title: Human Dermal Fibroblasts Demonstrate Positive Immunostaining for Neuron- and Glia- Specific Proteins.
abstract: In stem cell cultures from adult human tissue, undesirable contamination with fibroblasts is frequently present. The presence of fibroblasts obscures the actual number of stem cells and may result in extracellular matrix production after transplantation. Identification of fibroblasts is difficult because of the lack of specific fibroblast markers. In our laboratory, we isolate and expand neural-crest-derived stem cells from human hair follicle bulges and investigate their potential to differentiate into neural cells. To establish cellular identities, we perform immunohistochemistry with antibodies specific for glial and neuronal markers, and use fibroblasts as negative control. We frequently observe that human adult dermal fibroblasts also express some glial and neuronal markers. In this study, we have sought to determine whether our observations represent actual expression of these markers or result from cross-reactivity. Immunohistochemistry was performed on human adult dermal fibroblasts using acknowledged glial and neuronal antibodies followed by verification of the data using RT-qPCR. Human adult dermal fibroblasts showed expression of the glia-specific markers SOX9, glial fibrillary acidic protein and EGR2 (KROX20) as well as for the neuron-specific marker class III β-tubulin, both at the protein and mRNA level. Furthermore, human adult dermal fibroblasts showed false-positive immunostaining for S100β and GAP43 and to a lower extent for OCT6. Our results indicate that immunophenotyping as a tool to determine cellular identity is not as reliable as generally assumed, especially since human adult dermal fibroblasts may be mistaken for neural cells, indicating that the ultimate proof of glial or neuronal identity can only be provided by their functionality.
authors.affiliation: Department of Otorhinolaryngology and Head & Neck Surgery, Leiden University Medical Center, Leiden, the Netherlands.
authors.forename: C J
authors.initials: CJ
authors.lastname: Janmaat
authors.affiliation: Percuros B.V., Enschede, the Netherlands.|Department of Radiology, Leiden University Medical Center, Leiden, the Netherlands.
authors.forename: K E
authors.initials: KE
authors.lastname: de Rooij
authors.affiliation: Department of Otorhinolaryngology and Head & Neck Surgery, Leiden University Medical Center, Leiden, the Netherlands.
authors.forename: H
authors.initials: H
authors.lastname: Locher
authors.affiliation: Department of Otorhinolaryngology and Head & Neck Surgery, Leiden University Medical Center, Leiden, the Netherlands.
authors.forename: S C
authors.initials: SC
authors.lastname: de Groot
authors.affiliation: Department of Otorhinolaryngology and Head & Neck Surgery, Leiden University Medical Center, Leiden, the Netherlands.
authors.forename: J C M J
authors.initials: JC
authors.lastname: de Groot
authors.affiliation: Department of Otorhinolaryngology and Head & Neck Surgery, Leiden University Medical Center, Leiden, the Netherlands.
authors.forename: J H M
authors.initials: JH
authors.lastname: Frijns
authors.affiliation: Department of Otorhinolaryngology and Head & Neck Surgery, Leiden University Medical Center, Leiden, the Netherlands.
authors.forename: M A
authors.initials: MA
authors.lastname: Huisman
mesh_terms: D000818:Animals; D015415:Biomarkers / Q000378:metabolism; D002478:Cells, Cultured; D051767:Early Growth Response Protein 2 / Q000235:genetics / Q000378:metabolism; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism*; D019922:GAP-43 Protein / Q000235:genetics / Q000378:metabolism; D006801:Humans; D007150:Immunohistochemistry; D051379:Mice; D009457:Neuroglia / Q000166:cytology / Q000378:metabolism*; D009474:Neurons / Q000166:cytology / Q000378:metabolism*; D027701:Organic Cation Transport Proteins / Q000235:genetics / Q000378:metabolism; D061251:Primary Cell Culture / Q000379:methods*; D009418:S100 Proteins / Q000235:genetics / Q000378:metabolism; D055758:SOX9 Transcription Factor / Q000235:genetics / Q000378:metabolism
chemical_list: D015415:Biomarkers; C497326:EGR2 protein, human; D051767:Early Growth Response Protein 2; D019922:GAP-43 Protein; D027701:Organic Cation Transport Proteins; D009418:S100 Proteins; C467737:SLC22A16 protein, human; D055758:SOX9 Transcription Factor; C527932:SOX9 protein, human