Anti-inflammatory mediators ST2 and SIGIRR are induced by diphenyldifluoroketone EF24 in lipopolysaccharide-stimulated dendritic cells.

The objective of this study was to investigate the effect of EF24, an NF-κB-inhibitor, on the expression of negative regulators in IL-1R pathway, namely ST2 and SIGIRR. Murine JAWS II dendritic cells (DC) were challenged with lipopolysaccharide (LPS, 100 ng/ml) for 4 h, followed by treatment with 10 μM EF24 for 1 h. ST2 and SIGIRR expression was monitored by qRT-PCR and immunoblotting. ST2L and MyD88 interaction was studied by co-immunoprecipitation, and IL-33, a ST2L ligand, was assayed by ELISA. Activation of transcription factor SP1 was examined by confocal microscopy, immunoblotting, and EMSA. The effect of EF24 on accumulation of ubiquitinated proteins in DCs and proteolysis of fluorogenic peptides by purified proteasome was studied. We found that EF24 upregulated the expression of ST2 and SIGIRR and decreased the interaction of the membrane-bound ST2 (ST2L) with MyD88, and significantly reduced IL-33 levels in LPS-stimulated DCs. Simultaneously it increased the activation of transcription factor SP1and restored the basal level of ubiquitinated proteins in LPS-stimulated DCs. Moreover, EF24 inhibited trypsin- and chymotrypsin-like activity of proteasome by directly interacting with 26S proteasome. The results suggest that EF24 activates endogenous anti-inflammatory arm of IL-1R signaling, most likely by stabilizing SP1 against proteasomal degradation.