To determine whether steroids inhibit the production of inflammatory cytokines by the inhibition of nuclear factor kappaB (NF-kappaB) activation in fibroblast-like rheumatoid synoviocytes (FLSs) under inflammatory conditions, and to determine whether steroids stimulate the induction of synthesis of the inhibitory protein IkappaB-alpha in the anti-inflammatory immune response of these cells.
Expression of the interleukin-6 (IL-6) and interleukin-1beta (IL-1beta) genes was measured by semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR), and the secreted IL-6 was measured with the enzyme-linked immunosorbent assay. Inhibition of the NF-kappaB activation was examined with the electrophoretic mobility shift assay (EMSA). In order to study dexamethasone (DEX)-dependent regulation of IkappaB-alpha expression, we performed Western blotting before and after stimulation with tumour necrosis factor alpha (TNF-alpha).
The inflammatory cytokine study showed that DEX suppressed gene expression and the production of protein in FLSs. EMSA demonstrated that identical amounts of NF-kappaB were present in the nucleus of the FLSs stimulated by TNF-alpha, with or without pretreatment with DEX. Treatment of FLSs with DEX did not induce an increase in IkappaB-alpha sufficient to prevent nuclear translocation of NF-kappaB on stimulation with TNF-alpha.
DEX may suppress the production of inflammatory cytokines, such as IL-6 and IL-1beta, but it neither prevents the translocation of NF-kappaB to the nucleus nor induces the synthesis of IkappaB-alpha protein in FLSs stimulated by TNF-alpha.