Epigenetic regulation of TIMP-3 expression in human macrophages via physiological and pharmacological modification of histone3 lysine27 (H3K27).

BACKGROUND

Tissue inhibitor of metalloproteinases-3 (TIMP-3) prevents over-activity of metalloproteinases, thereby reducing the severity of several immune/ inflammatory diseases in which T-lymphocyte-stimulated macrophage invasion plays an essential role. However, the underlying mechanism of TIMP-3 expression and its modulation by anti-inflammatory agents are poorly understood.

METHODS AND RESULTS

Using RT-qPCR measurements in human macrophages, we showed that TIMP-3 steady-state and pre-spliced mRNAs are induced by the Thelper2 cytokine, interleukin-4 (IL-4) but downregulated by the pro-inflammatory Thelper1 cytokine, interferon-γ (IFN-γ), an inhibitory effect confirmed at the protein level. These effects of IL-4 and IFN-γ were independent of changes in the lineage determining transcription factors, CEBP-β, SP-1 and PU1/ETS, suggesting that epigenetic regulation might be important. We investigated the action of GSK-J4, an inhibitor of the histone3 lysine27 (H3K27) demethylase (KDM6) enzymes. GSK-J4 decreased basal and IL-4 induced TIMP-3 levels, concomitant with increased H3K27 tri-methylation (H3K27me) by Chromatin Immunoprecipitation. Moreover, using adenovirus-mediated silencing, TIMP-3 expression was found to be specifically regulated by KDM6B. GSK-J4 had no effect on IFN-γ induced inhibition of TIMP-3 mRNA, which was instead accompanied by a fall in H3K27 acetylation at the TIMP-3 promoter. The histone deacetylase (HDAC) inhibitor, MS-275, completely reversed this fall and partially restored TIMP-3 expression.

CONCLUSIONS

In conclusion GSK-J4 reduces TIMP-3 expression and might therefore paradoxically increase macrophage invasion. By contrast, the HDAC inhibitor MS-275 antagonised the depression of TIMP-3 expression by IFN-γ, which should reduce macrophage invasion.