Chlorophyllin suppresses interleukin-1 beta expression in lipopolysaccharide-activated RAW 264.7 cells

Abstract

Our previous findings demonstrated that chlorophyllin (CHL) inhibits inducible nitric oxide gene expression in macrophages. In the present study, we show that CHL inhibited IL-1beta production and its mRNA expression in a lipopolysaccharide (LPS)-stimulated murine macrophage cell-line, RAW 264.7. The inhibitory effect of CHL on IL-1beta gene expression was further supported by an in vitro transfection assay using a pIL-1(870 bp)-CAT construct, where CHL inhibited the activation of the IL-1beta promoter. Furthermore, CHL attenuated the activation of NF-kappaB, NF-IL6 and AP-1, which are known to be responsible for IL-1beta gene expression, as determined by an electrophoretic mobility shift assay and an in vitro transfection assay using p(NF-kappaB)3-CAT, p(NF-IL6)3-CAT, and p(AP-1)3-CAT, respectively. However, it was evident that the inhibitory activity of CHL on IL-1beta expression in the LPS-stimulated macrophages was independent of CRE/ATF. The immunoblot experiment demonstrated that CHL also caused a substantial decrease in the phosphorylation of p38 MAP kinase in LPS-stimulated RAW 264.7. These results suggest that CHL inhibits IL-1beta production in macrophages stimulated with LPS at transcriptional level by blocking the phosphorylation of p38 and by suppressing the activation of transcription factors, NF-kappaB, NF-IL6, and AP-1.