炎性细胞因子抑制剂高通量筛选方法的研究  

Study of high throughput screening for inhibitors of inflammatory cytokine

作　　者：张莉[1] 杜冠华[1] 

机构地区：[1]中国医学科学院.中国协和医科大学药物研究所药物筛选中心,北京100050

出　　处：《中国药理学通报》2003年第11期1260-1263,共4页Chinese Pharmacological Bulletin

基　　金：" 8 63"创新药物和中药现代化重大专项资金资助;No2 0 0 2AA2Z3 43B;国家自然科学基金资助;No 3 0 0 1161940 ;No 3 0 17114 8

摘　　要：目的　建立能够同时作用于炎症相关细胞因子的药物筛选模型 ,探讨多指标综合筛选方法 ,为发现新的抗炎先导化合物提供快速有效的方法。方法　采用人血白细胞为研究对象 ,以脂多糖 (LPS)刺激诱发炎性因子生成 ,探讨最佳刺激条件 ,建立多指标检测方法和综合评价药物作用的分析方法。观察不同浓度脂多糖 (0 5～ 5 0mg·L-1)及不同作用时间 (0～ 4h)炎性因子IL 1、IL 8、TNF α生成水平的变化。同时观察 5 LO抑制剂NDGA (nordihydroguaiareticacid)和COX 2抑制剂Diclofenac在多种细胞因子筛选模型上的应用。结果　脂多糖可浓度依赖性 (0 5～ 5 0mg·L-1)和时间依赖性 (0～ 4h)地刺激人血白细胞释放IL 1、IL 8和TNF α。脂多糖 (终浓度 5mg·L-1)刺激人血白细胞 4h有较高释放 ,与对照组比差异明显。NDGA和Diclofenac在脂多糖刺激 4h对炎性因子释放有显著抑制作用。结论　这种脂多糖体外刺激人血白细胞释放多种炎性因子的筛选模型可用于高通量大规模筛选具有抗炎作用的先导化合物 。AIM The aim of our study was to establish drug screening models which can evaluate samples' effects on cytokines related with inflammtion, capable of fast and efficient screening of anti-inflammatory lead compounds on the release of inflammtory cytokines. METHODS Heparinized human blood leukocytes was evaluated as a model to study the effects of various classes of anti-inflammatory lead compounds on cytokine release/biosynthesis from leukocytes. Human blood leukocytes was stimulated with LPS (final concentration 0.5～50 mg·L -1), with or without test drugs (diclofenac, a cytooxygenase inhibitor, nordihydroguaiaretic acid NDGA, a 5-lipoxygenase inhibitor) for 1～4 h to induce cytokine release. RESULTS Human blood leukocytes stimulated with LPS could product IL-1, IL-8 and TNF-α in a dose-dependent manner. Human blood leukocytes was stimulated with LPS(5 mg·L -1) for 4h to induce cytokine release. TNF-α, IL-1 and IL-8 time-course profiles were determined in culture media, using bioassays and ELISA. LPS-mediated release of IL-1 and TNF-α was significantly suppressed by NDGA and Diclofenac. In LPS stimulated blood, NDGA and Diclofenac inhibited the release of TNF-α(IC 50 of 149 μmol·L -1 and 23.88 μmol·L -1) or IL-1 (IC 50 of 222.57 μmol·L -1 and 126 μmol·L -1). CONCLUSION This human blood leukocytes screening system in vitro has the potential to screen new cytokine release inhibitors and sites of action of new anti-inflammatory lead compounds, and increases the screening efficiency.

关 键 词：IL-1 TNF-α IL-8 脂多糖 人血白细胞 

分 类 号：R392.12[医药卫生—免疫学] R965.1[医药卫生—基础医学]