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Separating NADH and NADPH fluorescence in live cells and tissues using FLIM.
Blacker, Thomas S; Mann, Zoe F; Gale, Jonathan E; Ziegler, Mathias; Bain, Angus J; Szabadkai, Gyorgy; Duchen, Michael R.
Afiliación
  • Blacker TS; 1] Centre for Mathematics and Physics in the Life Sciences and Experimental Biology, University College London, London WC1E 6BT, UK [2] Research Department of Cell & Developmental Biology, University College London, London WC1E 6BT, UK [3] Department of Physics and Astronomy, University College
  • Mann ZF; 1] Research Department of Cell & Developmental Biology, University College London, London WC1E 6BT, UK [2] UCL Ear Institute, University College London, London WC1X 8EE, UK.
  • Gale JE; 1] Research Department of Cell & Developmental Biology, University College London, London WC1E 6BT, UK [2] UCL Ear Institute, University College London, London WC1X 8EE, UK.
  • Ziegler M; Department of Molecular Biology, University of Bergen, N-5008 Bergen, Norway.
  • Bain AJ; Department of Physics and Astronomy, University College London, London WC1E 6BT, UK.
  • Szabadkai G; 1] Research Department of Cell & Developmental Biology, University College London, London WC1E 6BT, UK [2] Department of Biomedical Sciences, University of Padua and CNR Neuroscience Institute, Padua 35121, Italy [3].
  • Duchen MR; 1] Research Department of Cell & Developmental Biology, University College London, London WC1E 6BT, UK [2].
Nat Commun ; 5: 3936, 2014 May 29.
Article en En | MEDLINE | ID: mdl-24874098
NAD is a key determinant of cellular energy metabolism. In contrast, its phosphorylated form, NADP, plays a central role in biosynthetic pathways and antioxidant defence. The reduced forms of both pyridine nucleotides are fluorescent in living cells but they cannot be distinguished, as they are spectrally identical. Here, using genetic and pharmacological approaches to perturb NAD(P)H metabolism, we find that fluorescence lifetime imaging (FLIM) differentiates quantitatively between the two cofactors. Systematic manipulations to change the balance between oxidative and glycolytic metabolism suggest that these states do not directly impact NAD(P)H fluorescence decay rates. The lifetime changes observed in cancers thus likely reflect shifts in the NADPH/NADH balance. Using a mathematical model, we use these experimental data to quantify the relative levels of NADH and NADPH in different cell types of a complex tissue, the mammalian cochlea. This reveals NADPH-enriched populations of cells, raising questions about their distinct metabolic roles.
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Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Asunto principal: Oxidación-Reducción / Cóclea / Imagen Óptica / Glucólisis / NAD / NADP Límite: Animals / Humans Idioma: En Revista: Nat Commun Asunto de la revista: BIOLOGIA / CIENCIA Año: 2014 Tipo del documento: Article Pais de publicación: Reino Unido
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Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Asunto principal: Oxidación-Reducción / Cóclea / Imagen Óptica / Glucólisis / NAD / NADP Límite: Animals / Humans Idioma: En Revista: Nat Commun Asunto de la revista: BIOLOGIA / CIENCIA Año: 2014 Tipo del documento: Article Pais de publicación: Reino Unido