Logan Angela, Cochemé Helena M, Li Pun Pamela Boon, Apostolova Nadezda, Smith Robin A J, Larsen Lesley, Larsen David S, James Andrew M, Fearnley Ian M, Rogatti Sebastian, Prime Tracy A, Finichiu Peter G, Dare Anna, Chouchani Edward T, Pell Victoria R, Methner Carmen, Quin Caroline, McQuaker Stephen J, Krieg Thomas, Hartley Richard C, Murphy Michael P
MRC Mitochondrial Biology Unit, Hills Road, Cambridge CB2 0XY, UK.
Biochim Biophys Acta. 2014 Feb;1840(2):923-30. doi: 10.1016/j.bbagen.2013.05.026. Epub 2013 May 30.
The ability to measure the concentrations of small damaging and signalling molecules such as reactive oxygen species (ROS) in vivo is essential to understanding their biological roles. While a range of methods can be applied to in vitro systems, measuring the levels and relative changes in reactive species in vivo is challenging.
One approach towards achieving this goal is the use of exomarkers. In this, exogenous probe compounds are administered to the intact organism and are then transformed by the reactive molecules in vivo to produce a diagnostic exomarker. The exomarker and the precursor probe can be analysed ex vivo to infer the identity and amounts of the reactive species present in vivo. This is akin to the measurement of biomarkers produced by the interaction of reactive species with endogenous biomolecules.
Our laboratories have developed mitochondria-targeted probes that generate exomarkers that can be analysed ex vivo by mass spectrometry to assess levels of reactive species within mitochondria in vivo. We have used one of these compounds, MitoB, to infer the levels of mitochondrial hydrogen peroxide within flies and mice. Here we describe the development of MitoB and expand on this example to discuss how better probes and exomarkers can be developed. This article is part of a Special Issue entitled Current methods to study reactive oxygen species - pros and cons and biophysics of membrane proteins. Guest Editor: Christine Winterbourn.
在体内测量诸如活性氧（ROS）等小的损伤性和信号分子的浓度，对于理解它们的生物学作用至关重要。虽然一系列方法可应用于体外系统，但在体内测量活性物质的水平及其相对变化具有挑战性。
实现这一目标的一种方法是使用外源性标志物。在这种方法中，将外源性探针化合物施用于完整生物体，然后它们在体内被活性分子转化以产生诊断性外源性标志物。外源性标志物和前体探针可在体外进行分析，以推断体内存在的活性物质的种类和数量。这类似于测量活性物质与内源性生物分子相互作用产生的生物标志物。
我们实验室已开发出线粒体靶向探针，这些探针能产生可通过质谱在体外进行分析的外源性标志物，以评估体内线粒体中活性物质的水平。我们已使用其中一种化合物MitoB来推断果蝇和小鼠体内线粒体过氧化氢的水平。在此，我们描述MitoB的开发过程，并以此为例展开讨论如何开发更好的探针和外源性标志物。本文是名为“研究活性氧的当前方法——利弊及膜蛋白生物物理学”的特刊的一部分。客座编辑：克里斯汀·温特伯恩。
