Inhibition of lipopolysaccharide-induced cyclooxygenase-2 transcription by 6-(methylsulfinyl) hexyl isothiocyanate, a chemopreventive compound from Wasabia japonica (Miq.) Matsumura, in mouse macrophages

Abstract

6-(Methylsulfinyl)hexyl isothiocyanate (6-MITC) is a chemopreventive compound occurring in Wasabi ( Wasabia japonica (Miq.) Matsumura), which is a very popular pungent spice in Japan. We investigated the effects of 6-MITC on the expression of cyclooxygenase-2 (COX-2) in lipopolysaccharide (LPS)-activated murine macrophage RAW264 cells. Treatment with 6-MITC suppressed LPS-mediated induction of COX-2 protein in a dose-dependent manner. Transfections with various COX-2 promoter reporter constructs revealed that the inhibitory effects of 6-MITC on COX-2 gene expression were directed by the core promoter elements including nuclear factor κB (NF-κB), CCAAT/enhancer-binding protein (C/EBP) and cyclic AMP-response element (CRE) sites. Western blotting analysis showed that 6-MITC inhibited LPS-induced activation of MAPK (ERK, p38 kinase and JNK) and transcriptional factors (CREB, c-Jun and C/EBPδ) binding the core elements of COX-2 promoter, substantiating the involvement of these signal transduction pathways in the regulation of COX-2 expression by 6-MITC. Moreover, Western blotting experiments with MAPK-specific inhibitors (U0126 for MEK1/2, SB203580 for p38 kinase and SP600125 for JNK) demonstrated that 6-MITC suppressed LPS-induced COX-2 expression by blocking the activation of JNK-mediated AP-1 and ERK/p38 kinase-mediated CREB or C/EBPδ. Finally, the structure–activity study revealed that the inhibitory potency of methylsulfinyl isothiocyanates (MITCs) depended on the methyl chain length. These findings demonstrate for the first time that 6-MITC is an effective agent to attenuate COX-2 production, and enhance our understanding of the anti-inflammation properties of 6-MITC.