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THU0010 Transcriptional profiling of rheumatoid arthritis fibroblast-like synoviocytes identifies a subset of inflammatory genes with au-rich 3’-untranslated regions which are suppressed by histone deacetylase inhibitors via reduction of MRNA stability
  1. A.M. Grabiec1,2,
  2. A.M. Willemsen3,
  3. A.H.C. van Kampen3,
  4. P.P. Tak1,
  5. K.A. Reedquist1,2
  1. 1Division of Clinical Immunology and Rheumatology
  2. 2Department of Experimental Immunology
  3. 3Bioinformatics Laboratory, Department of Clinical Epidemiology, Biostatistics and Bioinformatics, Academic Medical Center, University of Amsterdam, Amsterdam, Netherlands


Background The production of inflammatory cytokines is tightly regulated by epigenetic and non-epigenetic mechanisms, among which acetylation and deacetylation of histones and non-histone proteins plays a prominent role. Histone deacetylase (HDAC) inhibitors (HDACi) suppress cytokine production by rheumatoid arthritis (RA) patient immune and stromal cells, and are potent therapeutics in animal arthritis models. Recently, the HDACi ITF2357 has demonstrated initial clinical efficacy in the treatment of systemic onset juvenile idiopathic arthritis. In our previous work we identified acceleration of mRNA degradation as a mechanism contributing to HDACi suppression of IL-6 production.

Objectives This study was undertaken to characterize the role of reduced mRNA stability in HDACi-mediated regulation of RA stromal synovial cell inflammatory activation.

Methods RA fibroblast-like synoviocytes (FLS) were treated with IL-1β in the presence or absence of 100-250 nM ITF2357 and total RNA was extracted. mRNA expression and stability of IL-1β-responsive genes was analyzed using a low density quantitative PCR array system. The frequencies of AU-rich elements (AREs) in the 3’-untranslated regions (UTRs) of the analyzed genes were determined using R Scripts (Bioconductor). Production of IL-8, MMP-1, MMP-3 and TIMP-1 was measured by ELISA.

Results FLS exposure to ITF2357 significantly reduced the mRNA levels of 85% of genes induced by IL-1β, including cytokines (IL-6, TNFα, IL-1α, IL-1β), chemokines (CCL2, IL-8, CXCL2, CXCL3, CXCL6, CXCL9-11), matrix-degrading enzymes (MMP-1, MMP-3, MMP-13), and intracellular molecules regulating cellular inflammatory responses (COX-2, IRAK2, NFKB1, IRF1). At the same time ITF2357 failed to modulate expression of genes non-responsive to IL-1β. Consistent with mRNA expression data, ITF2357 dose-dependently suppressed protein secretion of IL-8, MMP-1 and MMP-3, without affecting TIMP-1 production. Analyses of mRNA stability confirmed accelerated decay of IL-6 mRNA after FLS treatment with HDACi, and identified a number of other transcripts, including IL-8, COX-2, CXCL2, Bcl-XL and ADAMTS1, which are regulated by HDACi in a similar fashion. 3’ UTRs of these transcripts were characterized by significantly higher frequencies of AREs than other HDACi-sensitive genes in RA FLS. In contrast, mRNA kinetics experiments revealed other sets of genes, including MMPs, CXCL9-11, IL-1β, and E-selectin, which were suppressed by HDACi independently of effects on mRNA degradation.

Conclusions We demonstrate that HDACi prevent induction of the vast majority of IL-1β -inducible genes in RA FLS. HDACi suppress a subset of these genes via the acceleration of mRNA degradation, identifying this as an important general mechanism by which HDACi mediate their anti-inflammatory effects. However, our analyses also reveal distinct mechanisms by which HDACi can also regulate gene expression in RA FLS, awaiting further molecular characterization. Our findings provide additional evidence that therapies targeting HDAC activity may be useful in suppressing inflammation in RA.

Disclosure of Interest None Declared

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