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Human embryonic stem cells derived keratinocyte as an in vitro research model for the study of immune response
Author(s) -
Kidwai Fahad Karim,
Jokhun Doorgesh Sharma,
Movahednia Mohammad Mehdi,
Yeo Jin Fei,
Tan Kai Soo,
Cao Tong
Publication year - 2013
Publication title -
journal of oral pathology and medicine
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.887
H-Index - 83
eISSN - 1600-0714
pISSN - 0904-2512
DOI - 10.1111/jop.12054
Subject(s) - keratinocyte , keratinocyte growth factor , biology , cell culture , innate immune system , microbiology and biotechnology , tumor necrosis factor alpha , immune system , immunology , receptor , growth factor , biochemistry , genetics
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 (h ESC s) derived keratinocytes ( H 9‐ K ert) 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 (i NOS ) and tumour necrosis factor‐alpha ( TNF ‐α), in H 9‐ K ert and immortalized human keratinocyte cell line ( H a C a T ) were analysed at m RNA 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 ( NF ĸ B ) was assayed in these cells by transiently transfecting the cells with NF ĸ B reporter plasmid. Activation of NF ĸ B following treatment with heat‐killed P orphyromonas gingivalis ( P . gingivalis) , an oral pathogen, was determined by assaying for the reporter, secreted alkaline phosphatase activity. Results The expression of TLR s, cytokines and activation of NF ĸ B following bacterial stimulation showed in both H 9‐ K ert and the widely used H a C a T keratinocyte cell line was similar. Conclusion Overall, our results support the potential application of h ESC s 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.