Human embryonic stem cells derived keratinocyte as an in vitro research model for the study of immune response

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. H...

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Bibliographic Details
Published in:Journal of oral pathology & medicine 2013-09, Vol.42 (8), p.627-634
Main Authors: Kidwai, Fahad Karim, Jokhun, Doorgesh Sharma, Movahednia, Mohammad Mehdi, Yeo, Jin Fei, Tan, Kai Soo, Cao, Tong
Format: Article
Language:English
Subjects:
Alkaline phosphatase
Alkaline Phosphatase - analysis
Biological and medical sciences
Cell Line
Cell Lineage
Culture Media
Dentistry
Embryonic Stem Cells - physiology
Fibroblasts - cytology
Genetic Vectors - genetics
human embryonic stem cells
human embryonic stem cells derived keratinocyte
Humans
Immunity, Innate - immunology
innate immunity
Interleukin-6 - analysis
Interleukin-8 - analysis
Keratinocytes - cytology
Keratinocytes - immunology
Medical sciences
NF-kappa B - analysis
Nitric Oxide Synthase Type II - analysis
Otorhinolaryngology. Stomatology
Porphyromonas gingivalis
Porphyromonas gingivalis - immunology
Real-Time Polymerase Chain Reaction
Reverse Transcriptase Polymerase Chain Reaction
Toll-Like Receptor 2 - analysis
Toll-Like Receptor 4 - analysis
toll-like receptors
Tumor Necrosis Factor-alpha - analysis
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Summary: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. Conclusion 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.
ISSN:0904-2512
1600-0714
DOI:10.1111/jop.12054