title: Various members of the Toll-like receptor family contribute to the innate immune response of human epidermal keratinocytes.
abstract: Toll-like receptors (TLRs) are important pattern recognition molecules that activate the nuclear factor (NF)-kappaB pathway leading to the production of antimicrobial immune mediators. As keratinocytes represent the first barrier against exogenous pathogens in human skin, we investigated their complete functional TLR1-10 expression profile. First, reverse transcription-polymerase chain reaction (PCR) analysis revealed a very similar pattern of TLR mRNA expression when comparing freshly isolated human epidermis and cultured primary human keratinocytes. Thus, further experiments were carried out with primary keratinocytes in comparison with the spontaneously immortalized human keratinocyte cell line HaCaT. The quantitative expression of TLR1-10 mRNA in real-time PCR of primary human keratinocytes and HaCaT cells was analysed. Both cell types constitutively expressed TLR2, TLR3, TLR5, and to a lesser extent TLR10. TLR4 was only found in HaCaT cells, TLR1 to a higher degree in primary keratinocytes. In line with this, LPS induced mRNA expression of CD14 and TLR4 only in HaCaT cells. After stimulation with various TLR ligands, the NF-kappaB-activated chemokine interleukin-8 (IL-8) was measured. In primary keratinocytes and HaCaT cells the TLR3 ligand poly (I:C) was the most potent stimulator of IL-8 secretion. The TLR ligands peptidoglycan, Pam3Cys and flagellin which bind to TLR2, TLR1/TLR2 heterodimer, and TLR5, respectively, also induced IL-8 secretion, whereas no IL-8 was induced by LPS, R-848, loxoribine and cytosine guanine dinucleotide-containing oligodeoxynucleotide. A corresponding pattern was found in the RelA NF-kappaB translocation assay after ligand stimulation of primary keratinocytes. These studies provide substantial evidence for a functional TLR expression and signalling profile of normal human keratinocytes contributing to the antimicrobial defence barrier of human skin.
authors.affiliation: Division of Environmental Dermatology and Allergy GSF/TUM, GSF National Research Center for Environment and Health, Neuherberg, Germany. koellisch@gsf.de
authors.forename: Gabriele
authors.initials: G
authors.lastname: Köllisch
authors.forename: Behnam Naderi
authors.initials: BN
authors.lastname: Kalali
authors.forename: Verena
authors.initials: V
authors.lastname: Voelcker
authors.forename: Reinhard
authors.initials: R
authors.lastname: Wallich
authors.forename: Heidrun
authors.initials: H
authors.lastname: Behrendt
authors.forename: Johannes
authors.initials: J
authors.lastname: Ring
authors.forename: Stefan
authors.initials: S
authors.lastname: Bauer
authors.forename: Thilo
authors.initials: T
authors.lastname: Jakob
authors.forename: Martin
authors.initials: M
authors.lastname: Mempel
authors.forename: Markus
authors.initials: M
authors.lastname: Ollert
mesh_terms: D001692:Biological Transport; D018929:Cell Culture Techniques; D002461:Cell Line, Transformed; D003713:Dendritic Cells / Q000187:drug effects / Q000276:immunology; D004817:Epidermis; D015870:Gene Expression; D006151:Guanosine / Q000031:analogs & derivatives* / Q000494:pharmacology; D006801:Humans; D007093:Imidazoles / Q000494:pharmacology; D016209:Interleukin-8 / Q000276:immunology; D015603:Keratinocytes / Q000187:drug effects / Q000276:immunology* / Q000382:microbiology; D008024:Ligands; D008070:Lipopolysaccharides / Q000494:pharmacology; D008562:Membrane Glycoproteins / Q000235:genetics / Q000378:metabolism*; D016328:NF-kappa B / Q000378:metabolism; D012333:RNA, Messenger / Q000032:analysis; D011956:Receptors, Cell Surface / Q000235:genetics / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013268:Stimulation, Chemical; D051194:Toll-Like Receptor 1; D051201:Toll-Like Receptor 10; D051195:Toll-Like Receptor 2; D051196:Toll-Like Receptor 3; D051197:Toll-Like Receptor 4; D051198:Toll-Like Receptor 5; D051193:Toll-Like Receptors
chemical_list: D007093:Imidazoles; D016209:Interleukin-8; D008024:Ligands; D008070:Lipopolysaccharides; D008562:Membrane Glycoproteins; D016328:NF-kappa B; D012333:RNA, Messenger; D011956:Receptors, Cell Surface; C495342:TLR2 protein, human; C495344:TLR3 protein, human; C495345:TLR4 protein, human; C495346:TLR5 protein, human; D051194:Toll-Like Receptor 1; D051201:Toll-Like Receptor 10; D051195:Toll-Like Receptor 2; D051196:Toll-Like Receptor 3; D051197:Toll-Like Receptor 4; D051198:Toll-Like Receptor 5; D051193:Toll-Like Receptors; D006151:Guanosine; C068562:loxoribine; C402365:resiquimod