[국내논문]Aqueous extract of Lycii fructus suppresses inflammation through the inhibition of nuclear factor kappa B signal pathway in murine raw 264.7 macrophages원문보기
Lycii fructus is the fruit of Lycium chinense Miller and is part of the Solanaceae family. Lycii fructus produces various effects such as hypotensive, hypoglycemic, anti-pyretic, and anti-stress activities. Lycii fructus is known to contain betaine, carotene, nicotinic acid, zeaxanthin, and cerebros...
Lycii fructus is the fruit of Lycium chinense Miller and is part of the Solanaceae family. Lycii fructus produces various effects such as hypotensive, hypoglycemic, anti-pyretic, and anti-stress activities. Lycii fructus is known to contain betaine, carotene, nicotinic acid, zeaxanthin, and cerebroside. In the present study, the effects of Lycii fructus aqueous extract on lipopolysaccharide (LPS)-induced inflammation in murine raw 264.7 macrophage cells were investigated. In this study we utilized the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, reverse transcriptionpolymerase chain reaction (RT-PCR), Western blotting, and nitric oxide (NO) detection. Lycii fructus aqueous extract suppressed NO production by inhibiting the LPS-induced expressions of inducible nitric oxide synthase (iNOS) and tumor necrosis factor-alpha (TNF-$\alpha$) mRNA and iNOS protein in murine raw 264.7 macrophage cells. Also, Lycii fructus aqueous extract suppressed the activation of nuclear factor-kappa B (NF-${\kappa}B$) in the nucleus. These results demonstrated that Lycii fructus aqueous extract causes an anti-inflammatory effect that was likely produced by the suppression of iNOS expression through the down-regulation of NF-$\hat{e}B$ binding activity.
Lycii fructus is the fruit of Lycium chinense Miller and is part of the Solanaceae family. Lycii fructus produces various effects such as hypotensive, hypoglycemic, anti-pyretic, and anti-stress activities. Lycii fructus is known to contain betaine, carotene, nicotinic acid, zeaxanthin, and cerebroside. In the present study, the effects of Lycii fructus aqueous extract on lipopolysaccharide (LPS)-induced inflammation in murine raw 264.7 macrophage cells were investigated. In this study we utilized the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, reverse transcriptionpolymerase chain reaction (RT-PCR), Western blotting, and nitric oxide (NO) detection. Lycii fructus aqueous extract suppressed NO production by inhibiting the LPS-induced expressions of inducible nitric oxide synthase (iNOS) and tumor necrosis factor-alpha (TNF-$\alpha$) mRNA and iNOS protein in murine raw 264.7 macrophage cells. Also, Lycii fructus aqueous extract suppressed the activation of nuclear factor-kappa B (NF-${\kappa}B$) in the nucleus. These results demonstrated that Lycii fructus aqueous extract causes an anti-inflammatory effect that was likely produced by the suppression of iNOS expression through the down-regulation of NF-$\hat{e}B$ binding activity.
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제안 방법
7 macrophages were investigated. The procedures preformed in this study included 3- (4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, reverse transcription-polymerase chain reaction (RT-PCR), Western blot and NO detection.
For TNF-α, the PCR procedure was carried out using a PTC-0150 MiniCycler (Bio-Rad, Hercules, CA, USA) under the following conditions: initial denaturation at 94 °C for 5 min, followed by 35 amplification cycles, each consisting of denaturation at 94 °C for 30 sec, annealing at 60 °C for 30 sec, and extension at 72 °C for 30 sec, with an additional extension step at the end of the procedure at 72 °C for 10 min.
Analysis of the expression of iNOS protein was performed in order to provide an estimate of the relative level of expression of this protein.
RT-PCR analysis to determine the mRNA levels of TNF-α and iNOS was performed in order to provide an estimate of the relative expressions of these genes.
대상 데이터
For cyclophilin, the internal control used in the study, the primer sequences were 5'-ACCCCACCGTGTTCTTCGAC-3' (a 20-mer sense oligonucleotide) and 5'- CATTTGCCATGGACAAGATG-3' (a 20-mer antisense oligonucleotide).
Lipopolysacharride (LPS), pyrrolidine dithiocarbamate (PDTC), 3-(4,5-dimethyl thiazol-2yl)-2,5-diphenyl tetrazolium bromide (MTT), sodium dodecyl sulfate (SDS), hydrochloric acid (HCl), diethylpyrocarbonate (DEPC), 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid (HEPES), sodium chloride, potassium chloride, glycerol, Triton X-100, magnesium chloride hexahydrate), ethylenediaminetetraacetic acid (EDTA), ethyleneglycol-bis-(β-aminoethylether)-N,N'-tetraacetic acid (EGTA), phenylmethylsulfonyl fluoride (PMSF), eupeptin, pepstatin, sodium ortho vanadate, sodium floride, Dithiothreitol (DTT), aprotinin, Nonidet P - 40, tris-hydroxymethylaminomethane (Tris-base), agarose, ethidium bromide (EtBr), acrylamide, bisacrylamide, and almost drugs used in this study were purchased from Sigma Chemical Co. (St. Louis, MO, USA).
데이터처리
The data were analyzed by one-way ANOVA followed by Duncan’s post-hoc test using SPSS version 12.0.
성능/효과
The present results showed that the aqueous extract of Lycii fructus exerted no cytotoxic effects at concentrations less than 1,000 µg/ml.
The present results showed that LPS enhanced the expression of NF-κB (p65) protein while the aqueous extract of Lycii fructus and PDTC decreased the LPS-induced expression of NF-κB (p65) protein in the nuclear fraction.
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