Identification of anti-inflammatory target genes of Rhizoma coptidis extract in lipopolysaccharide-stimulated RAW264.7 murine macrophage-like cells
Identification of anti-inflammatory target genes of Rhizoma coptidis extract in lipopolysaccharide-stimulated RAW264.7 murine macrophage-like cells
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DOI：
10.1016/j.jep.2010.05.022
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发表时间：
2010-07-20
影响因子：
5.4
通讯作者：
Lee, Na Gyong
中科院分区：
文献类型：
--
作者：
Kim, Jong Min;Jung, Hyun Ah;Lee, Na Gyong
研究目的：黄连在传统东方医学中被广泛用于治疗炎症性疾病。目的：研究黄连提取物(CEX)在脂多糖(LPS)刺激的RAW264.7小鼠巨噬细胞样细胞中的抗炎靶基因。材料和方法：用CEX处理RAW264.7细胞6h，并用寡核苷酸DNA芯片分析基因表达谱的变化。用半定量逆转录聚合酶链式反应对芯片分析结果进行验证。为证实CEX的抗炎活性，采用夹心ELISA法测定细胞因子释放浓度，Griess试剂检测NO的产生，免疫印迹法检测iNOS的表达水平。结果：基因芯片分析显示，脂多糖激活RAW264.7细胞后，许多已知与炎症反应相关的基因的mRNA表达发生了显著变化。CEX可抑制多种细胞因子/趋化因子、细胞表面分子、黏附分子和生长因子的表达。酶联免疫吸附试验也显示IL-1α、GMCSF和IL-6的分泌减少，但未见肿瘤坏死因子-α的分泌。结论：CEX通过在转录水平抑制参与炎症反应的多种促炎细胞因子和细胞表面分子的表达而发挥抗炎作用。这些数据支持CEX作为抗炎药的传统用法，并应为了解CEX的药理作用提供有用的信息。(C)2010爱思唯尔爱尔兰有限公司。保留所有权利。
Aim of the study: Rhizoma coptidis is used widely in traditional Oriental medicine to treat inflammatory diseases. The aim of this study was to identify the anti-inflammatory target genes of Rhizoma coptidis extract (CEX) in lipopolysaccharide (LPS)-stimulated RAW264.7 murine macrophage-like cells.Materials and methods: RAW264.7 cells were treated with CEX in the absence or presence of LPS for 6 h, and changes in gene expression profiles were analyzed using oligonucleotide DNA microarrays. The results of microarray analysis were validated by semiquantitative reverse transcription-polymerase chain reaction. To confirm the anti-inflammatory activity of CEX, the concentrations of cytokines released into the media were measured by sandwich ELISA, NO production was assessed using the Griess reagent, and iNOS expression levels were determined using immunoblot analysis.Results: Microarray analysis revealed that activation of RAW264.7 cells with LPS elicited marked changes in mRNA expression of numerous genes known to be associated with inflammatory responses. Treatment of the cells with CEX suppressed the expression of various cytokines/chemokines, cell surface molecules, adhesion molecules, and growth factors. An ELISA also showed a decrease in the secretion of IL-1 alpha, GMCSF, and IL-6 but not of TNF-alpha. iNOS protein expression and NO production were also reduced by CEX treatment.Conclusions: The data obtained in this study demonstrate that CEX exerts its anti-inflammatory effect by inhibiting the expression of various proinflammatory cytokines and cell surface molecules involved in inflammatory responses at the transcriptional level. These data support the traditional use of CEX as an anti-inflammatory agent and should provide useful information for the understanding of the pharmacological effects of CEX. (C) 2010 Elsevier Ireland Ltd. All rights reserved.