Letter | Published:

Intestinal crypt homeostasis revealed at single-stem-cell level by in vivo live imaging

Nature volume 507, pages 362365 (20 March 2014) | Download Citation


The rapid turnover of the mammalian intestinal epithelium is supported by stem cells located around the base of the crypt1. In addition to the Lgr5 marker, intestinal stem cells have been associated with other markers that are expressed heterogeneously within the crypt base region1,2,3,4,5,6. Previous quantitative clonal fate analyses have led to the proposal that homeostasis occurs as the consequence of neutral competition between dividing stem cells7,8,9. However, the short-term behaviour of individual Lgr5+ cells positioned at different locations within the crypt base compartment has not been resolved. Here we establish the short-term dynamics of intestinal stem cells using the novel approach of continuous intravital imaging of Lgr5-Confetti mice. We find that Lgr5+ cells in the upper part of the niche (termed ‘border cells’) can be passively displaced into the transit-amplifying domain, after the division of proximate cells, implying that the determination of stem-cell fate can be uncoupled from division. Through quantitative analysis of individual clonal lineages, we show that stem cells at the crypt base, termed ‘central cells’, experience a survival advantage over border stem cells. However, through the transfer of stem cells between the border and central regions, all Lgr5+ cells are endowed with long-term self-renewal potential. These findings establish a novel paradigm for stem-cell maintenance in which a dynamically heterogeneous cell population is able to function long term as a single stem-cell pool.

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The authors would like to thank A. de Graaff from the Hubrecht Imaging Center for imaging support, all members of the van Rheenen group for useful discussions and the Hubrecht Institute animal caretakers for animal support. This work was supported by a Vidi fellowship (91710330; J.v.R.) and equipment grants (175.010.2007.00 and 834.11.002; J.v.R.) from the Dutch Organization of Scientific Research (NWO), a grant from the Dutch Cancer Society (KWF; HUBR 2009-4621; J.v.R.), a grant from the Association for International Cancer Research (AICR; 13-0297; J.v.R.), and the Wellcome Trust (grant number 098357/Z/12/Z; B.D.S.).

Author information

Author notes

    • Laila Ritsma
    •  & Saskia I. J. Ellenbroek

    These authors contributed equally to this work.


  1. Cancer Genomics Netherlands, Hubrecht Institute-KNAW and University Medical Centre Utrecht, Uppsalalaan 8, 3584 CT Utrecht, The Netherlands

    • Laila Ritsma
    • , Saskia I. J. Ellenbroek
    • , Anoek Zomer
    • , Hans Clevers
    •  & Jacco van Rheenen
  2. University Medical Centre Utrecht, Universiteitsweg 100, 3584 CG Utrecht, The Netherlands

    • Hugo J. Snippert
  3. Department of Molecular Biology, Genentech Inc., 1 DNA Way, South San Francisco, California 94080, USA

    • Frederic J. de Sauvage
  4. Cavendish Laboratory, Department of Physics, J. J. Thomson Avenue, University of Cambridge, Cambridge CB3 0HE, UK

    • Benjamin D. Simons
  5. The Wellcome Trust/Cancer Research UK Gurdon Institute, University of Cambridge, Tennis Court Road, Cambridge CB2 1QN, UK

    • Benjamin D. Simons
  6. The Wellcome Trust/Medical Research Council Stem Cell Institute, University of Cambridge, Tennis Court Road, Cambridge CB2 1QN, UK

    • Benjamin D. Simons


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J.v.R. and L.R. conceived the study. L.R. optimized the surgical and imaging procedure. L.R., S.I.J.E., A.Z. and H.J.S. performed imaging experiments. L.R., H.J.S., B.D.S. and S.I.J.E. performed analyses. F.J.d.S. provided the Lgr5DTR:eGFP mice and B.D.S. did all biophysical modelling. L.R. and S.I.J.E. made the figures. J.v.R. and H.C. supervised the study. All authors discussed results and participated in preparation of the manuscript.

Competing interests

The authors declare no competing financial interests.

Corresponding authors

Correspondence to Hans Clevers or Jacco van Rheenen.

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  1. 1.

    Supplementary Information

    This file contains Supplementary Notes and an additional reference.


  1. 1.

    3D reconstruction of a crypt containing Lgr5+ CBC cells

    Lgr5+ CBC cells are shown in green, non-CBC cells are shown in red and Collagen 1 is shown in blue.

  2. 2.

    Dynamics of Lgr5+ CBC cells at the crypt base

    Left, time series of Lgr5+ CBC cells in a crypt. In the cartoons, Lgr5+ CBC cells are highlighted and the moving cells are colour coded. In the right two videos cell tracks are shown by lines and the centre of cells is indicated by dots. The time is indicated in hours. Scale bar, 20 µm.

  3. 3.

    Dividing and moving Lgr5+ CBC cells

    Three days after Tamoxifen administration, mice were imaged in real-time. Lgr5+ CBC cells within the entire stem-cell compartment of a single crypt are projected onto one plane and are shown over time. Note the outlined cells that divide and move.

  4. 4.

    Lgr5+ CBC cells get expelled from the stem-cell niche

    Three days after Tamoxifen administration, mice were imaged in real-time. Lgr5+ CBC cells within the entire stem-cell compartment of a single crypt are projected onto one plane and they are shown over time. Note the outlined cells that divide, move, and disappear.

  5. 5.

    Heterogeneous recovery of stem cells following targeted ablation

    In mice where the human DT receptor (DTR) fused to eGFP was knocked in the Lgr5 locus (Lgr5DTR:eGFP), Lgr5+ cells were fully ablated using diphtheria toxin (DT) injection. The recovery was monitored by acquiring a z-stack at 58hrs. The video shows the z-stack. The empty crypts are indicated with yellow. The crypts containing one recovered cell are indicated with red. The crypts containing more than 1 recovered cell are indicated with grey.

  6. 6.

    Cohesion of clusters of Lgr5+ cells

    In mice where the human DT receptor (DTR) fused to eGFP was knocked in the Lgr5 locus (Lgr5DTR:eGFP), Lgr5+ cells were fully ablated using diphtheria toxin (DT) injection. The video shows a 3D reconstruction of clusters of recovered Lgr5+ cells from different sizes found 72hrs after ablation. Note the cohesion of these clusters, which is suggesting their origin lies in clonal expansion of cells.

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