Differentiating the Stem Cell Pool of Human Hair Follicle Outer Root Sheath into Functional Melanocytes

  • Marie Schneider
  • Christina Dieckmann
  • Katrin Rabe
  • Jan-Christoph Simon
  • Vuk Savkovic
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1210)

Abstract

Bench-to-Bedside concepts for regenerative therapy place significant weight on noninvasive approaches, with harvesting of the starting material as a header. This is particularly important in autologous treatments, which use one’s bodily constituents for therapy. Precisely the stretch between obtaining therapeutic elements invasively and noninvasively places non-intrusive “sampling” rather than “biopsy” in the center of the road map of developing an autologous regenerative therapy. We focus on such a noninvasively available source of adult stem cells that we carry with us throughout our life, available at our fingertips—or shall we say hair roots, by a simple plucking of hair: the human hair follicle. This chapter describes an explant procedure for cultivating melanocytes differentiated from the stem cell pool of the hair follicle Outer Root Sheath (ORS). In vivo, the most abundant derivatives of the heterogeneous ORS stem cell pool are epidermal cells—melanocytes and keratinocytes which complete their differentiation—either spontaneously or upon picking up regenerative cues from damaged skin—and migrate from the ORS towards the adjacent regenerating area of the epidermis. We have taken advantage of the ORS developmental potential by optimizing explant primary culture, expansion and melanogenic differentiation of resident ORS stem cells towards end-stage melanocytes in order to obtain functional melanocytes noninvasively for the purposes of transplantation and use them for the treatment of depigmentation disorders. Our protocol specifies sampling of hair with their ORS, follicle medium–air interface primary culture, stimulation of cell outgrowth, adherent culture and differentiation of ORS stem cells and precursors towards fully functional melanocytes. Along with cultivation, we describe selection techniques for establishing and maintaining a pure melanocyte population and methods suitable for determining melanocyte identity.

Key words

Melanocytes Hair follicle Outer Root Sheath ORS Stem cells Skin regeneration Pigmentation Vitiligo Precursors Differentiation 

Notes

Acknowledgments

The work presented in this paper was made possible by funding from the German Federal Ministry of Education and Research (BMBF 1315883) and the German Research Council SFB TRR 67 project B3 to JCS. We are grateful to Dr. Michael Cross and Pavel Suba for valuable language remarks.

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Copyright information

© Springer Science+Business Media New York 2014

Authors and Affiliations

  • Marie Schneider
    • 1
  • Christina Dieckmann
    • 1
  • Katrin Rabe
    • 1
  • Jan-Christoph Simon
    • 2
  • Vuk Savkovic
    • 1
  1. 1.Translationszentrum für Regenerative MedizinUniversität LeipzigLeipzigGermany
  2. 2.Klinik und Poliklinik für Dermatologie, Venerologie und AllergologieUniversitätsklinikum LeipzigLeipzigGermany

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