An improved, standardised protocol for the isolation, enrichment and targeted neural differentiation of Nestin+ progenitors from adult human dermis

Exp Dermatol. 2010 Jun;19(6):549-55. doi: 10.1111/j.1600-0625.2009.01041.x. Epub 2010 Jan 21.

Abstract

Human skin-derived Nestin+ cells serve as a convenient source for autologous, adult, pluripotent progenitor cells that offer new therapeutic possibilities in cell-based regenerative medicine. However, the isolation of human Nestin+ cells has tended to be of very low efficiency and to produce highly variable cell yields. Here we report a standardised protocol that facilitates the isolation and enrichment of Nestin+ progenitor cells from enzymatically digested adult human scalp dermis. The use of distinct media like Dulbecco's modified Eagle medium supplemented with foetal bovine serum or, alternatively, serum-free, supplemented neural stem cell medium greatly affected cell morphology, proliferation and differentiation (e.g. towards a neural versus mesenchymal phenotype). Finally, Nestin+ cells were isolated from a heterogeneous dermis-derived progenitor cell population, which proliferates within clones or floating microspheres under defined serum-free culture conditions. Supplementation of the medium with epidermal growth factor and basic fibroblast growth factor as well as coating with fibronectin allowed the highest enrichment level of Nestin+ progenitors and differentiation towards neural fate. These methodological advances should greatly facilitate the isolation, culture and targeted differentiation of primary, adult human scalp skin dermis-derived Nestin+ cells.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Actins / metabolism
  • Adult
  • Adult Stem Cells / cytology*
  • Adult Stem Cells / drug effects
  • Adult Stem Cells / metabolism*
  • Aged
  • Cell Culture Techniques / methods*
  • Cell Differentiation / drug effects*
  • Cell Proliferation / drug effects
  • Cell Separation / methods*
  • Culture Media / pharmacology
  • Culture Media, Serum-Free / pharmacology
  • Dermis / cytology*
  • Female
  • Fibronectins / pharmacology
  • Gene Expression / genetics
  • Humans
  • Intermediate Filament Proteins / metabolism*
  • Male
  • Mesoderm / cytology
  • Middle Aged
  • Nerve Tissue Proteins / metabolism*
  • Nestin
  • Neurofilament Proteins / genetics
  • Neurofilament Proteins / metabolism
  • Neurons / cytology*
  • Neurons / metabolism

Substances

  • Actins
  • Culture Media
  • Culture Media, Serum-Free
  • Fibronectins
  • Intermediate Filament Proteins
  • NES protein, human
  • Nerve Tissue Proteins
  • Nestin
  • Neurofilament Proteins