Abstract

A time-resolved spectroscopic imaging system is built to study the fluorescence characteristics of nicotinamide adenine dinucleotide (NADH), an important metabolic coenzyme and endogenous fluorophore in cells. The system provides a unique approach to measure fluorescence signals in different cellular organelles and cytoplasm. The ratios of free over protein-bound NADH signals in cytosol and nucleus are slightly higher than those in mitochondria. The mitochondrial fluorescence contributes about 70% of overall cellular fluorescence and is not a completely dominant signal. Furthermore, NADH signals in mitochondria, cytosol, and the nucleus respond to the changes of cellular activity differently, suggesting that cytosolic and nuclear fluorescence may complicate the well-known relationship between mitochondrial fluorescence and cellular metabolism.

© 2008 Optical Society of America

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W. Ying, Antioxid. Redox. Sign. 10, 1 (2008).
[CrossRef]

M. S. Yang, D. Li, T. Lin, J. J. Zheng, W. Zheng, and J. Y. Qu, Toxicology 247, 6 (2008).
[CrossRef] [PubMed]

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M. C. Skala, K. M. Riching, D. K. Bird, A. Gendron-Fitzpatrick, J. Eichhoff, K. W. Eliceiri, P. J. Keely, and N. Ramanujam, J. Biomed. Opt. 12, 024014 (2007).
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2004

A. C. Croce, A. Ferrigno, M. Vairetti, R. Bertone, I. Freitas, and G. Bottiroli, Photochem. Photobiol. Sci. 3, 920 (2004).
[CrossRef] [PubMed]

2003

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[CrossRef] [PubMed]

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S. Huang, A. A. Heikal, and W. W. Webb, Biophys. J. 82, 2811 (2002).
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[PubMed]

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J. Beuthan, O. Minet, and G. Muller, Ann. N.Y. Acad. Sci. 838, 150 (1998).
[CrossRef] [PubMed]

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T. V. Zglinicki, C. Edwall, E. Ostlund, B. Lind, M. Nordberg, N. R. Ringertz, and J. Wroblewski, J. Cell. Sci. 103, 1073 (1992).

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J. Beuthan, O. Minet, and G. Muller, Ann. N.Y. Acad. Sci. 838, 150 (1998).
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M. C. Skala, K. M. Riching, D. K. Bird, A. Gendron-Fitzpatrick, J. Eichhoff, K. W. Eliceiri, P. J. Keely, and N. Ramanujam, J. Biomed. Opt. 12, 024014 (2007).
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C. C. Fjeld, W. T. Birdsong, and R. H. Goodman, Proc. Natl. Acad. Sci. U.S.A. 100, 9202 (2003).
[CrossRef] [PubMed]

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A. C. Croce, A. Ferrigno, M. Vairetti, R. Bertone, I. Freitas, and G. Bottiroli, Photochem. Photobiol. Sci. 3, 920 (2004).
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M. K. Campbell, Biochemistry, (Saunders College Publishing, 1995).
[PubMed]

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A. C. Croce, A. Ferrigno, M. Vairetti, R. Bertone, I. Freitas, and G. Bottiroli, Photochem. Photobiol. Sci. 3, 920 (2004).
[CrossRef] [PubMed]

Edwall, C.

T. V. Zglinicki, C. Edwall, E. Ostlund, B. Lind, M. Nordberg, N. R. Ringertz, and J. Wroblewski, J. Cell. Sci. 103, 1073 (1992).

Eichhoff, J.

M. C. Skala, K. M. Riching, D. K. Bird, A. Gendron-Fitzpatrick, J. Eichhoff, K. W. Eliceiri, P. J. Keely, and N. Ramanujam, J. Biomed. Opt. 12, 024014 (2007).
[CrossRef] [PubMed]

Eliceiri, K. W.

M. C. Skala, K. M. Riching, D. K. Bird, A. Gendron-Fitzpatrick, J. Eichhoff, K. W. Eliceiri, P. J. Keely, and N. Ramanujam, J. Biomed. Opt. 12, 024014 (2007).
[CrossRef] [PubMed]

D. K. Bird, L. Yan, K. M. Vrotsos, K. W. Eliceiri, E. M. Vaughan, P. J. Keely, J. G. White, and N. Ramanujam, Cancer Res. 65, 8766 (2005).
[CrossRef] [PubMed]

Ferrigno, A.

A. C. Croce, A. Ferrigno, M. Vairetti, R. Bertone, I. Freitas, and G. Bottiroli, Photochem. Photobiol. Sci. 3, 920 (2004).
[CrossRef] [PubMed]

Fjeld, C. C.

C. C. Fjeld, W. T. Birdsong, and R. H. Goodman, Proc. Natl. Acad. Sci. U.S.A. 100, 9202 (2003).
[CrossRef] [PubMed]

Freitas, I.

A. C. Croce, A. Ferrigno, M. Vairetti, R. Bertone, I. Freitas, and G. Bottiroli, Photochem. Photobiol. Sci. 3, 920 (2004).
[CrossRef] [PubMed]

Gendron-Fitzpatrick, A.

M. C. Skala, K. M. Riching, D. K. Bird, A. Gendron-Fitzpatrick, J. Eichhoff, K. W. Eliceiri, P. J. Keely, and N. Ramanujam, J. Biomed. Opt. 12, 024014 (2007).
[CrossRef] [PubMed]

Goodman, R. H.

C. C. Fjeld, W. T. Birdsong, and R. H. Goodman, Proc. Natl. Acad. Sci. U.S.A. 100, 9202 (2003).
[CrossRef] [PubMed]

Q. Zhang, D. W. Piston, and R. H. Goodman, Science 295, 1895 (2002).
[PubMed]

Heikal, A. A.

S. Huang, A. A. Heikal, and W. W. Webb, Biophys. J. 82, 2811 (2002).
[CrossRef] [PubMed]

Huang, S.

S. Huang, A. A. Heikal, and W. W. Webb, Biophys. J. 82, 2811 (2002).
[CrossRef] [PubMed]

Keely, P. J.

M. C. Skala, K. M. Riching, D. K. Bird, A. Gendron-Fitzpatrick, J. Eichhoff, K. W. Eliceiri, P. J. Keely, and N. Ramanujam, J. Biomed. Opt. 12, 024014 (2007).
[CrossRef] [PubMed]

D. K. Bird, L. Yan, K. M. Vrotsos, K. W. Eliceiri, E. M. Vaughan, P. J. Keely, J. G. White, and N. Ramanujam, Cancer Res. 65, 8766 (2005).
[CrossRef] [PubMed]

Li, D.

M. S. Yang, D. Li, T. Lin, J. J. Zheng, W. Zheng, and J. Y. Qu, Toxicology 247, 6 (2008).
[CrossRef] [PubMed]

Lin, T.

M. S. Yang, D. Li, T. Lin, J. J. Zheng, W. Zheng, and J. Y. Qu, Toxicology 247, 6 (2008).
[CrossRef] [PubMed]

Lind, B.

T. V. Zglinicki, C. Edwall, E. Ostlund, B. Lind, M. Nordberg, N. R. Ringertz, and J. Wroblewski, J. Cell. Sci. 103, 1073 (1992).

Minet, O.

J. Beuthan, O. Minet, and G. Muller, Ann. N.Y. Acad. Sci. 838, 150 (1998).
[CrossRef] [PubMed]

Muller, G.

J. Beuthan, O. Minet, and G. Muller, Ann. N.Y. Acad. Sci. 838, 150 (1998).
[CrossRef] [PubMed]

Nordberg, M.

T. V. Zglinicki, C. Edwall, E. Ostlund, B. Lind, M. Nordberg, N. R. Ringertz, and J. Wroblewski, J. Cell. Sci. 103, 1073 (1992).

Ostlund, E.

T. V. Zglinicki, C. Edwall, E. Ostlund, B. Lind, M. Nordberg, N. R. Ringertz, and J. Wroblewski, J. Cell. Sci. 103, 1073 (1992).

Piston, D. W.

Q. Zhang, D. W. Piston, and R. H. Goodman, Science 295, 1895 (2002).
[PubMed]

Qu, J. Y.

M. S. Yang, D. Li, T. Lin, J. J. Zheng, W. Zheng, and J. Y. Qu, Toxicology 247, 6 (2008).
[CrossRef] [PubMed]

Y. Wu, W. Zheng, and J. Y. Qu, Opt. Lett. 31, 3122 (2006).
[CrossRef] [PubMed]

Ramanujam, N.

M. C. Skala, K. M. Riching, D. K. Bird, A. Gendron-Fitzpatrick, J. Eichhoff, K. W. Eliceiri, P. J. Keely, and N. Ramanujam, J. Biomed. Opt. 12, 024014 (2007).
[CrossRef] [PubMed]

D. K. Bird, L. Yan, K. M. Vrotsos, K. W. Eliceiri, E. M. Vaughan, P. J. Keely, J. G. White, and N. Ramanujam, Cancer Res. 65, 8766 (2005).
[CrossRef] [PubMed]

Riching, K. M.

M. C. Skala, K. M. Riching, D. K. Bird, A. Gendron-Fitzpatrick, J. Eichhoff, K. W. Eliceiri, P. J. Keely, and N. Ramanujam, J. Biomed. Opt. 12, 024014 (2007).
[CrossRef] [PubMed]

Ringertz, N. R.

T. V. Zglinicki, C. Edwall, E. Ostlund, B. Lind, M. Nordberg, N. R. Ringertz, and J. Wroblewski, J. Cell. Sci. 103, 1073 (1992).

Schulze, H. O.

S. Barban and H. O. Schulze, J. Biol. Chem. 236, 1887 (1961).
[PubMed]

Skala, M. C.

M. C. Skala, K. M. Riching, D. K. Bird, A. Gendron-Fitzpatrick, J. Eichhoff, K. W. Eliceiri, P. J. Keely, and N. Ramanujam, J. Biomed. Opt. 12, 024014 (2007).
[CrossRef] [PubMed]

Vairetti, M.

A. C. Croce, A. Ferrigno, M. Vairetti, R. Bertone, I. Freitas, and G. Bottiroli, Photochem. Photobiol. Sci. 3, 920 (2004).
[CrossRef] [PubMed]

Vaughan, E. M.

D. K. Bird, L. Yan, K. M. Vrotsos, K. W. Eliceiri, E. M. Vaughan, P. J. Keely, J. G. White, and N. Ramanujam, Cancer Res. 65, 8766 (2005).
[CrossRef] [PubMed]

Vrotsos, K. M.

D. K. Bird, L. Yan, K. M. Vrotsos, K. W. Eliceiri, E. M. Vaughan, P. J. Keely, J. G. White, and N. Ramanujam, Cancer Res. 65, 8766 (2005).
[CrossRef] [PubMed]

Webb, W. W.

S. Huang, A. A. Heikal, and W. W. Webb, Biophys. J. 82, 2811 (2002).
[CrossRef] [PubMed]

White, J. G.

D. K. Bird, L. Yan, K. M. Vrotsos, K. W. Eliceiri, E. M. Vaughan, P. J. Keely, J. G. White, and N. Ramanujam, Cancer Res. 65, 8766 (2005).
[CrossRef] [PubMed]

Wroblewski, J.

T. V. Zglinicki, C. Edwall, E. Ostlund, B. Lind, M. Nordberg, N. R. Ringertz, and J. Wroblewski, J. Cell. Sci. 103, 1073 (1992).

Wu, Y.

Yan, L.

D. K. Bird, L. Yan, K. M. Vrotsos, K. W. Eliceiri, E. M. Vaughan, P. J. Keely, J. G. White, and N. Ramanujam, Cancer Res. 65, 8766 (2005).
[CrossRef] [PubMed]

Yang, M. S.

M. S. Yang, D. Li, T. Lin, J. J. Zheng, W. Zheng, and J. Y. Qu, Toxicology 247, 6 (2008).
[CrossRef] [PubMed]

Ying, W.

W. Ying, Antioxid. Redox. Sign. 10, 1 (2008).
[CrossRef]

Zglinicki, T. V.

T. V. Zglinicki, C. Edwall, E. Ostlund, B. Lind, M. Nordberg, N. R. Ringertz, and J. Wroblewski, J. Cell. Sci. 103, 1073 (1992).

Zhang, Q.

Q. Zhang, D. W. Piston, and R. H. Goodman, Science 295, 1895 (2002).
[PubMed]

Zheng, J. J.

M. S. Yang, D. Li, T. Lin, J. J. Zheng, W. Zheng, and J. Y. Qu, Toxicology 247, 6 (2008).
[CrossRef] [PubMed]

Zheng, W.

M. S. Yang, D. Li, T. Lin, J. J. Zheng, W. Zheng, and J. Y. Qu, Toxicology 247, 6 (2008).
[CrossRef] [PubMed]

Y. Wu, W. Zheng, and J. Y. Qu, Opt. Lett. 31, 3122 (2006).
[CrossRef] [PubMed]

Ann. N.Y. Acad. Sci.

J. Beuthan, O. Minet, and G. Muller, Ann. N.Y. Acad. Sci. 838, 150 (1998).
[CrossRef] [PubMed]

Antioxid. Redox. Sign.

W. Ying, Antioxid. Redox. Sign. 10, 1 (2008).
[CrossRef]

Biophys. J.

S. Huang, A. A. Heikal, and W. W. Webb, Biophys. J. 82, 2811 (2002).
[CrossRef] [PubMed]

Cancer Res.

D. K. Bird, L. Yan, K. M. Vrotsos, K. W. Eliceiri, E. M. Vaughan, P. J. Keely, J. G. White, and N. Ramanujam, Cancer Res. 65, 8766 (2005).
[CrossRef] [PubMed]

J. Biol. Chem.

S. Barban and H. O. Schulze, J. Biol. Chem. 236, 1887 (1961).
[PubMed]

J. Biomed. Opt.

M. C. Skala, K. M. Riching, D. K. Bird, A. Gendron-Fitzpatrick, J. Eichhoff, K. W. Eliceiri, P. J. Keely, and N. Ramanujam, J. Biomed. Opt. 12, 024014 (2007).
[CrossRef] [PubMed]

J. Cell. Sci.

T. V. Zglinicki, C. Edwall, E. Ostlund, B. Lind, M. Nordberg, N. R. Ringertz, and J. Wroblewski, J. Cell. Sci. 103, 1073 (1992).

Opt. Lett.

Photochem. Photobiol. Sci.

A. C. Croce, A. Ferrigno, M. Vairetti, R. Bertone, I. Freitas, and G. Bottiroli, Photochem. Photobiol. Sci. 3, 920 (2004).
[CrossRef] [PubMed]

Proc. Natl. Acad. Sci. U.S.A.

C. C. Fjeld, W. T. Birdsong, and R. H. Goodman, Proc. Natl. Acad. Sci. U.S.A. 100, 9202 (2003).
[CrossRef] [PubMed]

Science

Q. Zhang, D. W. Piston, and R. H. Goodman, Science 295, 1895 (2002).
[PubMed]

Toxicology

M. S. Yang, D. Li, T. Lin, J. J. Zheng, W. Zheng, and J. Y. Qu, Toxicology 247, 6 (2008).
[CrossRef] [PubMed]

Other

M. K. Campbell, Biochemistry, (Saunders College Publishing, 1995).
[PubMed]

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Figures (2)

Fig. 1
Fig. 1

Representative images of SiHa cell culture in pseudocolor: (a) SPF image; (b) TPF image.

Fig. 2
Fig. 2

Pseudocolor images and decay associate spectra (DAS) signals of three cellular compartments: (a) SPF image of SiHa cells; (b) extracted mitochondrial compartment; (c) cytosolic compartment; (d) nuclear compartment; (e) normalized short lifetime DAS of SiHa cells and fluorescence spectrum of pure NADH; (f) Normalized long lifetime DAS of SiHa cells and fluorescence spectrum of NADH bound with lactate dehydrogenase (LDH).

Tables (1)

Tables Icon

Table 1 A 1 A 2 Ratio and Lifetimes Measured from SiHa Cell Cultures a, b, c

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