Helioxanthin inhibits interleukin-1β-induced MIP-1β production by reduction of c-jun expression and binding of the c-jun/CREB1 complex to the AP-1/CRE site of the MIP-1β promoter in Huh7 cells

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An elevated level of macrophage inflammatory protein-1β (MIP-1β) induced by IL-1β has been correlated with chronic hepatic inflammatory disease. However, molecular mechanism of IL-1β-induced MIP-1β expression in hepatic cells is obscure. Previously, we reported the mechanism of the anti-hepatitis B virus (HBV) activity of helioxanthin (HE-145). Here, we demonstrated that HE-145 inhibited IL-1β-induced MIP-1β expression in a dose-dependent manner in Huh7 cells. To understand the mode of action of HE-145, we first examined how IL-1β induced MIP-1β expression at the molecular level. Using selective inhibitors, we found that JNK and p38 pathways participated in IL-1β-induced MIP-1β expression. HE-145 specifically suppressed IL-1β-induced c-jun mRNA and protein expression and prevented c-jun-mediated AP-1 DNA-binding activity, whereas it had no effect on IL-1β-induced activation of JNK, p38 and ATF2. Further studies indicated that HE-145 may downregulate c-jun mRNA expression directly at transcriptional level without requirement of de novo protein synthesis. Mutational analysis and supershift assays indicated that IL-1β stimulated c-jun and CREB1 binding to the essential AP-1/CRE site of the MIP-1β promoter. The inhibitory effect of HE-145 on IL-1β-induced MIP-1β promoter activity was completely reversed by overexpressing c-jun. Electrophoretic mobility shift assay (EMSA) and chromatin immunoprecipitation (ChIP) assay consistently revealed that HE-145 reduced c-jun binding to the AP-1/CRE site in vitro and in vivo. Our results established a major role for c-jun in IL-1β-induced MIP-1β expression in hepatic cells. The reduction in IL-1β-induced c-jun expression and subsequent binding of the c-jun/CREB1 complex to AP-1/CRE site mainly contributed to the inhibitory action of HE-145 on IL-1β-induced MIP-1β production.

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