Developing chemical mass spectrometry probes to assess the production of reactive oxygen species in vivo

开发化学质谱探针来评估体内活性氧的产生

• 批准号: BB/I012923/1
• 负责人: Michael Murphy
• 金额: $ 3.9万
• 依托单位: MRC Centre Cambridge
• 依托单位国家: 英国
• 项目类别: Research Grant
• 财政年份: 2011
• 资助国家: 英国
• 起止时间: 2011 至 无数据
• 项目状态: 已结题
• 来源: https://gtr.ukri.org/projects?ref=BB%2FI012923%2F1
• 关键词: Developing; chemical; mass; spectrometry; probes

Within our bodies, and the bodies of all animals, the oxygen we breathe undergoes a side reaction which leads to the production of small molecules called 'reactive oxygen species' (ROS). These ROS are responsible for the oxidative damage associated with ageing. Indeed, it has been argued that they cause the ageing process itself and the conditions, such as nuerodegeneration, associated with it. ROS are also used in important signaling pathways within living organisms. However, while there is a lot of circumstantial evidence pointing to the importance of ROS within living creatures, there is also a considerable amount of uncertainty about the extent of their role. This is because ROS are very short lived and difficult to measure, so while they can be quantified in the test tube, it is currently not possible to work out how much ROS are present inside an organism. Because of this difficulty, we are often uncertain about how important ROS are in a range of important biological processes, such as ageing, disease, growth and cancer. To overcome this difficulty we will develop a new method using molecular probes that are designed to go to mitochondria, the parts of the cells that make up our bodies which are believed to be the main sources of ROS. The targeting to mitochondria relies on a simple physicochemical effect whereby lipophilic cations capable of crossing membranes accumulate in the mitochondrial matrix. Once there, the probe molecules will react with the ROS to form distinctive products. The ratio of the starting molecule to its product will then give an indication of the amount of ROS in isolated mitochondria and in the mitochondria of whole cells and animals. Similar probes are designed to remain outside cells so that ROS can be detected in the extracellular environment, particularly in the circulation. Our approach has to be very sensitive in order to assess the small changes that occur within organisms. To facilitate this, the probes are designed so that they can be detected in complex biological mixtures selectively and sensitively, even when present at very small amounts, by a technique called liquid chromatography-tandem mass spectrometry. Through the new probes' combination of targeting to particular locations in the cell or extracellular environment along with their built in feature that allows very sensitive detection by mass spectrometry, we will be able measure ROS in experimental animals and thereby work out for the first time the detailed contributions of ROS to many important biological processes, most significantly to the process of ageing itself. Antioxidants targeted to remain outside cells will also be developed as an intervention that will allow us to assess how the specific contribution of oxidative stress in the circulation and extracellular matrix affects the process of ageing as a whole.

在我们的身体和所有动物的体内，我们呼吸的氧气会发生侧反应，从而导致产生称为“活性氧”（ROS）的小分子。这些ROS负责与衰老有关的氧化损伤。确实，有人认为它们会导致与之相关的衰老过程本身和诸如nuerodegeneration之类的条件。 ROS也用于活生物体内的重要信号通路。但是，尽管有很多间接证据表明ROS在生物中的重要性，但其角色的程度也存在很大的不确定性。这是因为ROS的生命非常短且难以测量，因此尽管可以在试管中进行量化，但目前无法确定生物体中存在多少ROS。由于这种困难，我们通常不确定ROS在一系列重要的生物学过程中的重要性，例如衰老，疾病，生长和癌症。为了克服这一困难，我们将使用旨在进入线粒体的分子探针开发一种新方法，线粒体是构成我们身体的细胞部分，这些细胞构成了我们的身体，这些部分被认为是ROS的主要来源。线粒体对线粒体的靶向依赖于一种简单的物理化学效应，从而使跨膜的亲脂性阳离子积聚在线粒体基质中。到达那里后，探针分子将与ROS反应形成独特的产物。然后，起始分子与其产物的比率将表明在分离的线粒体以及整个细胞和动物的线粒体中的ROS量。类似的探针被设计为保留在细胞外部，以便在细胞外环境中，尤其是在循环中检测到ROS。我们的方法必须非常敏感，以评估生物体中发生的小变化。为了促进这一点，设计了探针，以便可以通过一种称为液相色谱倾斜质谱法的技术选择性地和敏感地在复杂的生物混合物中检测到它们。通过新的探针将靶向在细胞或细胞外环境中的特定位置的组合以及它们的内置功能，允许通过质谱进行非常敏感的检测，我们将能够测量实验动物中的ROS，从而首次在ROS对许多重要的生物学过程中详细贡献，最重要的是对老化本身的生物学过程很重要。靶向留在外部细胞的抗氧化剂也将作为一种干预措施开发，该干预措施将使我们能够评估氧化应激在循环和细胞外基质中的特定贡献如何影响整个衰老的过程。

项目成果

Ischaemic accumulation of succinate controls reperfusion injury through mitochondrial ROS.

• DOI: 10.1038/nature13909
• 发表时间: 2014-11-20
• 期刊: NATURE
• 影响因子: 64.8
• 作者: Chouchani, Edward T.;Pell, Victoria R.;Gaude, Edoardo;Aksentijevic, Dunja;Sundier, Stephanie Y.;Robb, Ellen L.;Logan, Angela;Nadtochiy, Sergiy M.;Ord, Emily N. J.;Smith, Anthony C.;Eyassu, Filmon;Shirley, Rachel;Hu, Chou-Hui;Dare, Anna J.;James, Andrew M.;Rogatti, Sebastian;Hartley, Richard C.;Eaton, Simon;Costa, Ana S. H.;Brookes, Paul S.;Davidson, Sean M.;Duchen, Michael R.;Saeb-Parsy, Kourosh;Shattock, Michael J.;Robinson, Alan J.;Work, Lorraine M.;Frezza, Christian;Krieg, Thomas;Murphy, Michael P.
• 通讯作者: Murphy, Michael P.

A mitochondria-targeted mass spectrometry probe to detect glyoxals: implications for diabetes.

• DOI: 10.1016/j.freeradbiomed.2013.11.025
• 发表时间: 2014-02
• 期刊: FREE RADICAL BIOLOGY AND MEDICINE
• 影响因子: 7.4
• 作者: Pun, Pamela Boon Li;Logan, Angela;Darley-Usmar, Victor;Chacko, Balu;Johnson, Michelle S.;Huang, Guang W.;Rogatti, Sebastian;Prime, Tracy A.;Methner, Carmen;Krieg, Thomas;Fearnley, Ian M.;Larsen, Lesley;Larsen, David S.;Menger, Katja E.;Collins, Yvonne;James, Andrew M.;Kumar, G. D. Kishore;Hartley, Richard C.;Smith, Robin A. J.;Murphy, Michael P.
• 通讯作者: Murphy, Michael P.

In vivo levels of mitochondrial hydrogen peroxide increase with age in mtDNA mutator mice.

• DOI: 10.1111/acel.12212
• 发表时间: 2014-08
• 期刊: Aging cell
• 影响因子: 7.8
• 作者: Logan A;Shabalina IG;Prime TA;Rogatti S;Kalinovich AV;Hartley RC;Budd RC;Cannon B;Murphy MP
• 通讯作者: Murphy MP

Expanding the palette of phenanthridinium cations.

• DOI: 10.1002/chem.201304241
• 发表时间: 2014-03-24
• 期刊: Chemistry (Weinheim an der Bergstrasse, Germany)
• 作者: Cairns AG;Senn HM;Murphy MP;Hartley RC
• 通讯作者: Hartley RC

Cardioprotection by S-nitrosation of a cysteine switch on mitochondrial complex I.

• DOI: 10.1038/nm.3212
• 发表时间: 2013-06
• 期刊: Nature medicine
• 影响因子: 82.9