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Paneth cells constitute the niche for Lgr5 stem cells in intestinal crypts

Nature volume 469, pages 415–418 (2011)Cite this article

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Abstract

Homeostasis of self-renewing small intestinal crypts results from neutral competition between Lgr5 stem cells, which are small cycling cells located at crypt bottoms1,2. Lgr5 stem cells are interspersed between terminally differentiated Paneth cells that are known to produce bactericidal products such as lysozyme and cryptdins/defensins3. Single Lgr5-expressing stem cells can be cultured to form long-lived, self-organizing crypt–villus organoids in the absence of non-epithelial niche cells4. Here we find a close physical association of Lgr5 stem cells with Paneth cells in mice, both in vivo and in vitro. CD24+ Paneth cells express EGF, TGF-α, Wnt3 and the Notch ligand Dll4, all essential signals for stem-cell maintenance in culture. Co-culturing of sorted stem cells with Paneth cells markedly improves organoid formation. This Paneth cell requirement can be substituted by a pulse of exogenous Wnt. Genetic removal of Paneth cells in vivo results in the concomitant loss of Lgr5 stem cells. In colon crypts, CD24+ cells residing between Lgr5 stem cells may represent the Paneth cell equivalents. We conclude that Lgr5 stem cells compete for essential niche signals provided by a specialized daughter cell, the Paneth cell.

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Figure 1: Geometric distribution pattern of Paneth cells and Lgr5 stem cells.
Figure 2: Paneth cells express CD24 and support growth of Lgr5 stem cells.
Figure 3: Paneth cells produce Wnt3 and other essential niche signals for Lgr5 stem cells.
Figure 4: Paneth cells regulate numbers of intestinal stem cells in vivo.

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Gene Expression Omnibus

Data deposits

The data for the microarray analysis were deposited to the GEO database under accession number GSE25109.

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Acknowledgements

We thank H. Begthel, J. Korving and S. van den Brink for technical assistance; J. Gordon for providing small intestinal sections from Cr2-tox177 mice; L. Lum for providing IWP1; and A. Abo for R-spondin 1.

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Authors and Affiliations

  1. Hubrecht Institute, KNAW and University Medical Center Utrecht, Uppsalalaan 8, 3584CT Utrecht, the Netherlands ,

    Toshiro Sato, Johan H. van Es, Hugo J. Snippert, Daniel E. Stange, Robert G. Vries, Maaike van den Born, Nick Barker, Marc van de Wetering & Hans Clevers

  2. Division of Gastroenterology, Cincinnati Children’s Hospital, Medical Center, MLC 2010, 3333 Burnet Avenue, Cincinnati, 45229, Ohio, USA

    Noah F. Shroyer

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  1. Toshiro Sato
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Contributions

T.S. and H.C. conceived and designed the experiments. T.S., J.H.v.E., H.J.S., R.G.V., M.v.d.B., N.B. and M.v.d.W. performed the experiments, and T.S., J.H.v.E., H.J.S., D.E.S. and H.C. analysed the data. N.F.S. provided Gfi1−/− mice intestines. T.S. and H.C. wrote the manuscript.

Corresponding author

Correspondence to Hans Clevers.

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Competing interests

H.C. is an inventor on several patents involving the culture system in this paper, as is T.S.

Supplementary information

Supplementary Figures

The file contains Supplementary Figures 1-6 with legends. (PDF 2696 kb)

Supplementary Movie 1

The movie shows a growing organoid from an Lgr5-EGFP-ires-CreERT2 crypt. The organoid was filmed on a live-imaging confocal microscope for 2.5 days. Note the appearance of GFP-positive Lgr5 stem cells near differential interference contrast-positive Paneth cells at sites of crypt budding. (AVI 5811 kb)

Supplementary Movie 2

The movie shows time course of reassociation of sorted single stem cells and single Paneth cells. 1500 single RFP+ stem cells were recombined with 1500 single YFP+ Paneth cells and filmed over 10 days. Of note, we observed one YFP+ budding structure formation originated from contaminated stem cells in YFP+ Paneth cells. (AVI 24322 kb)

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Sato, T., van Es, J., Snippert, H. et al. Paneth cells constitute the niche for Lgr5 stem cells in intestinal crypts. Nature 469, 415–418 (2011). https://doi.org/10.1038/nature09637

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