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Aged rat heart mitochondrial dysfunction: proteome and lipidome dynamics

老年大鼠心脏线粒体功能障碍：蛋白质组和脂质组动力学

基本信息

批准号：
8969074
负责人：
Charles Leslie Hoppel
金额：
$ 23.78万
依托单位：
CASE WESTERN RESERVE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2015
资助国家：
美国
起止时间：
2015-09-30 至 2017-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8969074
关键词：
Accounting Acetylcarnitine Address Adult Affect Age Aging Anabolism Animals Binding Biochemical Cardiac Cardiolipins Cardiovascular Diseases Cardiovascular system Cell membrane Complex Cytochromes b Data Defect Deuterium Oxide Development Dinitrophenols Docosahexaenoic Acids Elderly Electron Transport Electron Transport Complex III Energy Metabolism Environment Essential Fatty Acids Fatty Acids Fostering Glues Glycerol Goals Heart Impairment Individual Injury Inner mitochondrial membrane Kinetics Knowledge Label Left Life Linoleic Acids Lipids Measures Membrane Metabolic Methods Minor Mitochondria Mitochondrial Proteins Modeling Molecular Morbidity - disease rate Myofibrils Organelles Oxidative Phosphorylation Phospholipids Production Proteins Proteome Public Health Publishing Rattus Replacement Therapy Reporting Research Respiration Risk Factors Site Structure System Techniques Testing Vertebral column Work age effect age related aged antioxidant therapy base complex IV heart metabolism in vivo inhibitor/antagonist insight mitochondrial dysfunction mortality novel novel strategies prevent public health relevance respiratory restoration tool

项目摘要

DESCRIPTION (provided by applicant): Mitochondrial function deteriorates with aging. We have shown that aging is associated with decreased mitochondrial protein, mitochondrial oxidative phosphorylation, complex III (CIII) and IV activity, and is associated with decrease in major cardiolipin species [CL(C18:2)4] and increase in docosahexaenoic acid containing CL [CL(C18:2)3, (C22:6)1]. The activity of complex IV is restored in the presence of phospholipids. These age-associated changes are observed in interfibrillar (IFM) mitochondria, but not in subsarcolemmal mitochondria (SSM). Because the activity of mitochondrial electron transport chain complexes is profoundly affected by the lipid environment of the mitochondrial inner membrane, specifically CL, we proposed that there is increased CL turnover/remodeling. To experimentally test our hypothesis we will use a novel heavy water (2H2O) metabolic labeling technique we have developed for a global lipidome and proteome turnover studies in free living animals. In Aim 1 we determine the effect of aging on phospholipid kinetics in SSM and IFM from 6 and 24 months old rats with particular focus on the turnover of the distinctive CL species [CL(C18:2)3, (C22:6)1] specific for mitochondria of aged rats. The glycerol moiety of CL will be used to assess CL biosynthesis and the turnover of minor nonessential fatty acids in the 6 month and docosahexaenoic acid in the 24 month old animals. Linoleic acid, an essential fatty acid, will not be labeled, which has been a problem in the two published studies. In contrast, we propose to use the glycerol moiety, which will be labeled and is the backbone while the fatty acids are part of the remodeling. In Aim 2 we determine the turnover and remodeling of CL associated with free CIII and CIII incorporated into supercomplexes in IFM in comparison to SSM, which are unaffected by aging. Is CL kinetics in cardiac IFM from aged rats restored to that observed in SSM and to IFM for 6 month old animals following restoration of complex III activity with acetyl-carnitine? Does the turnover of CIII protein, in specific cytochrome b, increase with restoration of CIII activity and of mitochondrial oxidative phosphorylation in IFM? The goal of this R21 is to collect preliminary data relevant to the mechanism of age-associated decreased mitochondrial function by determining the turnover of mitochondrial phospholipids and CL in specific using a novel heavy water (2H2O) metabolic labeling technique we have developed for a global proteome turnover studies in free living animals. Application of this novel approach to assess the effect of aging on mitochondrial function will provide valuable insight into the molecular mechanism of cardiac aging. The understanding gained from our proposed work will foster the development of strategies to counter the negative impact of aging on cardiac metabolism and to delay or slow the rate of cardiovascular mortality and morbidity.

描述(申请人提供)：线粒体功能随年龄增长而恶化。我们发现，衰老与线粒体蛋白、线粒体氧化磷酸化、复合体III(CIII)和IV的活性降低有关，与主要心磷脂物种[CL(C18：2)4]的减少和含有CL[CL(C18：2)3，(C22：6)1]的二十二碳六烯酸的增加有关。在磷脂存在的情况下，络合物IV的活性恢复。这些与年龄相关的变化在纤维间线粒体(IFM)中观察到，但在肌膜下线粒体(SSM)中没有观察到。由于线粒体电子传递链复合体的活性受到线粒体内膜脂环境的深刻影响，特别是CL，我们认为存在CL周转/重塑的增加。为了在实验上验证我们的假设，我们将使用一种新的重水(2H2O)代谢标记技术，我们为自由生活动物中的全球脂体和蛋白质组周转研究开发了一种新的技术。在目的1中，我们研究了衰老对6月龄和24月龄大鼠SSM和IFM磷脂动力学的影响，重点研究了老年大鼠线粒体特有的CL[CL(C18：2)3，(C22：6)1]的更新。CL的甘油部分将被用来评估CL的生物合成和6月龄时少量非必需脂肪酸的周转以及24月龄动物的二十二碳六烯酸。亚油酸，一种基本的脂肪酸，将不会被贴上标签，这在两项已发表的研究中一直是一个问题。相反，我们建议使用甘油部分，它将被标记为骨架，而脂肪酸是重塑的一部分。在目标2中，我们确定了与不受老化影响的SSM相比，IFM中与游离CIII和CIII结合在超复合体中的CL的周转和重塑。在乙酰肉碱恢复复合体III活性后，老年大鼠心脏IFM的CL动力学是否恢复到SSM观察到的，6月龄动物的IFM恢复到IFM？在IFM中，CIII蛋白在特定细胞色素b中的周转是否随着CIII活性和线粒体氧化磷酸化的恢复而增加？R21的目的是通过利用我们为自由生活动物的全球蛋白质组周转研究开发的一种新的重水(2H2O)代谢标记技术，通过具体测定线粒体磷脂和CL的周转来收集与年龄相关的线粒体功能下降机制相关的初步数据。应用这一新的方法来评估衰老对线粒体功能的影响，将为了解心脏衰老的分子机制提供有价值的见解。从我们拟议的工作中获得的理解将促进制定战略，以对抗衰老对心脏代谢的负面影响，并延缓或减缓心血管死亡率和发病率。