Activation of interleukin-6 gene expression through the NF-kappa B transcription factor.

The promoter region of the interleukin-6 (IL-6) gene has a putative NF-kappa B-binding site. We found that a fragment of the IL-6 promoter containing the site specifically binds highly purified NF-kappa B protein and the NF-kappa B protein in nuclear extracts of phorbol ester-induced Jurkat cells. Mutations of the NF-kappa B site abolished complex formation with both purified NF-kappa B and the nuclear extract protein. Transient expression of chloramphenicol acetyltransferase (CAT) plasmids containing the IL-6 promoter revealed very little activity of the promoter in U-937 monocytic cells and in HeLa cells before stimulation. However, stimulation of U-937 and HeLa cells by inducers of NF-kappa B led to a dramatic increase in CAT activity. Mutations in the NF-kappa B-binding site abolished inducibility of IL-6 promoter-cat constructs in U-937 cells by lipopolysaccharide, tumor necrosis factor alpha, the double-stranded RNA poly(IC), or phytohemagglutinin and in HeLa cells by tumor necrosis factor alpha and drastically reduced but did not completely eliminate inducibility in HeLa cells stimulated by double-stranded RNA poly(IC) or phorbol 12-myristate 13-acetate. These results suggest that NF-kappa B is an important mediator for activation of the IL-6 gene by a variety of IL-6 inducers in both U-937 and HeLa cells and that alternative inducible enhancer elements contribute in a cell-specific manner to IL-6 gene induction. Because NF-kappa B is involved in the control of a variety of genes activated upon inflammation, NF-kappa B may play a central role in the inflammatory response to infection and tissue injury.     

Matrix metalloproteinase (MMP)-12 regulates MMP-9 expression in interleukin-1beta-treated articular chondrocytes

Limited information is available on the expression and role of matrix metalloproteinase (MMP)-12 in chondrocytes. We characterized the expression mechanism of MMP-12 and possible function in chondrocytes. Interleukin (IL)-1beta induced the expression and activation of MMP-12 in primary culture chondrocytes and cartilage explants via mitogen-activated protein (MAP) kinase signaling pathways. Among MAP kinases, extracellular signal-regulated kinase and p38 kinase are necessary for MMP-12 expression, whereas c-jun N-terminal kinase is required for the activation of MMP-12. The possibility that MMP-12 acts as a modulator of other MMP was examined. MMP-12 alone did not affect other MMP expressions. However, MMP-12 enhanced expression and activation of MMP-9 in the presence of IL-1beta. Our results indicate that IL-1beta in chondrocytes induces the expression and activation of MMP-12, which, in turn, augments MMP-9 expression and activation.     

Lycopene inhibits matrix metalloproteinase-9 expression and down-regulates the binding activity of nuclear factor-kappa B and stimulatory protein-1

The carotenoid lycopene has been associated with decreased risks of several types of cancer, such as hepatoma. Although lycopene has been shown to inhibit metastasis, its mechanism of action is poorly understood. Here, we used SK-Hep-1 cells (from a human hepatoma) to test whether lycopene exerts its anti-invasion activity via down-regulation of the expression of matrix metalloproteinase (MMP)-9, an important enzyme in the degradation of basement membrane in cancer invasion. The activity and expressions of MMP-9 protein and mRNA were detected by gelatin zymography, Western blotting and RT-PCR, respectively. The binding abilities of nuclear factor-kappa B (NF-kappaB), activator protein-1 and stimulatory protein-1 (Sp1) to the binding sites in the MMP-9 promoter were measured by the electrophoretic mobility shift assay. We showed that lycopene (1-10 microM) significantly inhibited SK-Hep-1 invasion (P<.05) and that this effect correlated with the inhibition of MMP-9 at the levels of enzyme activity (r(2)=.94, P<.001), protein expression (r(2)=.80, P=.007) and mRNA expression (r(2)=.94, P<.001). Lycopene also significantly inhibited the binding abilities of NF-kappaB and Sp1 and decreased, to some extent, the expression of insulin-like growth factor-1 receptor (IGF-1R) and the intracellular level of reactive oxygen species (P<.05). The antioxidant effect of lycopene appeared to play a minor role in its inhibition of MMP-9 and invasion activity of SK-Hep-1 cells because coincubation of cells with lycopene plus hydrogen peroxide abolished the antioxidant effect but did not significantly affect the anti-invasion ability of lycopene. Thus, lycopene decreases the invasive ability of SK-Hep-1 cells by inhibiting MMP-9 expression and suppressing the binding activity of NF-kappaB and Sp1. These effects of lycopene may be related to the down-regulation of IGF-1R, while the antioxidant activity of lycopene appears to play a minor role.     

Caffeic acid phenethyl ester inhibits invasion and expression of matrix metalloproteinase in SK-Hep1 human hepatocellular carcinoma cells by targeting nuclear factor kappa B

Numerous studies have shown that the levels of matrix metalloproteinase (MMP)-2 and/or MMP-9 are associated with the invasive phenotypes of cancer cells. This study investigated the effects of caffeic acid phenethyl ester (CAPE), a chemopreventive phytochemical derived from honeybee propolis, on the invasive phenotype of SK-Hep1 human hepatocellular carcinoma cells (SK-Hep1 cells). CAPE effectively suppressed SK-Hep1 cell invasion in a dose-dependent manner. The constitutive expression of MMP-2 and MMP-9 in SK-Hep1 cells was almost completely abolished by treatment with 12.5 muM CAPE. CAPE also significantly inhibited nuclear factor kappa B (NF-kappaB) DNA-binding activity in SK-Hep1 cells. These results taken together suggest that CAPE exerts antimetastatic potential through inhibition of MMP-2 and MMP-9 expression, possibly by targeting NF-kappaB in hepatocellular carcinoma.