人参多糖对LPS诱导小鼠巨噬细胞炎症因子生成的抑制作用及其机制

Inhibitory Effect of Ginseng Radix et Rhizoma Polysaccharide on LPS-induced Inflammatory Factor Formation in Mouse Peritoneal Macrophages and Its Mechanism

目的：研究人参多糖对脂多糖（LPS）诱导小鼠巨噬细胞RAW264.7炎症反应的抑制作用及机制。方法：细胞以2×10^4个/m L的密度接种于6孔培养板,将细胞分为空白组,LPS组,LPS＋地塞米松组,LPS加人参多糖组（质量浓度分别为31.25,62.6,125,250,500 mg·L^-1）。用LPS（1 mg·L^-1）刺激生长良好的RAW264.7细胞24 h建立体外细胞炎症模型,Griess试剂法检测一氧化氮（NO）含量,酶联免疫吸附测定法（ELISA）检测细胞上清液中肿瘤坏死因子-α（TNF-α）和白细胞介素-6（IL-6）含量,实时荧光定量聚合酶链式反应（Real-time PCR）检测一氧化氮（NO）,TNF-α和IL-6 mRNA的表达,蛋白免疫印迹法（Western blot）检测细胞K基因结合核因子抗体核转录因子-κB（NF-κB）p65蛋白的表达。结果：与空白组比较,LPS模型组细胞因子NO,TNF-α,IL-6表达量显著升高（P〈0.01）,炎症模型建立成功。给予人参多糖干预后,与LPS组比较,31.25~500 mg·L-1的人参多糖可以明显降低LPS诱导的RAW264.7细胞释放炎症因子NO,TNF-α和IL-6水平（P〈0.05,P〈0.01）,抑制i NOS,TNF-α和IL-6 mRNA的转录（P〈0.05,P〈0.01）,与LPS组比较,125~500μmol·L-1可以显著抑制细胞核内NF-κB p65的表达（P〈0.05,P〈0.01）。结论：人参多糖能明显抑制巨噬细胞炎症因子NO,TNF-α和IL-6过度表达,从而抑制炎症反应,其机制可能与抑制相关炎症因子和mRNA的表达和抑制NF-κB信号通路相关。

Objective： To study the anti-inflammatory effect and mechanism of Ginseng Radix et Rhizoma polysaccharide on inflammation factors caused by lipopolysaccharide（ LPS）-induced macrophage in mice.Method： RAW264. 7 cells were cultured in 6-well plates with the concentration of 2 × 10^4 cells/m L. The cells was divided into blank group,model group,LPS plus dexamethasone group, LPS plus Ginseng Radix et Rhizoma polysaccharide groups（ 31. 25, 62. 6, 125, 250, 500 mg ·L^-1）. RAW264. 7 cells were polarized to inflammatory macrophage by treating with 1 mg·L^-1 LPS for 24 h. The content of nitricoxide（ NO） was assayed by Griess reagent. Enzyme linked immunosorbent assay（ ELISA） was used to detect the content of inflammatory mediators,such as tumor necrosis factor-α（ TNF-α） and interleukin-6（ IL-6） in cell supernatant. The expressions of i NOS,IL-6,TNF-α mRNA were detected by real-time quantitative PCR（ Real-time PCR）. And the expression of mitogen-activated protein kinase nuclear transcription factor（ NF）-κB p65 protein was determined by Western blot. Result： Compared with blank group,the expressions of NO,IL-6,TNF-α of model group were significantly higher（ P〈0. 01）. It meant that the inflammation model was successfully established.Compared with model group,31. 25-500 mg·L^-1 of Ginseng Radix et Rhizoma polysaccharide in LPS-induced RAW264. 7 cells greatly inhibited the release of inflammatory mediators,such as NO,TNF-α and IL-6 levels（ P〈0. 05,P〈0. 01）. Compared with the model group, 31. 25-500 mg·L-1 of Ginseng Radix et Rhizoma polysaccharide reduced the concentrations of i NOS, IL-6, TNF-α mRNA（ P〈0. 05, P〈0. 01）. And the expression of p65 protein was decreased（ P〈0. 05, P〈0. 01）. Conclusion： Ginseng Radix et Rhizoma polysaccharide can significantly inhibit the over expression of macrophage inflammatory factors NO,TNF-α and IL-6. Its mechanism may be related to the inhibition of the expressions of related inflammatory factors mRNA and the inhibition of NF-κB signaling pathway.