title: Human embryonic stem cells derived keratinocyte as an in vitro research model for the study of immune response.
abstract: BACKGROUND
  The innate immune response (IMR) is critical for the oral mucosa due to their continuous exposure to various oral pathogens. Keratinocytes play important role in IMR. Therefore, to date, keratinocytes from different sources have been used as in vitro research model for the study of IMR. However, current keratinocyte research models are hampered by the limited supply, patients' dependency and batch to batch variation. Therefore, in this study, we demonstrated the use of human embryonic stem cells (hESCs) derived keratinocytes (H9-Kert) as an alternative research model for the study of IMR.
  
  
  METHODS
  The expression kinetics of toll-like receptor (TLR) 2, TLR 4, interleukin (IL) -6, IL-8, inducible nitric oxide synthase (iNOS) and tumour necrosis factor-alpha (TNF-α), in H9-Kert and immortalized human keratinocyte cell line (HaCaT) were analysed at mRNA levels by both reverse transcription polymerase chain reaction (RT-PCR) and quantitative real-time RT-PCR. The activation of the inflammatory transcription factor nuclear factor kappa-b (NFĸB) was assayed in these cells by transiently transfecting the cells with NFĸB reporter plasmid. Activation of NFĸB following treatment with heat-killed Porphyromonas gingivalis (P. gingivalis), an oral pathogen, was determined by assaying for the reporter, secreted alkaline phosphatase activity.
  
  
  RESULTS
  The expression of TLRs, cytokines and activation of NFĸB following bacterial stimulation showed in both H9-Kert and the widely used HaCaT keratinocyte cell line was similar.
  
  
  CONCLUSIONS
  Overall, our results support the potential application of hESCs as an alternative limitless cell source for primary keratinocytes which can be used as consistent and dependable research tool with minimum variations and no donor's dependency.
authors: Oral Sciences Disciplines, Faculty of Dentistry, National University of Singapore, Singapore.
authors: Fahad Karim
authors: FK
authors: Kidwai
authors: Doorgesh Sharma
authors: DS
authors: Jokhun
authors: Mohammad Mehdi
authors: MM
authors: Movahednia
authors: Jin Fei
authors: JF
authors: Yeo
authors: Kai Soo
authors: KS
authors: Tan
authors: Tong
authors: T
authors: Cao
mesh_terms: D000469:Alkaline Phosphatase / Q000032:analysis; D002460:Cell Line; D019070:Cell Lineage; D003470:Culture Media; D053595:Embryonic Stem Cells / Q000502:physiology*; D005347:Fibroblasts / Q000166:cytology; D005822:Genetic Vectors / Q000235:genetics; D006801:Humans; D007113:Immunity, Innate / Q000276:immunology*; D015850:Interleukin-6 / Q000032:analysis; D016209:Interleukin-8 / Q000032:analysis; D015603:Keratinocytes / Q000166:cytology / Q000276:immunology*; D016328:NF-kappa B / Q000032:analysis; D052247:Nitric Oxide Synthase Type II / Q000032:analysis; D016966:Porphyromonas gingivalis / Q000276:immunology; D060888:Real-Time Polymerase Chain Reaction; D020133:Reverse Transcriptase Polymerase Chain Reaction; D051195:Toll-Like Receptor 2 / Q000032:analysis; D051197:Toll-Like Receptor 4 / Q000032:analysis; D014409:Tumor Necrosis Factor-alpha / Q000032:analysis
chemical_list: C508606:CXCL8 protein, human; D003470:Culture Media; C508600:IL6 protein, human; D015850:Interleukin-6; D016209:Interleukin-8; D016328:NF-kappa B; C495342:TLR2 protein, human; C495345:TLR4 protein, human; D051195:Toll-Like Receptor 2; D051197:Toll-Like Receptor 4; D014409:Tumor Necrosis Factor-alpha; C496319:NOS2 protein, human; D052247:Nitric Oxide Synthase Type II; D000469:Alkaline Phosphatase
keywords: human embryonic stem cells; human embryonic stem cells derived keratinocyte; innate immunity; toll-like receptors