Heart mitochondrial NOS and in vitro hypoxia/reperfusion

心脏线粒体NOS与体外缺氧/再灌注

基本信息

批准号：
7191504
负责人：
PEDRAM GHAFOURIFAR
金额：
$ 4.39万
依托单位：
OHIO STATE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2004
资助国家：
美国
起止时间：
2004-09-30 至 2007-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7191504
关键词：
aging antioxidants apoptosis cytochrome c electrodes enzyme activity free radical oxygen inhibitor /antagonist laboratory rat mitochondria myocardial ischemia /hypoxia nitration nitric oxide synthase oxidative stress peroxidation peroxynitrites reperfusion

项目摘要

DESCRIPTION (provided by applicant): 2. CARDIOVASCULAR AND CEREBROVASCULAR AGING-Nitric oxide (NO) and reactive oxygen species (ROS) play critical roles in hypoxia/reoxygenation in heart. Aged animals are more susceptible to oxidative assault during hypoxia/reoxygenation. Increased ROS, perturbed redox balance and apoptosis occur during heart hypoxia/reoxygenation. Mitochondria play a central role in oxidative damage and apoptosis. Mitochondria render one of the main cellular sources of ROS and they release key apoptogenic proteins such as cytochrome c. Recently we and others showed that mitochondria also produce NO and they possess mitochondrial NO synthase (mtNOS). NO generated by mtNOS can react with mitochondrial ROS to produce oxidizing species such as peroxynitrite. Mitochondrial peroxynitrite induces oxidative stress and releases the key mitochondrial pro-apoptosis protein, cytochrome c. The interplay between mitochondrial NO, ROS, peroxynitrite, and apoptosis components during hypoxia/reoxyqenation is not well understood. We have designed and established a novel in vitro model to study these factors with minimum cellular components that may confound the study. To mimic hypoxia we incubate isolated mitochondria in lowered oxygen concentrations, and to mimic hypoxia/reoxygenation we incubate isolated mitochondria in lowered oxygen concentrations followed by reoxygenation. Using mitochondria isolated from heart of young adult rats we found that hypoxia/reoxygenation, but not hypoxia pre se, causes the release of cytochrome c and induces oxidative stress in a reverse oxygen concentration-dependent manner. The release of cytochrome c Is prevented when mtNOS is inhibited or when mitochondria are provided exogenously with antioxidants: glutathione, vitamin C, or a vitamin E derivative peroxynitrite scavenger. We hypothesize that mtNOS is involved in hypoxia/reoxygenation-induced oxidative stress and cytochrome c release, and that mitochondria isolated from the heart of aged animals are more susceptible to hypoxia/reoxygenation-induced oxidative stress and cytochrome c release. We propose to address this hypothesis using our in vitro model. We will test heart mtNOS protein expression and activity during hypoxia/reoxygenation in young and awed rats. We will also test apoptosis, oxidative stress and peroxynitrite markers during hypoxia/reoxygenation in heart mitochondria of young and aged rats. The results of the proposed study will reveal novel insights into the molecular mechanisms underlying hypoxia/reoxygenation induced oxidative stress and apoptosis and aging.

描述（由申请人提供）：2。心血管和脑血管衰老的氧化氧化物（NO）和活性氧（ROS）在心脏中缺氧/重氧中起关键作用。老年动物在缺氧/重氧中更容易受到氧化攻击的影响。在心脏缺氧/重氧中，ROS增加，扰动的氧化还原平衡和凋亡发生。线粒体在氧化损伤和凋亡中起着核心作用。线粒体呈现ROS的主要细胞来源之一，它们释放了关键的凋亡蛋白，例如细胞色素c。最近，我们和其他人表明，线粒体也会产生NO，并且它们具有线粒体NO合酶（mTNOS）。 mtnos产生的不可能与线粒体ROS反应，产生氧化物种，例如过氧亚硝酸盐。线粒体过氧亚硝酸盐诱导氧化应激，并释放关键的线粒体促凋亡蛋白，细胞色素c。在缺氧/雷氧化过程中，线粒体NO，ROS，过氧亚硝酸盐和凋亡成分之间的相互作用尚不清楚。我们设计并建立了一种新型的体外模型，以研究可能混淆研究的最小细胞成分来研究这些因素。为了模仿缺氧，我们在降低的氧气浓度中孵育分离的线粒体，并模仿缺氧/重氧，我们在降低的氧气浓度中孵育分离的线粒体，然后再氧合。使用从年轻大鼠心脏中分离出的线粒体，我们发现缺氧/再氧化剂（而不是缺氧）会导致细胞色素C的释放，并以反氧浓度依赖的方式诱导氧化应激。当抑制MTNOS或用抗氧化剂外源性地提供线粒体时，可以预防细胞色素C：谷胱甘肽，维生素C或维生素E衍生物过氧硝酸盐清除剂。我们假设MTNO参与缺氧/重氧诱导的氧化应激和细胞色素c释放，并且从老年动物心脏中分离出的线粒体更容易受到低氧/再氧诱导的氧化应激和细胞染色体C的释放。我们建议使用我们的体外模型解决这一假设。我们将在年轻大鼠和敬畏大鼠的低氧/二氧化过程中测试心脏mTNOS蛋白的表达和活性。我们还将测试年轻大鼠和年龄大鼠心脏线粒体中缺氧/二氧化过程中缺氧/重氧期间的细胞凋亡，氧化应激和过氧亚硝酸盐标记。拟议的研究的结果将揭示对缺氧/重氧诱导的氧化应激以及凋亡和衰老的分子机制的新见解。

项目成果

期刊论文数量（11）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

mAtNOS1 induces apoptosis of human mammary adenocarcinoma cells.

DOI：
10.1016/j.lfs.2008.03.019
发表时间：
2008-05
期刊：
Life sciences
影响因子：
6.1
作者：
A. Parihar;M. Parihar;Zhonghai Chen;P. Ghafourifar
通讯作者：
A. Parihar;M. Parihar;Zhonghai Chen;P. Ghafourifar

Mammalian mitochondrial nitric oxide synthase: characterization of a novel candidate.

哺乳动物线粒体一氧化氮合酶：新候选者的表征。

DOI：
10.1016/j.febslet.2005.12.038
发表时间：
2006
期刊：
FEBS letters
影响因子：
3.5
作者：
Zemojtel,Tomasz;Kolanczyk,Mateusz;Kossler,Nadine;Stricker,Sigmar;Lurz,Rudi;Mikula,Ivan;Duchniewicz,Marlena;Schuelke,Markus;Ghafourifar,Pedram;Martasek,Pavel;Vingron,Martin;Mundlos,Stefan
通讯作者：
Mundlos,Stefan

Alpha-synuclein overexpression and aggregation exacerbates impairment of mitochondrial functions by augmenting oxidative stress in human neuroblastoma cells