• Biomolecules & Therapeutics
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• v.21 no.2
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• pp.146-152
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• 2013

This study examined the total polyphenol content of eight wild edible plants from Ethiopia and their effect on NO production in Raw264.7 cells. Owing to its relatively high polyphenol concentration and inhibition of NO production, the methanol extract of Adansonia digitata L. leaf (MEAD) was subjected to detailed evaluation of its antioxidant and anti-inflammatory effects. Antioxidant effects were assessed by measuring free-radical-scavenging activity using 1,1-diphenyl-2-picrylhydrazyl (DPPH) and oxygen-radical-absorbance capacity (ORAC) assays, while anti-inflammatory effects were assessed by measuring inducible nitric oxide synthase (iNOS) expression in lipopolysaccharide (LPS)-stimulated RAW264.7 cells. In the ORAC assay, MEAD was 10.2 times more potent than vitamin C at eliminating peroxyl radicals. In DPPH assay, MEAD also showed a strong ROS scavenging effect. MEAD significantly inhibited iNOS activity ($IC_{50}=28.6{\mu}g/ml$) of LPS-stimulated Raw264.7 cells. We also investigated the relationship between iNOS expression and nuclear factor kappa B (NF-${\kappa}B$) activation. MEAD inhibited $I{\kappa}B{\alpha}$ degradation and NF-${\kappa}B$ translocation from the cytosol to the nucleus in LPS-induced RAW264.7 cells without significant cytotoxic effects, as confirmed by MTT assay. These results suggest that MEAD inhibits anti-inflammatory iNOS expression, which might be related to the elimination of peroxyl radicals and thus the inhibition of $I{\kappa}B{\alpha}$-mediated NF-${\kappa}B$ signal transduction.

• Molecules and Cells
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• v.39 no.5
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• pp.403-409
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• 2016

NME1 is a well-known metastasis suppressor which has been reported to be downregulated in some highly aggressive cancer cells. Although most studies have focused on NME1, the NME1 gene also encodes the protein (NME1L) containing N-terminal 25 extra amino acids by alternative splicing. According to previous studies, NME1L has potent anti-metastatic activity, in comparison with NME1, by interacting with $IKK{\beta}$ and regulating its activity. In the present study, we tried to define the role of the N-terminal 25 amino acids of NME1L in $NF-{\kappa}B$ activation signaling. Unfortunately, the sequence itself did not interact with $IKK{\beta}$, suggesting that it may be not enough to constitute the functional structure. Further construction of NME1L fragments and biochemical analysis revealed that N-terminal 84 residues constitute minimal structure for homodimerization, $IKK{\beta}$ interaction and regulation of $NF-{\kappa}B$ signaling. The inhibitory effect of the fragment on cancer cell migration and $NF-{\kappa}B$-stimulated gene expression was equivalent to that of whole NME1L. The data suggest that the N-terminal 84 residues may be a core region for the anti-metastatic activity of NME1L. Based on this result, further structural analysis of the binding between NME1L and $IKK{\beta}$ may help in understanding the anti-metastatic activity of NME1L and provide direction to NME1L and $IKK{\beta}$-related anti-cancer drug design.

• BMB Reports
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• v.50 no.2
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• pp.103-108
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• 2017

Heme oxygenase (HO-1) catalyzes heme to carbon monoxide (CO), biliverdin/bilirubin, and iron and is known to prevent the pathogenesis of several human diseases. We assessed the beneficial effect of heme degradation products on osteoclastogenesis induced by receptor activator of NF-${\kappa}B$ ligand (RANKL). Treatment of RAW264.7 cells with CORM-2 (a CO donor) and bilirubin, but not with iron, decreased RANKL-induced osteoclastogenesis, with CORM-2 having a more potent anti-osteogenic effect. CORM-2 also inhibited RANKL-induced osteoclastogenesis and osteoclastic resorption activity in marrow-derived macrophages. Treatment with hemin, a HO-1 inducer, strongly inhibited RANKL-induced osteoclastogenesis in wild-type macrophages, but was ineffective in $HO-1^{+/-}$ cells. CORM-2 reduced RANKL-induced NFATc1 expression by inhibiting IKK-dependent NF-${\kappa}B$ activation and reactive oxygen species production. These results suggest that CO potently inhibits RANKL-induced osteoclastogenesis by inhibiting redox-sensitive NF-${\kappa}B$-mediated NFATc1 expression. Our findings indicate that HO-1/CO can act as an anti-resorption agent and reduce bone loss by blocking osteoclast differentiation.

• Korean Journal of Pharmacognosy
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• v.45 no.1
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• pp.28-34
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• 2014

Rhei Rhizoma (RR) and Moutan cortex (MC) have been reported to have anti-inflammatory effects. However, little is known about the effects of RR and MC on endothelial inflammation and insulin resistance (IR). This study aims to investigate whether the water extracts of RR and MC could exert protection against palmitic acid (PA)-induced inflammation and IR in human umbilical vein endothelial cells (HUVECs). HUVECs were pretreated for 6 h with RR or MC, and then exposed to PA for 24 h. The levels of interleukin-6 (IL-6) and tumor necrosis factor-${\alpha}$ (TNF-${\alpha}$) were determined by enzyme-linked immunosorbant assay kits. Western blot analysis was performed for activation of nuclear factor-${\kappa}B$ (NF-${\kappa}B$) and insulin receptor substrate-1 (IRS-1). In HUVECs stimulated with PA, both RR and MC significantly inhibited the production of TNF-${\alpha}$ and IL-6 and the activation of NF-${\kappa}B$. At the same concentrations, the inhibitory effects of RR were more potent than those of MC. PA reduced insulin-induced phosphorylation of IRS-1, which was reversed by RR and MC. The results suggest that RR and MC are effective in inhibiting PA-associated endothelial inflammation and ameliorating IR by beneficial regulation of NF-${\kappa}B$ and IRS-1 activation.

• Journal of Microbiology and Biotechnology
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• v.18 no.1
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• pp.95-103
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• 2008

Mushrooms are regarded as one of the well-known foods and biopharmaceutical materials with a great deal of interest. Polysaccharide ${\beta}$-glucan is the major component of mushrooms that displays various biological activities such as antidiabetic, anticancer, and antihyperlipidemic effects. In this study, we compared the immunostimulatory potency of polysaccharide fractions, prepared from liquid culture of pine-mushroom Tricholoma matsutake, with a potent immunogen lipopolysaccharide (LPS), and their molecular mechanisms on the functional activation of macrophages. We found that fraction II (TMF-II) was able to comparably upregulate or highly enhance the phenotypic functions of macrophages such NO production and cytokine (IL-$1{\beta}$, IL-6, IL-12, and TNF-${\alpha}$) expression, to LPS. TMF-II triggered the phosphorylation of $I{\kappa}B{\alpha}$, a critical step for NF-${\kappa}B$ activation and translocation. Of the upstream signaling enzymes tested, Src and Akt were thought to be the responsible upstream signaling components in induction of NO production, although TMF-II strongly upregulated the phosphorylation of all MAPK pathways. Therefore, our data suggest that T. matsutake-derived ${\beta}$-glucan may exert its immunostimulating activities with similar potency to LPS via activation of multiple signaling pathways linked to NF-${\kappa}B$ activation.

• Biomolecules & Therapeutics
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• v.20 no.1
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• pp.36-42
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• 2012

Baicalin, a main constituent of the rhizome of Scutellaria baicalensis, is metabolized to baicalein and oroxylin A in the intestine before its absorption. To understand the role of intestinal microflora in the pharmacological activities of baicalin, we investigated its anti-inflammatory effect in mice treated with and without antibiotics. Orally administered baicalin showed the anti-inflammatory effect in mice than intraperitoneally treated one, apart from intraperitoneally administered its metabolites, baicalein and oroxylin A, which potently inhibited LPS-induced inflammation. Of these metabolites, oroxylin A showed more potent anti-inflammatory effect. However, treatment with the mixture of cefadroxil, oxytetracycline and erythromycin (COE) significantly attenuated the anti-inflammatory effect of orally administered baicalin in mice. Treatment with COE also reduced intestinal bacterial fecal ${\beta}$-glucuronidase activity. The metabolic activity of human stools is significantly different between individuals, but neither between ages nor between male and female. Baicalin was metabolized to baicalein and oroxylin A, with metabolic activities of $1.427{\pm}0.818$ and $1.025{\pm}0.603$ pmol/min/mg wet weight, respectively. Baicalin and its metabolites also inhibited the expression of pro-inflammatory cytokines, TNF-${\alpha}$ and IL-$1{\beta}$, and the activation of NF-${\kappa}B$B in LPS-stimulated peritoneal macrophages. Of them, oroxylin A showed the most potent inhibition. Based on these findings, baicalin may be metabolized to baicalein and oroxylin A by intestinal microflora, which enhance its anti-inflammatory effect by inhibiting NF-${\kappa}B$ activation.

• Preventive Nutrition and Food Science
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• v.19 no.2
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• pp.89-97
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• 2014

Broccoli (Brassica oleracea var. italia) florets were extracted with 80% methanol and the extract was sequentially fractionated with n-hexane, ethyl acetate, n-butanol, and distilled water. The extract and the fractions were evaluated for total phenolic content, sulforaphane content, antioxidant activity, and anti-inflammatory activity in lipopolysaccharide (LPS)-stimulated RAW 264.7 cells. The total phenolic content and sulforaphane content of the ethyl acetate fraction (EF) were 35.5 mg gallic acid equivalents/g and $620.2{\mu}g/g$, respectively. These values were higher than those of the 80% methanol extract and organic solvent fractions. The oxygen radical absorbance capacity of the EF [$1,588.7{\mu}M$ Trolox equivalents (TE)/mg] was 11-fold higher than that of the distilled water fraction ($143.7{\mu}M\;TE/mg$). The EF inhibited nitric oxide release from LPS-stimulated RAW 264.7 cells in a dose-dependent manner and inhibited $I{\kappa}B-{\alpha}$ degradation and nuclear factor-${\kappa}B$ activation in LPS-stimulated RAW 264.7 cells. In conclusion, the EF of broccoli florets exerted potent antioxidant and anti-inflammatory effects.

• BMB Reports
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• v.39 no.6
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• pp.788-793
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• 2006

Bacterial DNA containing immunostimulatory CpG motifs can stimulate antigen-presenting cells to express co-stimulatory molecules and to produce various cytokines in vivo and in vitro. In this study, we fragmented macromolecular E.coli genomic DNA with DNase I, and analyzed the ability of the resulting DNA fragments to induce the NF-${\kappa}B$ activation and humoral immune response. Furthermore, using computational analysis and luciferase assay for synthetic ODNs based on the sequence of the immunostimulatory DNA fragments (DF-ODNs), an active component of DF-ODNs sequences was investigated. Experimental results demonstrated that DF-ODN is optimal for the NF-${\kappa}B$-responsive promoter activation in the mouse macrophage cell line and the humoral immune response in vivo. In agreement with the activity of the DF-ODNs processed by DNase I, a synthetic ODN based on the DF-ODN sequences is potent at inducing IL-12 mRNA expression in primary dendritic cells. These results suggest that the discovery and characterization of a highly active natural CpG-ODN may be achieved by the analyses of bacterial DNA fragments generated by a nuclease activity.

• Food Science and Biotechnology
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• v.18 no.1
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• pp.262-266
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• 2009

To evaluate the anticolitic effect of red ginseng (RG, the steamed root of Panax ginseng CA. Meyer, Araliaceae), RG and its representative constituents, ginsenosides Rg3 and Rh2, were orally administered to dextran sulfate sodium (DSS)-induced colitic mice and inflammatory markers investigated. RG and its constituents, ginsenosides Rg3 and Rh2, inhibited colon shortening and myeloperoxidase activity induced by DSS. The ginsenosides Rg3 and Rh2 inhibited mRNA expression of interleukin (IL)-$1{\beta}$ as well as protein levels of IL-$1{\beta}$ and IL-6. These ginsenosides also inhibited the activation of a transcription nuclear factor (NF)-${\kappa}B$. Ginsenoside Rh2 was a more potent inhibitor than ginsenoside Rg3. The anticolitic effects of these ginsenosides were comparable with sulfasalazine.

• IMMUNE NETWORK
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• v.7 no.1
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• pp.39-47
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• 2007

Background: Stromal cell-derived factor (SDF)-1 is a potent chemoattractant for activated T cells into the inflamed Rheumatoid arthritis (RA) synovium. To determine the effect of macrophage migration inhibitory factor (MIF) on the production of SDF-1 in the inflamed RA synovium. Methods: The expression of SDF-1 and MIF in RA and Osteoarthritis (OA) synovium was examined by immunohistochemical staining. The SDF-1 was quantified by RT-PCR and ELISA after RA fibroblast like synoviocyte (FLS) were treated with MIF in the presence and absence of inhibitors of intracellular signal molecules. The synovial fluid (SF) and serum levels of MIF and SDF-1 in RA, OA and healthy control were measured by ELISA. Results: Expression of SDF-1 and MIF in synovium was higher in RA patients than in OA patients. The production of SDF-1 was enhanced in RA FLS by MIF stimulation. Such effect of MIF was blocked by the inhibitors of NF-${\kappa}B$. Concentrations of SDF-1 in the serum and SF were higher in RA patients than in OA patients and healthy control. SDF-1 and MIF was overexpressed in RA FLS, and MIF could up-regulate the production of SDF-1 in RA FLS via NF-${\kappa}B$-mediated pathways. Conclusion: These results suggest that an inhibition of interaction between MIF from T cells and SDF-1 of FLS may provide a new therapeutic approach in the treatment of RA.