Quantification of the Metabolic State in Cell-Model of Parkinson’s Disease by Fluorescence Lifetime Imaging Microscopy

Chakraborty, Sandeep; Nian, Fang; Tsai, Jin Wu; Karmenyan, Artashes; Chiou, Arthur

doi:10.1038/srep19145

Cited by 60 publications

(49 citation statements)

References 59 publications

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“…Importantly, FLIM measurements of NAD(P)H also enable investigation of the cellular metabolic phenotype. Indeed, empirical evidence has associated a shorter average lifetime, and a higher contribution of the short lifetime component, to a more glycolytic phenotype (Bird et al, ; Chakraborty, Nian, Tsai, Karmenyan, & Chiou, ; Schneckenburger, Wagner, Weber, Strauss, & Sailer, ; Skala et al, ). Intriguingly, we observed a higher average lifetime in TgAD neurons compared to WT (+7%, p = 0.006, Figure f,g), as well as a higher proportion of NAD(P)H molecules in the bound state (+10%, p = 0.003, Figure h,i), indicating that TgAD neurons were, proportionally, more aerobic compared to WT, despite lower mitochondrial NAD(P)H levels.…”

Section: Resultsmentioning

confidence: 99%

Systems biology identifies preserved integrity but impaired metabolism of mitochondria due to a glycolytic defect in Alzheimer's disease neurons

et al. 2019

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Mitochondrial dysfunction is implicated in most neurodegenerative diseases, including Alzheimer's disease (AD). We here combined experimental and computational approaches to investigate mitochondrial health and bioenergetic function in neurons from a double transgenic animal model of AD (PS2APP/B6.152H). Experiments in primary cortical neurons demonstrated that AD neurons had reduced mitochondrial respiratory capacity. Interestingly, the computational model predicted that this mitochondrial bioenergetic phenotype could not be explained by any defect in the mitochondrial respiratory chain (RC), but could be closely resembled by a simulated impairment in the mitochondrial NADH flux. Further computational analysis predicted that such an impairment would reduce levels of mitochondrial NADH, both in the resting state and following pharmacological manipulation of the RC. To validate these predictions, we utilized fluorescence lifetime imaging microscopy (FLIM) and autofluorescence imaging and confirmed that transgenic AD neurons had reduced mitochondrial NAD(P)H levels at rest, and impaired power of mitochondrial NAD(P)H production. Of note, FLIM measurements also highlighted reduced cytosolic NAD(P)H in these cells, and extracellular acidification experiments showed an impaired glycolytic flux. The impaired glycolytic flux was identified to be responsible for the observed mitochondrial hypometabolism, since bypassing glycolysis with pyruvate restored mitochondrial health. This study highlights the benefits of a systems biology approach when investigating complex, nonintuitive molecular processes such as mitochondrial bioenergetics, and indicates that primary cortical neurons from a transgenic AD model have reduced glycolytic flux, leading to reduced cytosolic and mitochondrial NAD(P)H and reduced mitochondrial respiratory capacity.

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Section: Resultsmentioning

confidence: 99%

Systems biology identifies preserved integrity but impaired metabolism of mitochondria due to a glycolytic defect in Alzheimer's disease neurons

et al. 2019

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“…The assessment of the redox ratio of NADH/ FAD + was introduced more than 40 years ago and was used multiple times for estimating cellular energy metabolism (73). In this regard, FAD + is the most commonly used molecule, as the oxidized form exhibits autofluorescence.…”

Section: Nadh/fad + Imagingmentioning

confidence: 99%

“…In this regard, FAD + is the most commonly used molecule, as the oxidized form exhibits autofluorescence. The assessment of the redox ratio of NADH/ FAD + was introduced more than 40 years ago and was used multiple times for estimating cellular energy metabolism (73). Similar to NADH autofluorescence alone, the redox ratio can be associated qualitatively with metabolism after alteration of the mitochondrial respiratory system (74).…”

Section: Nadh/fad + Imagingmentioning

confidence: 99%

NADH Autofluorescence—A Marker on its Way to Boost Bioenergetic Research

et al. 2018

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More than 60 years ago, the idea was introduced that NADH autofluorescence could be used as a marker of cellular redox state and indirectly also of cellular energy metabolism. Fluorescence lifetime imaging microscopy of NADH autofluorescence offers a marker-free readout of the mitochondrial function of cells in their natural microenvironment and allows different pools of NADH to be distinguished within a cell. Despite its many advantages in terms of spatial resolution and in vivo applicability, this technique still requires improvement in order to be fully useful in bioenergetics research. In the present review, we give a summary of technical and biological challenges that have so far limited the spread of this powerful technology. To help overcome these challenges, we provide a comprehensible overview of biological applications of NADH imaging, along with a detailed summary of valid imaging approaches that may be used to tackle many biological questions. This review is meant to provide all scientists interested in bioenergetics with support on how to embed successfully NADH imaging in their research.

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“…Cofactors NADH and FAD which are present in the cell in an oxidized (NAD + , FAD) or reduced form (NADN, FADN 2 ) play the role of electron donors and acceptors in reactions of ATP formation [1][2][3][4][5][6][7].…”

Section: Cofactors Nadn and Fad And Their Role In Cellular Energy Metmentioning

confidence: 99%

Metabolic Imaging in the Study of Oncological Processes (Review)

Lukina¹,

Shirmanova²,

Sergeeva³

et al. 2016

Sovrem Tehnol Med

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There have been reviewed the main approaches to the study of energy metabolism in cancer cells, based on fluorescent imaging of NADH and FAD cofactors. The term "metabolic imaging" covers a range of modern fluorescent techniques for detecting NADH and FAD according to fluorescence intensity and/or lifetime. These cofactors play an important role in energy metabolism reactions acting as electron carriers and, being fluorescent, serve as a basis for metabolic process analysis in living cells and tissues without the use of additional coloring agents. Particular attention is paid to metabolic changes associated with carcinogenesis. Numerous examples of metabolic imaging application in cell cultures in vitro, animal and human tumors in vivo, as well as in patients' tumor biopsy samples have demonstrated its being highly demanded for biomedical research in the area of oncology.Key words: metabolic imaging; energy metabolism; fluorescence lifetime; redox ratio; NADH; FAD; oxidative phosphorylation; glycolysis; tumor cells.For contacts: Mariya M. lukina, e-mail: kuznetsova.m.m@yandex.ru Cells need energy to maintain homeostasis. All processes of cell activity, such as generating concentration gradients, cytoskeleton movement, DNA repair, transcription, translation and vesicular transport, are energy-dependent. Energy metabolism of normal cells is known to differ significantly from cell metabolism in case of a disease. Therefore, the metabolic status can serve as an indicator for diagnosis and visualization of response to pathological process treatment. Taking into account high incidence of oncological diseases, assessment of metabolic phenotype of tumor cells is particularly urgent.Development of optical visualization technologies has provided the possibility of noninvasive analysis of metabolic cofactors NADH and FAD in living tumor cells at high spatial resolution (up to several hundred Metabolic Imaging in the Study of Oncological Process nanometers) without using additional coloring agents and with no significant effect on biochemical and physiological condition of cells. The term "metabolic imaging" covers a range of modern fluorescent techniques which allow visualizing NADH and FAD according to their fluorescence intensity and/or lifetime.The present review characterizes the properties of energy metabolism in cancer cells, describes the two key approaches to the assessment of metabolic status: analysis of cofactor fluorescence intensity ratio (redox ratio) and their fluorescence lifetime. There have been presented numerous examples of metabolic imaging application in cell cultures in vitro, animal and human tumors in vivo, as well as in patients' tumor biopsy samples.

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Quantification of the Metabolic State in Cell-Model of Parkinson’s Disease by Fluorescence Lifetime Imaging Microscopy

Cited by 60 publications

References 59 publications

Systems biology identifies preserved integrity but impaired metabolism of mitochondria due to a glycolytic defect in Alzheimer's disease neurons

Systems biology identifies preserved integrity but impaired metabolism of mitochondria due to a glycolytic defect in Alzheimer's disease neurons

NADH Autofluorescence—A Marker on its Way to Boost Bioenergetic Research

Metabolic Imaging in the Study of Oncological Processes (Review)

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