Nonviral in situ green fluorescent protein labeling and culture of primary, adult human hair follicle epithelial progenitor cells.
In this article we show that cloning of the human K15 promoter before a green fluorescence protein (GFP)/geneticin-resistance cassette and transfection of microdissected, organ-cultured adult human scalp hair follicles generates specific K15 promoter-driven GFP expression in their stem cell-rich bulge region. K15-GFP+ cells can be visualized in situ by GFP fluorescence and 2-photon laser scanning microscopy. Vital K15-GFP+ progenitor cells can then be selected by using the criteria of their green fluorescence, adhesion to collagen type IV and fibronectin, and geneticin resistance. Propagated K15-GFP+ cells express epithelial progenitor markers, show the expected differential gene expression profile of human bulge epithelium, and form holoclones. This application of nonretroviral, K15 promoter-driven, GFP labeling to adult human hair follicles facilitates the characterization and manipulation of human epithelial stem cells, both in situ and in vitro, and should be transferable to other complex human tissues.
Tiede
Stephan
S
Department of Dermatology, University of Lübeck, D-23538 Lübeck, Germany. stephan.tiede@uk-sh.de
Koop
Norbert
N
Kloepper
Jennifer E
JE
Fässler
Reinhard
R
Paus
Ralf
R
D017209:Apoptosis / Q000235:genetics / Q000502:physiology; D049109:Cell Proliferation; D002470:Cell Survival / Q000235:genetics / Q000502:physiology; D002478:Cells, Cultured; D004847:Epithelial Cells / Q000166:cytology* / Q000378:metabolism*; D005455:Fluorescent Antibody Technique; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism*; D018859:Hair Follicle / Q000166:cytology*; D006801:Humans; D007150:Immunohistochemistry; D053569:Keratin-15 / Q000235:genetics / Q000378:metabolism; D020411:Oligonucleotide Array Sequence Analysis; D016133:Polymerase Chain Reaction; D011401:Promoter Regions, Genetic / Q000235:genetics; D013234:Stem Cells / Q000166:cytology* / Q000378:metabolism*
D053569:Keratin-15; D049452:Green Fluorescent Proteins