Fourier Transform Infrared Microspectroscopy Identifies Symmetric PO2 Modifications as a Marker of the Putative Stem Cell Region of Human Intestinal Crypts

Authors

  • Michael J. Walsh,

    1. Biomedical Sciences Unit, Department of Biological Sciences, Lancaster University, Bailrigg, Lancaster, United Kingdom
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  • Tariq G. Fellous,

    1. Centre for Diabetes and Metabolic Medicine, Queen Mary's School of Medicine and Dentistry, Institute of Cell and Molecular Science, London, United Kingdom
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  • Azzedine Hammiche,

    1. Department of Physics, Lancaster University, Bailrigg, Lancaster, United Kingdom
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  • Wey-Ran Lin,

    1. Department of Gastroenterology and Hepatology, Chang Gung Memorial Hospital, Chang Gung University College of Medicine, Taiwan
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  • Nigel J. Fullwood,

    1. Biomedical Sciences Unit, Department of Biological Sciences, Lancaster University, Bailrigg, Lancaster, United Kingdom
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  • Olaug Grude,

    1. Biomedical Sciences Unit, Department of Biological Sciences, Lancaster University, Bailrigg, Lancaster, United Kingdom
    2. Department of Physics, Lancaster University, Bailrigg, Lancaster, United Kingdom
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  • Fariba Bahrami,

    1. Synchrotron Radiation Department, Daresbury Laboratories, Science and Technologies Facilities Council, Warrington, United Kingdom
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  • James M. Nicholson,

    1. Synchrotron Radiation Department, Daresbury Laboratories, Science and Technologies Facilities Council, Warrington, United Kingdom
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  • Marine Cotte,

    1. European Synchrotron Radiation Facility, Grenoble, France
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  • Jean Susini,

    1. European Synchrotron Radiation Facility, Grenoble, France
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  • Hubert M. Pollock,

    1. Department of Physics, Lancaster University, Bailrigg, Lancaster, United Kingdom
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  • Mairi Brittan,

    1. Centre for Diabetes and Metabolic Medicine, Queen Mary's School of Medicine and Dentistry, Institute of Cell and Molecular Science, London, United Kingdom
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  • Pierre L. Martin-Hirsch,

    1. Biomedical Sciences Unit, Department of Biological Sciences, Lancaster University, Bailrigg, Lancaster, United Kingdom
    2. Sharoe Green Unit, Lancashire Teaching Hospitals NHS Trust, Preston, United Kingdom
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  • Malcolm R. Alison,

    1. Centre for Diabetes and Metabolic Medicine, Queen Mary's School of Medicine and Dentistry, Institute of Cell and Molecular Science, London, United Kingdom
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  • Francis L. Martin Ph.D.

    Corresponding author
    1. Biomedical Sciences Unit, Department of Biological Sciences, Lancaster University, Bailrigg, Lancaster, United Kingdom
    • Biomedical Sciences Unit, Department of Biological Sciences, Lancaster University, Bailrigg, Lancaster LA1 4YQ, U.K. Telephone: +44-1524-594505; Fax: +44-1524-593192
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Abstract

Complex biomolecules absorb in the mid-infrared (λ = 2–20 μm), giving vibrational spectra associated with structure and function. We used Fourier transform infrared (FTIR) microspectroscopy to “fingerprint” locations along the length of human small and large intestinal crypts. Paraffin-embedded slices of normal human gut were sectioned (10 μm thick) and mounted to facilitate infrared (IR) spectral analyses. IR spectra were collected using globar (15 μm × 15 μm aperture) FTIR microspectroscopy in reflection mode, synchrotron (≤10 μm × 10 μm aperture) FTIR microspectroscopy in transmission mode or near-field photothermal microspectroscopy. Dependent on the location of crypt interrogation, clear differences in spectral characteristics were noted. Epithelial-cell IR spectra were subjected to principal component analysis to determine whether wavenumber-absorbance relationships expressed as single points in “hyperspace” might on the basis of multivariate distance reveal biophysical differences along the length of gut crypts. Following spectroscopic analysis, plotted clusters and their loadings plots pointed toward symmetric (νs)PO2 (1,080 cm−1) vibrations as a discriminating factor for the putative stem cell region; this proved to be a more robust marker than other phenotypic markers, such as β-catenin or CD133. This pattern was subsequently confirmed by image mapping and points to a novel approach of nondestructively identifying a tissue's stem cell location. νsPO2, probably associated with DNA conformational alterations, might facilitate a means of identifying stem cells, which may have utility in other tissues where the location of stem cells is unclear.

Disclosure of potential conflicts of interest is found at the end of this article.

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