pp 1-14 | Cite as

Isolation and Enrichment of Newborn and Adult Skin Stem Cells of the Interfollicular Epidermis

Protocol
Part of the Methods in Molecular Biology book series

Abstract

The interfollicular epidermis regenerates from a heterogeneous population of basal cells undergoing either self-renewal or terminal differentiation, thereby balancing cell loss in tissue turnover or in wound repair. In this chapter, we describe a reliable and simple method for isolating interfollicular epithelial stem cells from the skin of newborn mice or from tail and ear skin of adult mice using fluorescence-activated cell sorting (FACS). We also provide a detailed protocol for culturing interfollicular epidermal stem cells and to assess their proliferative potential and self-renewing ability. These techniques are useful for directly evaluating epidermal stem cell function in normal mice under different conditions or in genetically modified mouse models.

Keywords

Adult stem cells CD34 CD71 Clonogenic assay Epidermis Flow cytometry Integrins Isolation Keratinocytes Progenitor cells 

Notes

Acknowledgments

This work was supported by the Telethon grant GGP16235 and the Italian Association for Cancer Research (AIRC IG2011-N.11369).

References

  1. 1.
    Gonzales KAU, Fuchs E (2017) Skin and its regenerative powers: an alliance between stem cells and their niche. Dev Cell 43(4):387–401.  https://doi.org/10.1016/j.devcel.2017.10.001CrossRefPubMedGoogle Scholar
  2. 2.
    Clayton E, Doupe DP, Klein AM, Winton DJ, Simons BD, Jones PH (2007) A single type of progenitor cell maintains normal epidermis. Nature 446(7132):185–189.  https://doi.org/10.1038/nature05574ADSCrossRefPubMedGoogle Scholar
  3. 3.
    Doupe DP, Klein AM, Simons BD, Jones PH (2010) The ordered architecture of murine ear epidermis is maintained by progenitor cells with random fate. Dev Cell 18(2):317–323.  https://doi.org/10.1016/j.devcel.2009.12.016CrossRefPubMedGoogle Scholar
  4. 4.
    Mascre G, Dekoninck S, Drogat B, Youssef KK, Brohee S, Sotiropoulou PA, Simons BD, Blanpain C (2012) Distinct contribution of stem and progenitor cells to epidermal maintenance. Nature 489(7415):257–262.  https://doi.org/10.1038/nature11393ADSCrossRefPubMedGoogle Scholar
  5. 5.
    Rompolas P, Mesa KR, Kawaguchi K, Park S, Gonzalez D, Brown S, Boucher J, Klein AM, Greco V (2016) Spatiotemporal coordination of stem cell commitment during epidermal homeostasis. Science 352(6292):1471–1474.  https://doi.org/10.1126/science.aaf7012ADSCrossRefPubMedPubMedCentralGoogle Scholar
  6. 6.
    Sada A, Jacob F, Leung E, Wang S, White BS, Shalloway D, Tumbar T (2016) Defining the cellular lineage hierarchy in the interfollicular epidermis of adult skin. Nat Cell Biol 18(6):619–631.  https://doi.org/10.1038/ncb3359CrossRefPubMedPubMedCentralGoogle Scholar
  7. 7.
    Lim X, Tan SH, Koh WL, Chau RM, Yan KS, Kuo CJ, van Amerongen R, Klein AM, Nusse R (2013) Interfollicular epidermal stem cells self-renew via autocrine Wnt signaling. Science 342(6163):1226–1230.  https://doi.org/10.1126/science.1239730ADSCrossRefPubMedPubMedCentralGoogle Scholar
  8. 8.
    Asare A, Levorse J, Fuchs E (2017) Coupling organelle inheritance with mitosis to balance growth and differentiation. Science 355(6324). pii: eaah4701).  https://doi.org/10.1126/science.aah4701
  9. 9.
    Jensen UB, Lowell S, Watt FM (1999) The spatial relationship between stem cells and their progeny in the basal layer of human epidermis: a new view based on whole-mount labelling and lineage analysis. Development 126(11):2409–2418PubMedGoogle Scholar
  10. 10.
    Jones PH, Watt FM (1993) Separation of human epidermal stem cells from transit amplifying cells on the basis of differences in integrin function and expression. Cell 73(4):713–724CrossRefPubMedGoogle Scholar
  11. 11.
    Tani H, Morris RJ, Kaur P (2000) Enrichment for murine keratinocyte stem cells based on cell surface phenotype. Proc Natl Acad Sci U S A 97(20):10960–10965ADSCrossRefPubMedPubMedCentralGoogle Scholar
  12. 12.
    Ferone G, Thomason HA, Antonini D, De Rosa L, Hu B, Gemei M, Zhou H, Ambrosio R, Rice DP, Acampora D, van Bokhoven H, Del Vecchio L, Koster MI, Tadini G, Spencer-Dene B, Dixon M, Dixon J, Missero C (2012) Mutant p63 causes defective expansion of ectodermal progenitor cells and impaired FGF signalling in AEC syndrome. EMBO Mol Med 4(3):192–205.  https://doi.org/10.1002/emmm.201100199CrossRefPubMedPubMedCentralGoogle Scholar
  13. 13.
    Blanpain C, Lowry WE, Geoghegan A, Polak L, Fuchs E (2004) Self-renewal, multipotency, and the existence of two cell populations within an epithelial stem cell niche. Cell 118(5):635–648.  https://doi.org/10.1016/j.cell.2004.08.012CrossRefPubMedGoogle Scholar
  14. 14.
    Jensen KB, Driskell RR, Watt FM (2010) Assaying proliferation and differentiation capacity of stem cells using disaggregated adult mouse epidermis. Nat Protoc 5(5):898–911.  https://doi.org/10.1038/nprot.2010.39CrossRefPubMedGoogle Scholar
  15. 15.
    Nowak JA, Fuchs E (2009) Isolation and culture of epithelial stem cells. Methods Mol Biol 482:215–232.  https://doi.org/10.1007/978-1-59745-060-7_14CrossRefPubMedPubMedCentralGoogle Scholar

Copyright information

© Springer Science+Business Media New York 2018

Authors and Affiliations

  • Stefano Sol
    • 1
    • 2
  • Dario Antonini
    • 1
    • 2
  • Caterina Missero
    • 1
    • 2
  1. 1.Department of BiologyUniversity of Naples Federico IINapoliItaly
  2. 2.CEINGE Biotecnologie Avanzate (Center for Genetic Engineering)NapoliItaly

Personalised recommendations