DNA demethylation at specific CpG sites in the IL1B promoter in response to inflammatory cytokines in human articular chondrocytes

Hashimoto, Ko, Oreffo, Richard O.C., Gibson, Marc B., Goldring, Mary B. and Roach, Helmstrud I. (2009) DNA demethylation at specific CpG sites in the IL1B promoter in response to inflammatory cytokines in human articular chondrocytes. Arthritis and Rheumatism, 60, (11), 3303-3313. (doi:10.1002/art.24882). (PMID:19877066).


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Objective: to determine whether changes in the DNA methylation status in the promoter region of the gene encoding interleukin-1? (IL-1?) account for expression of IL1B mRNA after long-term treatment of human articular chondrocytes with inflammatory cytokines.

Methods: IL-1? or TNF? combined with oncostatin M (OSM), or 5-aza-deoxycytidine (5-aza- dC) were added twice weekly for 4-5 weeks to primary cultures of normal human articular chondrocytes, obtained from patients with a femoral neck fracture. Expression of MMP13, IL1B??TNFA and DNMT1 were determined by SybrGreen-based qRT-PCR on genomic DNA and total RNA extracted from the same sample before and after culture. Bisulfite modification was used to identify which CpG sites in the IL1B promoter showed differential methylation between expressing and non-expressing cells. The percentages of cells that were methylated at that critical CpG site (-299bp) were quantified by a method that depended on methylation-sensitive restriction enzymes and real-time PCR. Secretion of IL-1? was assessed by enzyme-linked immunoabsorbent assay (ELISA) of the culture media.

Results: healthy chondrocytes did not express IL1B mRNA, but the levels were increased 5-fold by 5-aza-dC and 100- to 1000-fold by or TNF?/OSM. The % CpG methylation was decreased by 5-aza-dC, but reduced considerably more by IL-1? and almost abolished by TNF?/OSM. The mRNA was translated into protein in cytokine-treated chondrocytes.

Conclusion: these novel findings indicate that inflammatory cytokines can change DNA methylation status at key CpG sites, resulting in long-term induction of IL1B in human articular chondrocytes.

Item Type: Article
Digital Object Identifier (DOI): doi:10.1002/art.24882
Additional Information: This paper was selected by the Faculty of 1000 Medicine (www.f1000medicine.com), which is a literature awareness service that identifies and evaluates the most important articles published in Medicine based on the recommendations of a Faculty of over 2000 peer-nominated leading researchers and clinicians. The article's identification and inclusion provides recognition of its scientific merit and the positive contribution it makes to the medical literature.
ISSNs: 0004-3591 (print)
Keywords: epigenetics, DNA methylation, osteoarthritis, interleukin-1
Subjects: Q Science > Q Science (General)
R Medicine > R Medicine (General)
R Medicine > RZ Other systems of medicine
Divisions : University Structure - Pre August 2011 > School of Medicine > Developmental Origins of Health and Disease
ePrint ID: 72820
Accepted Date and Publication Date:
November 2009Published
Date Deposited: 23 Feb 2010
Last Modified: 31 Mar 2016 13:05
URI: http://eprints.soton.ac.uk/id/eprint/72820

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