Interleukin-6 promoter polymorphisms, chromatin remodelling and gene expression in a monocyte-derived macrophage model of inflammation.

lnterleukin-6 (IL-6) is a pro-inflammatory cytokine produced by macrophages, endothelial cells, fibroblasts, vascular smooth muscle cells, T lymphocytes and adipocytes. IL-6 is regulated at the level of transcription. There are multiple regulatory elements in the IL-6 promoter. Three single nucleotide polymorphisms at position -597, -572 and -174 and a variable region of A's and T's at -373 have been identified in the IL-6 proximal promoter. The THP-1 macrophage-like cell line was induced to express IL-6 mRNA in response to IFN-? and LPS. Interferon-gamma induced IL-6 mRNA expression was inhibited by treatment with either atorvastatin or pravastatin. Monocytes extracted from the blood of healthy volunteers of known IL-6 promoter haplotype (+GG9/11G and -GG9/11G) were differentiated to macrophages ex vivo. These cells were cultured in the presence of IL-1beta, LPS or IFN-gamma, to induce IL-6 mRNA. Quantitative PCR was used to quantitate the level of IL-6 mRNA. IL-6 mRNA in macrophages from individuals with the IL-6 promoter haplotype +GG9/11G was significantly induced in response to IL-1beta when compared to -GG9/11G individuals. A similar though non significant effect was detected in response to IFN-gamma. No significant IL-6 promoter haplotype-specific difference was observed in response to LPS. Chromatin remodeling of the IL-6 promoter occurred within 30 minutes of stimulation and was specific to the region of the IL-6 promoter and the stimulus applied to the cells. A DNase I hypersensitive site was identified in the region of the polymorphic AnTn variable tract and a potential DNA cruciform structure in the IL-6 promoter was resolved by T7 Endonuclease I. Atomic force microscopy was used to visualise DNA cruciform structure in IL- 6 promoter haplotype constructs. This work contributes to the understanding of the molecular mechanisms underlying the normal inter-individual differences in the macrophage IL-6 inflammatory response