Iterative Three-Dimensional Epidermis Bioengineering and Xenografting to Assess Long-Term Regenerative Potential in Human Keratinocyte Precursor Cells

  • Nicolas O. FortunelEmail author
  • Emmanuelle Bouissou-Cadio
  • Julien Coutier
  • Michèle T. Martin
Part of the Methods in Molecular Biology book series (MIMB, volume 2109)


The functional definition of somatic adult stem cells is based on their regenerative capacity, which allows tissue regeneration throughout life. Thus, refining methodologies to characterize this capacity is of great importance for progress in the fundamental knowledge of specific keratinocyte subpopulations but also for preclinical and clinical research, considering the high potential of keratinocytes in cell therapy. We present here a methodology which we define as iterative xenografting, which originates in the classical model of human skin substitute xenografts onto immunodeficient recipient mice. The principle of this functional assay is first to perform primary xenografts to assess graft take and the quality of epidermal differentiation. Then, human keratinocytes are extracted from primary graft samples to perform secondary xenografts, to assess the presence and preservation of functional keratinocyte stem cells with long-term regenerative potential. In the example of experiments shown, iterative skin xenografting was used to document the high regenerative potential of epidermal holoclone keratinocytes.


Epidermis Epidermis reconstruction Keratinocyte Regeneration Stem cell Xenograft 



We wish to thank S. Bouet and A. Boukadiri (histology platform, UMR 1313 GABI, INRA/CEA, Jouy en Josas). We thank J.-J. Lataillade and M. Trouillas (IRBA, INSERM U1197, Clamart) for helpful discussions on skin substitute bioengineering. This work was supported by grants from CEA and INSERM (UMR967), the Délégation Générale de l’Armement (DGA), and the “Les Gueules Cassées” foundation.


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Copyright information

© Springer Science+Business Media New York 2019

Authors and Affiliations

  • Nicolas O. Fortunel
    • 1
    • 2
    • 3
    • 4
    Email author
  • Emmanuelle Bouissou-Cadio
    • 1
    • 2
    • 3
    • 4
  • Julien Coutier
    • 1
    • 2
    • 3
    • 4
  • Michèle T. Martin
    • 1
    • 2
    • 3
    • 4
  1. 1.Laboratoire de Génomique et Radiobiologie de la KératinopoïèseCEA/DRF/IBFJ/IRCMParisFrance
  2. 2.INSERM U967ParisFrance
  3. 3.Université Paris-DiderotParisFrance
  4. 4.Université Paris-SaclayParisFrance

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