四氢黄连碱体外抗炎作用及机理研究  

Study on Anti-inflammatory Actions of Tetrahydrocoptisine in Vitro and Its Mechanism

作　　者：黄慧敏 王红梅 张斌强 陈琴华 HUANG Hui-Min;WANG Hong-Mei;ZHANG Bin-Qiang;CHEN Qin-Hua(Affiliated Dongfeng Hospital,Hubei University of Medicine,Shiyan 442008 Hubei,China)

机构地区：[1]湖北省湖北医药学院附属东风医院,湖北十堰442008

出　　处：《广州中医药大学学报》2019年第1期98-104,共7页Journal of Guangzhou University of Traditional Chinese Medicine

基　　金：湖北省教育厅青年人才项目(编号:Q20162113);十堰市科学技术研究与开发项目(编号:18Y78)

摘　　要：【目的】观察四氢黄连碱(THC)的体外抗炎作用及其相关机理。【方法】将体外培养的小鼠腹腔巨噬细胞分为空白对照组,模型对照组,四氢黄连碱低、中、高剂量组。以脂多糖(LPS)刺激小鼠原代腹腔巨噬细胞建立体外细胞炎症模型,作为模型对照组。四氢黄连碱低、中、高剂量组分别以不同浓度(终浓度为1、2、3μmol/L)的四氢黄连碱预培养小鼠巨噬细胞24 h后,加入LPS(终浓度为10μg/mL)。空白对照组加入相同体积的细胞培养液。继续培养12 h后,采用硝酸还原酶法检测细胞上清液中一氧化氮(NO)的含量;实时定量PCR(q PCR)法检测细胞内肿瘤坏死因子α(TNF-α)、白细胞介素6(IL-6)mRNA的表达;酶联免疫吸附分析(ELISA)检测细胞上清液中TNF-α、IL-6含量;ELISA检测细胞内核因子kappa B(NF-κB)与DNA的结合活性。【结果】与空白对照组比较,模型对照组小鼠巨噬细胞上清液中NO、TNF-α、IL-6含量,巨噬细胞TNF-α、IL-6 mRNA表达及NF-κBp65与DNA的结合活性均显著升高(P <0.001);经四氢黄连碱预处理小鼠巨噬细胞24 h后,上述指标均显著降低,且呈剂量依赖性(P <0.05或P <0.01或P <0.001)。【结论】四氢黄连碱具有显著的体外抗炎作用,其机理可能与阻断细胞内NF-κB信号通路的激活,从而下调促炎因子NO、TNF-α及IL-6的表达和分泌有关。Objective To observe the anti-inflammatory actions of tetrahydrocoptisine (THC) and its mechanism. Methods The macrophages isolated from mouse abdominal cavity were cultured and then were divided into blank control group,model control group,and low-,middle-and high-dose THC groups.The macrophages in the model control group were stimulated by lipopolysaccharide (LPS)to induce into cellular inflammation model in vitro.And the macrophages of THC groups were precultured with different concentrations of THC (at final concentrations of 1,2,3μmol/L)in low-,middle-and high-dose THC groups for 24h,and then were cultured with LPS (at final concentration of 10μg/mL),while the blank control group and the model control group were given the same volume of cell culture medium.After another 12-hour continuous culture,the content of nitric oxide (NO)in cell culture supernatant of various groups was measured by nitric reductase method,the mRNA expression levels of tumor necrosis factor alpha (TNF-α)and interleukin 6(IL-6)in mouse macrophages were detected by quantitative real-time PCR (qPCR),the levels of TNF-α and IL-6 in cell culture supernatant were detected by enzyme-linked immunosorbent assay (ELISA),and the activity of nuclear factor kappa B (NF-κB) binding to DNA in mouse macrophages was also detected by ELISA.Results Compared with the blank control group,the contents of NO,TNF-α and IL-6 in cell culture supernatant,and the mRNA expression levels of TNF-α and IL-6 as well as the activity of NF-κBp65 binding to DNA in mouse macrophages were increased significantly in the model control group (P <0.001).After THC pretreatment for 24 hours,the levels of the above indexes were decreased (P <0.05,P <0.01or P <0.001)in a dose-dependent manner.Conclusion THC has obvious anti-inflammatory effects in vitro,and its mechanism may be related with blocking the activation of NF-κB signaling pathways,which results into the down-regulation of the expression and secretion of NO,TNF-α and IL-6.

关 键 词：四氢黄连碱 巨噬细胞 炎症因子 细胞炎症模型 细胞培养 

分 类 号：R285.5[医药卫生—中药学]