Competitive Repopulation Assay of Long-Term Epidermal Stem Cell Regeneration Potential

  • Carmen Segrelles
  • Karla Santos-de-Frutos
  • Jesús M. Paramio
  • Corina LorzEmail author
Part of the Methods in Molecular Biology book series (MIMB, volume 2109)


Epidermal stem cells are responsible for normal tissue homeostasis and contribute to tissue regeneration during injury. Several assays measuring stem cell frequency and function can be used to assess epidermal stem cell potential. However, the ultimate assay that accounts for stemness is the capacity to sustain in vivo long-term tissue regeneration and maintenance. We can use this type of analysis to interrogate whether a specific genetic alteration (e.g., activation or inactivation of any gene thought to be involved in stem cell quiescence or proliferation) confers increased or decreased stem cell potential.


Epidermal stem cells Keratinocyte transplantation Skin regeneration Silicone chambers 



Work in our laboratory is supported by FEDER cofounded ISCIII grants [CB16/12/00228 and PI18/00263] and Foundation of Fanconi Anemia grant [2018/127].


  1. 1.
    Segrelles C, Garcia-Escudero R, Garin MI, Aranda JF, Hernandez P, Ariza JM et al (2014) Akt signaling leads to stem cell activation and promotes tumor development in epidermis. Stem Cells 32(7):1917–1928. CrossRefPubMedGoogle Scholar
  2. 2.
    Lorz C, Garcia-Escudero R, Segrelles C, Garin MI, Ariza JM, Santos M et al (2010) A functional role of RB-dependent pathway in the control of quiescence in adult epidermal stem cells revealed by genomic profiling. Stem Cell Rev 6(2):162–177. CrossRefPubMedCentralGoogle Scholar
  3. 3.
    Wu WY, Morris RJ (2005) Method for the harvest and assay of in vitro clonogenic keratinocytes stem cells from mice. Methods Mol Biol 289:79–86PubMedGoogle Scholar
  4. 4.
    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. CrossRefPubMedGoogle Scholar
  5. 5.
    Lorz C, Segrelles C, Garin M, Paramio JM (2010) Isolation of adult mouse stem keratinocytes using magnetic cell sorting (MACS). Methods Mol Biol 585:1–11. CrossRefPubMedGoogle Scholar
  6. 6.
    Strachan LR, Ghadially R (2010) Limiting dilution analysis of murine epidermal stem cells using an in vivo regeneration assay. Methods Mol Biol 585:421–432. CrossRefPubMedPubMedCentralGoogle Scholar
  7. 7.
    Strachan LR, Scalapino KJ, Lawrence HJ, Ghadially R (2008) Rapid adhesion to collagen isolates murine keratinocytes with limited long-term repopulating ability in vivo despite high clonogenicity in vitro. Stem Cells 26(1):235–243. CrossRefPubMedGoogle Scholar
  8. 8.
    Schneider TE, Barland C, Alex AM, Mancianti ML, Lu Y, Cleaver JE et al (2003) Measuring stem cell frequency in epidermis: a quantitative in vivo functional assay for long-term repopulating cells. Proc Natl Acad Sci U S A 100(20):11412–11417. CrossRefPubMedPubMedCentralGoogle Scholar
  9. 9.
    Segrelles C, Lu J, Hammann B, Santos M, Moral M, Cascallana JL et al (2007) Deregulated activity of Akt in epithelial basal cells induces spontaneous tumors and heightened sensitivity to skin carcinogenesis. Cancer Res 67(22):10879–10888. CrossRefPubMedGoogle Scholar
  10. 10.
    Okabe M, Ikawa M, Kominami K, Nakanishi T, Nishimune Y (1997) ‘Green mice’ as a source of ubiquitous green cells. FEBS Lett 407(3):313–319CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media New York 2019

Authors and Affiliations

  • Carmen Segrelles
    • 1
    • 2
    • 3
  • Karla Santos-de-Frutos
    • 1
    • 2
    • 3
  • Jesús M. Paramio
    • 1
    • 2
    • 3
  • Corina Lorz
    • 1
    • 2
    • 3
    Email author
  1. 1.Molecular Oncology Unit, CIEMAT (ed 70A)MadridSpain
  2. 2.Molecular OncologyUniversity Hospital 12 de Octubre, Research Institute 12 de Octubre i+12MadridSpain
  3. 3.Centro de Investigación Biomédica en Red de Cáncer (CIBERONC)MadridSpain

Personalised recommendations