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Isolation and in vitro expansion of human colonic stem cells

Nature Medicine volume 17, pages 1225–1227 (2011)Cite this article

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Abstract

Here we describe the isolation of stem cells of the human colonic epithelium. Differential cell surface abundance of ephrin type-B receptor 2 (EPHB2) allows the purification of different cell types from human colon mucosa biopsies. The highest EPHB2 surface levels correspond to epithelial colonic cells with the longest telomeres and elevated expression of intestinal stem cell (ISC) marker genes. Moreover, using culturing conditions that recreate the ISC niche, a substantial proportion of EPHB2-high cells can be expanded in vitro as an undifferentiated and multipotent population.

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Figure 1: Characterization of epithelial colon cells purified from human mucosa samples according to different EPHB2 surface levels.
Figure 2: Single EPHB2high human colonic crypt cells form in vitro spheroids with long-term proliferation and multilineage differentiation capacity.

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  • 20 September 2011

     In the version of this article initially published online, the author list was missing the name Herbert Auer, and the name of Mercedes Gallardo was incorrectly given as María M. Gallardo. The error has been corrected for the print, PDF and HTML versions of this article.

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Acknowledgements

We thank Genentech for the EphB2-specific antibody Mab 2H9 and the Institute for Research in Biomedicine transcriptomic facility, electronic microscopy service (Universitat de Barcelona) and Cell separation unit (UB) for expert assistance. We thank L. Lopez Vilaro and E. Calderón Gómez for help with mucosas. This work is supported by grants to E.B. from the CONSOLIDER and Plan Nacional I+D+i Programs from Spanish Ministry of Science and Innovation (MICINN) and the European Research Council (ERC-FP7). Work was furthermore supported by grants from Instituto de Salud Carlos III FEDER (RD09/0076/00036). The 'Xarxa de Bancs de tumors' is sponsored by Pla Director d'Oncologia de Catalunya (XBTC). The M.A.B. laboratory is founded by the MICINN, the EU, the Botin and Lilly Foundations, and the Korber European Science Award to M.A.B. P.J. holds a Marie Curie Fellowship (FP7).

Author information

Author notes

  1. Peter Jung and Toshiro Sato: These authors contributed equally to this work.

Authors and Affiliations

  1. Oncology program, Institute for Research in Biomedicine, Barcelona, Spain

    Peter Jung, Anna Merlos-Suárez, Francisco M Barriga, Elena Sancho & Eduard Batlle

  2. Hubrecht Institute and University Medical Center Utrecht, Utrecht, The Netherlands

    Toshiro Sato & Hans Clevers

  3. Department of Gastroenterology, School of Medicine, Keio University, Shinjuku-ku, Tokyo, Japan

    Toshiro Sato

  4. Department of Pathology, Hospital Universitari del Mar, Universitat Autònoma de Barcelona, Barcelona, Spain

    Mar Iglesias

  5. Biostatistics and Bioinformatics Unit, Institute for Research in Biomedicine, Barcelona, Spain

    David Rossell

  6. Functional Genomics Core Facility, Institute for Research in Biomedicine, Barcelona, Spain

    Herbert Auer

  7. Telomeres and Telomerase Group, Molecular Oncology Program, Spanish National Cancer Research Center, Madrid, Spain

    Mercedes Gallardo & Maria A Blasco

  8. Institució Catalana de Recerca i Estudis Avançats, Barcelona, Spain.,

    Eduard Batlle

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  1. Peter Jung
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Contributions

E.B. and P.J., study concept and design and writing of the manuscript. T.S. and H.C., development of original human colon culture method. P.J., acquisition, analysis and interpretation of data and modifications of the human CoSC culture protocol. A.M.-S. and F.M.B., profiling of human colon stem cells. D.R., bioinformatic analysis. E.S., conceptual and logistic support. M.M.G. and M.A.B., telomere length measurements. M.I., sample preparation and development of EphB2 sorting method.

Corresponding author

Correspondence to Eduard Batlle.

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Competing interests

The authors declare no competing financial interests.

Supplementary information

Supplementary Text and Figures

Supplementary Figures 1–11, Supplementary Data, Supplementary Table 1 and Supplementary Methods (PDF 2109 kb)

Supplementary Table 2

List of genes whose expression is enriched in EPHB2-high cells. (XLS 36 kb)

Supplementary Table 3

Genome-wide expression analysis of EPHB2-high, EPHB2-medium, EPHB2-low and EPHB2-negative cells from 3 independent colon mucosa samples. (XLS 20006 kb)

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Jung, P., Sato, T., Merlos-Suárez, A. et al. Isolation and in vitro expansion of human colonic stem cells. Nat Med 17, 1225–1227 (2011). https://doi.org/10.1038/nm.2470

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