics; Drug or chemical Tissue Distribution; Electron Transport; Environmental Exposure; Enzyme Inhibition; Epidemiologic Studies; Excretory function; Exposure to; Fishes; Genetic; Heterogeneity; Home environment; Human; Impairment; Insecticides; Lead; Membrane; Metabolism; Metals; Mitochondria; Mitochondrial DNA; Mitochondrial Diseases; Mitochondrial Proteins; Modeling; Molecular; Muscle; Mutation; Mycotoxins; Nerve Degeneration; Nuclear; Parkinson Disease; Peripheral; Pesticides; Phenotype; Physiological; Property; Rattus; Resistance; Risk Factors; Rotenone; Sampling; Skeletal Muscle; Solvents; Substantia nigra structure; Symptoms; Syndrome; Testing; Time; Tissues; Toxic Environmental Substances; Toxic effect; Toxin; Vegetables; base; clinical phenotype; disease diagnosis; dopamine system; genetic association; inhibitor/antagonist; interest; killings; mitochondrial dysfunction; pre-clinical; response; sample collection

Demonstrating or verifying a current or past exposure to an environmental mitochondrial toxin is extraordinarily difficult. For any given toxin, tissue distribution and pharmacokinetics of the toxin may be unknown. Low-level exposure may not produce a clinical phenotype, and depending on the assay chosen, functional mitochondrial impairment or enzyme inhibition may not persist beyond the acute exposure (or metabolism/excretion of the toxin). Thus, there is a pressing need to develop a biomarker for exposure to environmental mitochondrial inhibitors that is (i) sensitive, (i) at least semi- quantitative, (iii) enduring after toxin exposure has ceased, (iv) stable after specimen collection, and (v) highly reproducible. Another problem is that after exposure to certain mitochondrial toxins, some tissues are selectively vulnerable to damage while others are resistant. For example, when rats are exposed to rotenone chronically, they develop selective degeneration of the nigrostriatal dopamine system similar to Parkinson disease. The molecular and physiological basis for such heterogeneity in biological response is obscure and no biomarker of selective vulnerability exists. Our preliminary data suggest that mtDNA damage in blood or skeletal muscle may provide a biomarker of past or ongoing toxin exposure, and nuclear DNA (nDNA) damage may be a preclinical biomarker of selective vulnerability. For these studies, we will use an extremely sensitive PCR-based assay of DNA damage (both mtDNA & nDNA) that simultaneously allows assessment of multiple forms of damage, and further allows assessment of mtDNA and nDNA damage in the same samples, without a need for mitochondrial isolation. The Specific Aims of this proposal are: 1. (a) Determine how soon after rotenone exposure mtDNA damage can be detected in blood and muscle. (b) Determine the duration, after a single exposure, that mtDNA damage can be detected. 2. Determine the minimal level of complex I inhibition that is required to cause detectable mtDNA damage. 3. Determine whether nuclear DNA (nDNA) damage is a marker of tissues that are selectively vulnerable to mitochondrial toxin-induced degeneration.!! 4. (a) Determine whether peripheral mtDNA damage is a common feature of systemically active complex I inhibitors. (b) Determine whether mtDNA damage is a common feature of other ETC inhibitors, including those acting at complexes II-IV. Preliminary results suggest our assay will provide a biomarker that is relatively simple, extremely sensitive, quantitative, enduring after exposure has ceased, stable after collection, and highly reproducible.

证明或验证当前或过去暴露于环境线粒体毒素是 非常困难。对于任何给定的毒素，毒素的组织分布和药代动力学可能 是未知的。低级暴露可能不会产生临床表型，并取决于测定 选择的，功能性线粒体损伤或酶抑制可能不会持续到急性之外 暴露（或毒素的代谢/排泄）。因此，迫切需要开发生物标志物 为了暴露于（i）敏感的环境线粒体抑制剂，（i）至少半 定量，（iii）在毒素暴露后持续持续，（iv）标本收集后稳定，（v） 高度可重现。另一个问题是，暴露于某些线粒体毒素后，有些 组织有选择地遭受损害，而其他组织则具有抗性。例如，当老鼠是 他们长期暴露于鱼藤酮，形成斑纹纹状体多巴胺的选择性变性 类似于帕金森氏病的系统。这种异质性的分子和生理基础 生物反应晦涩难懂，没有选择性脆弱性的生物标志物。我们的初步 数据表明，血液或骨骼肌中的mtDNA损害可能会提供生物标志物 过去或持续的毒素暴露，核DNA（NDNA）损害可能是 选择性脆弱性的临床前生物标志物。对于这些研究，我们将使用极端 基于敏感的PCR基于DNA损伤的测定（MTDNA和NDNA），同时允许 评估多种形式的损害，并进一步评估mtDNA和NDNA损伤 在同一样品中，无需线粒体隔离。该提案的具体目的是： 1。（a）确定紫红酮暴露后多久可以在血液和 肌肉。 （b）确定单次暴露后的持续时间，可以检测到mtDNA损伤。 2。确定引起可检测的mtDNA所需的复合I抑制的最小水平 损害。 3。确定核DNA（NDNA）损伤是否是有选择性的组织的标记 容易受到线粒体毒素诱导的变性的影响。 4。（a）确定外围mtDNA损伤是否是系统性活动的常见特征 复杂的I抑制剂。 （b）确定mtDNA损伤是否是其他损害的常见特征 抑制剂，包括作用于复合物II-IV的抑制剂。 初步结果表明我们的测定将提供一个相对简单的生物标志物 敏感，定量，暴露后持久停止，收集后稳定，高度 可再现。