Mik Egbert G, Johannes Tanja, Zuurbier Coert J, Heinen Andre, Houben-Weerts Judith H P M, Balestra Gianmarco M, Stap Jan, Beek Johan F, Ince Can
Department of Physiology, Academic Medical Center, University of Amsterdam, Amsterdam, The Netherlands.
Biophys J. 2008 Oct;95(8):3977-90. doi: 10.1529/biophysj.107.126094. Epub 2008 Jul 18.
Mitochondrial oxygen tension (mitoPO(2)) is a key parameter for cellular function, which is considered to be affected under various pathophysiological circumstances. Although many techniques for assessing in vivo oxygenation are available, no technique for measuring mitoPO(2) in vivo exists. Here we report in vivo measurement of mitoPO(2) and the recovery of mitoPO(2) histograms in rat liver by a novel optical technique under normal and pathological circumstances. The technique is based on oxygen-dependent quenching of the delayed fluorescence lifetime of protoporphyrin IX. Application of 5-aminolevulinic acid enhanced mitochondrial protoporphyrin IX levels and induced oxygen-dependent delayed fluorescence in various tissues, without affecting mitochondrial respiration. Using fluorescence microscopy, we demonstrate in isolated hepatocytes that the signal is of mitochondrial origin. The delayed fluorescence lifetime was calibrated in isolated hepatocytes and isolated perfused livers. Ultimately, the technique was applied to measure mitoPO(2) in rat liver in vivo. The results demonstrate mitoPO(2) values of approximately 30-40 mmHg. mitoPO(2) was highly sensitive to small changes in inspired oxygen concentration around atmospheric oxygen level. Ischemia-reperfusion interventions showed altered mitoPO(2) distribution, which flattened overall compared to baseline conditions. The reported technology is scalable from microscopic to macroscopic applications, and its reliance on an endogenous compound greatly enhances its potential field of applications.
线粒体氧张力（mitoPO₂）是细胞功能的关键参数，在各种病理生理情况下被认为会受到影响。尽管有许多评估体内氧合的技术，但尚无用于在体内测量mitoPO₂的技术。在此，我们报告了一种新型光学技术在正常和病理情况下对大鼠肝脏mitoPO₂的体内测量以及mitoPO₂直方图的恢复情况。该技术基于原卟啉IX延迟荧光寿命的氧依赖性猝灭。应用5-氨基乙酰丙酸可提高线粒体原卟啉IX水平，并在不影响线粒体呼吸的情况下在各种组织中诱导氧依赖性延迟荧光。使用荧光显微镜，我们在分离的肝细胞中证明该信号源自线粒体。在分离的肝细胞和分离的灌注肝脏中对延迟荧光寿命进行了校准。最终，该技术被应用于体内测量大鼠肝脏中的mitoPO₂。结果显示mitoPO₂值约为30 - 40 mmHg。mitoPO₂对大气氧水平附近吸入氧浓度的微小变化高度敏感。缺血再灌注干预显示mitoPO₂分布发生改变，与基线条件相比总体变平。所报道的技术可从微观应用扩展到宏观应用，并且其对内源性化合物的依赖极大地扩展了其潜在应用领域。
