Assessment of Mitochondrial Metabolic Oxidative State in Living Cardiomyocytes with Cardiomyocyte Autofluorescence_West China Medical Journal

Authors：

1.the Department of Cardiovascular Surgery, Beijing University Shenzhen Hospital, Shenzhen Guangdong 518036, China; 2.the Department of Pediatrics, Research Centre of CHU SainteJustine, University of Montreal, Montreal, QC. Canada, H3T 1C5; 3.the Department of Biophotonics, International Laser Centre, Bratislava, Slovakia 81219;

Corresponding author：

Keywords：

烟酰胺腺嘌呤(磷酸)二核苷酸; 自发荧光; 光谱分辨的荧光寿命; 线粒体; 存活心肌细胞

Video：

Export PDF Favorites Scan Get Citation

Abstract Full text Figures/Tables Video References Cited by

目的:应用心肌自发荧光(AF)研究心肌线粒体氧化代谢状态,监测线粒体功能改变的早期信号。方法:烟酰胺腺嘌呤(磷酸)二核苷酸［NAD(P)H］作为荧光探针,用光谱分辨的时间相关单光子计数(TCSPC)记录375nm紫外激光激发的心肌AF光谱和荧光寿命,测试影响线粒体呼吸时AF动态衰减。结果:在420~560nm光谱区域,至少需用3个荧光寿命池0.4~0.7ns,1.2~1.9ns和8.0~13.0ns描述细胞AF。线粒体呼吸阻断剂鱼藤酮可显著增加AF强度,缩短平均荧光寿命。氧化磷酸化解偶联剂二硝基酚可显著降低AF强度,在520nm处增宽荧光光谱,延长平均荧光寿命。这些结果和NADH荧光动力学离体实验(in vitro)有可比性。结论:光谱分辨的荧光寿命技术测定心肌NAD(P)H荧光有很好的重复性,在细胞水平上增加了心肌氧化代谢或线粒体功能障碍的知识,为临床诊断和治疗线粒体功能障碍开拓了新视野。

Citation： CHENG Ying,REN Mingming,LIU Yunqi,et al.. Assessment of Mitochondrial Metabolic Oxidative State in Living Cardiomyocytes with Cardiomyocyte Autofluorescence. West China Medical Journal, 2009, 24(7): 1767-1771. doi: Copy

1.
2.	CHANCE B, SCHOENER B, OSHINO R, et al. Oxidationreduction ratio studies of mitochondria in freezetrapped samples. NADH and flavoprotein fluorescence signals[J]. J Biol Chem,1979,254(11):4764-4771..
3.	BLINOVA K, CARROLL S, BOSE S, et al. Distribution of mitochondrial NADH fluorescence lifetimes: steadystate kinetics of matrix NADH interactions[J]. Biochemistry,2005,44(7):2585-2594..
4.	ENG J, LYNCH R M, BALABAN R S. Nicotinamide adenine dinucleotide fluorescence spectroscopy and imaging of isolated cardiac myocytes[J]. Biophys J,1989,55(4):621-630..
5.	CHANCE B, THORELL B. Fluorescence measurements of mitochondrial pyridine nucleotide in aerobiosis and anaerobiosis[J]. Nature,1959,184:931-934..
6.	BASSIENCAPSA V, FOURON J C, COMTE B, et al. Structural, functional and metabolic remodeling of rat left ventricular myocytes in normal and in sodiumsupplemented pregnancy[J]. Cardiovasc Res,2006,69(2):423-431..
7.	CHORVAT D JR, CHORVATOVA A. Spectrally resolved timecorrelated single photon counting: a novel approach for characterization of endogenous fluorescence in isolated cardiac myocytes[J]. Eur Biophys J,2006,36:73-83..
8.	ANEBA S, CHENG Y, MATEASIK A, et al. Probing of cardiomyocyte metabolism by spectrally resolved lifetime detection of NAD(P)H fluorescence[J]. Comput Cardiol,2007,34:349-352..
9.	VISHWASRAO H D, HEIKAL A A, KASISCHKE K A, et al. Conformational dependence of intracellular NADH on metabolic state revealed by associated fluorescence anisotropy[J]. J Biol Chem,2005,280(26):25119-25126..
10.	WAKITA M, NISHIMURA G, TAMURA M. Some characteristics of the fluorescence lifetime of reduced pyridine nucleotides in isolated mitochondria, isolated hepatocytes, and perfused rat liver in situ[J]. J Biochem (Tokyo),1995,118(6):1151-1160..
11.	BRAUTIGAM C A, CHUANG J L, TOMCHICK D R, et al. Crystal structure of human dihydrolipoamide dehydrogenase: NAD+/NADH binding and the structural basis of diseasecausing mutations[J]. J Mol Biol,2005,350(3):543-552..
12.	ROMASHKO D N, MARBAN E, O’ROURKE B. Subcellular metabolic transients and mitochondrial redox waves in heart cells[J]. Proc Natl Acad Sci USA,1998,95(4):1618-1623..
13.	CHORVAT D JR, BASSIENCAPSA V, CAGALINEC M, et al. Mitochondrial autofluorescence induced by visible light in single cardiac myocytes studied by spectrally resolved confocal microscopy[J]. Laser Physics,2004,14(2):220-230..
14.	CHORVAT D JR, KIRCHNEROVA J, CAGALINEC M, et al. Spectral unmixing of flavin autofluorescence components in cardiac myocytes[J]. Biophys J,2005,89(6):L55-L57..
15.	CHORVAT D, ELZWIEI F, BASSIENCAPSA V, et al. Assessment of lowIntensity fluorescence signals in living cardiac cells using timeresolved laser spectroscopy[J]. Comput Cardiol,2007,34:353-357..
16.	SEKAR R B, PERIASAMY A. Fluorescence resonance energy transfer (FRET) microscopy imaging of live cell protein localizations[J]. J Cell Biol,2003,160(5):629-633..
17.	CHENG Y, DAHDAH N, PORIER N, et al. Spectrally and timeresolved study of NADH autofluorescence in cardiac myocytes from human biopsies[M]. Proceedings of SPIE: the International Society for Optical Engineering,2007:6771(Advanced Photon Counting Techniques II): 677104-1-67710413..

1.
2. CHANCE B, SCHOENER B, OSHINO R, et al. Oxidationreduction ratio studies of mitochondria in freezetrapped samples. NADH and flavoprotein fluorescence signals[J]. J Biol Chem,1979,254(11):4764-4771..
3. BLINOVA K, CARROLL S, BOSE S, et al. Distribution of mitochondrial NADH fluorescence lifetimes: steadystate kinetics of matrix NADH interactions[J]. Biochemistry,2005,44(7):2585-2594..
4. ENG J, LYNCH R M, BALABAN R S. Nicotinamide adenine dinucleotide fluorescence spectroscopy and imaging of isolated cardiac myocytes[J]. Biophys J,1989,55(4):621-630..
5. CHANCE B, THORELL B. Fluorescence measurements of mitochondrial pyridine nucleotide in aerobiosis and anaerobiosis[J]. Nature,1959,184:931-934..
6. BASSIENCAPSA V, FOURON J C, COMTE B, et al. Structural, functional and metabolic remodeling of rat left ventricular myocytes in normal and in sodiumsupplemented pregnancy[J]. Cardiovasc Res,2006,69(2):423-431..
7. CHORVAT D JR, CHORVATOVA A. Spectrally resolved timecorrelated single photon counting: a novel approach for characterization of endogenous fluorescence in isolated cardiac myocytes[J]. Eur Biophys J,2006,36:73-83..
8. ANEBA S, CHENG Y, MATEASIK A, et al. Probing of cardiomyocyte metabolism by spectrally resolved lifetime detection of NAD(P)H fluorescence[J]. Comput Cardiol,2007,34:349-352..
9. VISHWASRAO H D, HEIKAL A A, KASISCHKE K A, et al. Conformational dependence of intracellular NADH on metabolic state revealed by associated fluorescence anisotropy[J]. J Biol Chem,2005,280(26):25119-25126..
10. WAKITA M, NISHIMURA G, TAMURA M. Some characteristics of the fluorescence lifetime of reduced pyridine nucleotides in isolated mitochondria, isolated hepatocytes, and perfused rat liver in situ[J]. J Biochem (Tokyo),1995,118(6):1151-1160..
11. BRAUTIGAM C A, CHUANG J L, TOMCHICK D R, et al. Crystal structure of human dihydrolipoamide dehydrogenase: NAD+/NADH binding and the structural basis of diseasecausing mutations[J]. J Mol Biol,2005,350(3):543-552..
12. ROMASHKO D N, MARBAN E, O’ROURKE B. Subcellular metabolic transients and mitochondrial redox waves in heart cells[J]. Proc Natl Acad Sci USA,1998,95(4):1618-1623..
13. CHORVAT D JR, BASSIENCAPSA V, CAGALINEC M, et al. Mitochondrial autofluorescence induced by visible light in single cardiac myocytes studied by spectrally resolved confocal microscopy[J]. Laser Physics,2004,14(2):220-230..
14. CHORVAT D JR, KIRCHNEROVA J, CAGALINEC M, et al. Spectral unmixing of flavin autofluorescence components in cardiac myocytes[J]. Biophys J,2005,89(6):L55-L57..
15. CHORVAT D, ELZWIEI F, BASSIENCAPSA V, et al. Assessment of lowIntensity fluorescence signals in living cardiac cells using timeresolved laser spectroscopy[J]. Comput Cardiol,2007,34:353-357..
16. SEKAR R B, PERIASAMY A. Fluorescence resonance energy transfer (FRET) microscopy imaging of live cell protein localizations[J]. J Cell Biol,2003,160(5):629-633..
17. CHENG Y, DAHDAH N, PORIER N, et al. Spectrally and timeresolved study of NADH autofluorescence in cardiac myocytes from human biopsies[M]. Proceedings of SPIE: the International Society for Optical Engineering,2007:6771(Advanced Photon Counting Techniques II): 677104-1-67710413..

West China Medical Journal

Assessment of Mitochondrial Metabolic Oxidative State in Living Cardiomyocytes with Cardiomyocyte Autofluorescence

Abstract Full text Figures/Tables Video References Cited by

Format

Content