15-Deoxy-Δ^12,14-prostaglandin J₂inhibits IL-1β-induced cyclooxygenase-2 expression in human synovial fibroblasts through a histone deacetylase-independent mechanism

Objective

15-Deoxy-Δ^12,14-prostaglandin J₂ (15d-PGJ₂) is a natural ligand for peroxisome proliferator-activated receptor gamma (PPARγ) and has been reported to inhibit the expression of a number of inflammatory genes in several cell types. However, its effects on cyclooxygenase-2 (COX-2) expression remains controversial. In the present study, we investigated the effects of 15d-PGJ₂ on IL-1β-induced COX-2 expression in human synovial fibroblasts.

Methods

COX-2 protein and mRNA expression were evaluated using western blotting and real-time PCR analysis, respectively. The COX-2 promoter activity was analyzed in transient transfection experiments. Chromatin immunoprecipitation assays were performed to evaluate the level of histone acetylation and the recruitment of HDAC1, HDAC2, HDAC3, and p300 to the COX-2 promoter.

Results

15d-PGJ₂ inhibited IL-1β-induced COX-2 protein and mRNA expression, as well as COX-2 gene promoter activation. The suppression of COX-2 protein expression was abrogated by the PPARγ antagonist, GW9662, suggesting that this effect is mediated by PPARγ. The induction of COX-2 by IL-1β is associated with hyperacetylation of histone H3 and H4 at the COX-2 promoter. Interestingly, 15d-PGJ₂ selectively blocked IL-1β-induced histone H3 acetylation. This reduction was demonstrated to not correlate with the recruitment of histone deacetylase (HDAC) to the COX-2 promoter. Also, treatment with the specific HDAC inhibitor, trichostatin A, did not relieve the suppressive effect of 15d-PGJ₂, indicating that HDACs are not involved in the inhibitory effect of15d-PGJ₂ on COX-2 expression. Furthermore, 15d-PGJ₂ blocked IL-1β-induced recruitment of the histone acetylase (HAT) p300 to the COX-2 promoter, which may be the mechanism for decreased histone H3 acetylation and COX-2 expression. In line with this, overexpression of p300, but not of a mutant p300 lacking HAT activity, relieved the inhibitory effect of 15d-PGJ2 on COX-₂ promoter activation.

Conclusion

Our data suggest that 15d-PGJ₂ can inhibit IL-1β-induced COX-2 expression in a PPARγ-dependent, HDAC-independent mechanism, probably by interfering with the HAT p300.