TLR4在白念珠菌刺激小鼠巨噬细胞释放TNF-α和NO中的作用研究

目的　探求Toll样受体 4(TLR4)在白念珠菌刺激小鼠巨噬细胞系RAW 2 64 .7释放肿瘤坏死因子 α(TNF α)和一氧化氮 (NO)中的作用。方法　①体外培养RAW 2 64 .7,设置处理组 (抗TLR4单克隆抗体组 )、阴性对照组与空白对照组 ,分别给予白念珠菌刺激。②刺激 1h ,3h ,6h ,8h后用酶联免疫吸附试验检测各组分泌的TNF α水平。③刺激 6h后Griess法检测各组产生NO的水平。结果　①刺激 1h后抗体浓度为 2 0 μg/ml处理组分泌的TNF α水平低于阴性对照组 (P <0 .0 5 ) ,3h ,6h ,8h后各浓度处理组TNF α水平均低于阴性对照组 (P均 <0 .0 1) ,且呈浓度依赖性 ;② 6h后各浓度处理组NO水平与阴性对照组比较差异无显著性 (P >0 .0 5 )。结论　TLR4在白念珠菌刺激小鼠巨噬细胞释放TNF α中发挥作用 ,而对NO释放没有直接作用。

A Study on the Role of Toll-like4 Receptor(TLR4) in the Production of TNF-α and NO by Murine Macrophage Stimulated by Candida Albicans

Objective To investigate the role of Toll like receptor4 (TLR4)in the production of tumor necrosis factor alpha (TNF a) and Nitric Oxide (NO) by murine macrophage stimulated with Candida albicans. Methods ①RAW 264.7 cells were preincubated with anti TLR4 monoclonal antibodies in different concentrations in disposal groups;while in blank control groups an isotype matched control monoclonal antibodies was used. ②TNF α concentrations in the supernatant of each group were measured by enzyme linked immunosorbent assay after stimulated with heat killed Candida albicans for 1 h,3 h,6 h,8 h.③The NO level in the supernatants were determined by Griess reaction after stimulated with mactivated Candida albicans for 6h.Results ①The TNF α concentration was significant lower than that of the negative control group after stimulated for 1 h in the disposal group of 20 μg/ml anti TLR4 monoclonal antibodies( P <0.05).However,after stimulated for 3 h,6 h,8 h,the TNF-α concentrations in the supernatant of the each disposal group were much lower than that of the negative control groups( P <0.01).②There were no significant differences in NO level between the disposal groups and the control group after stimulated for 6 h( P >0.05).Conclusion TLR4 plays a role in the production of TNF α but not NO by murine macrophage after stimulated with Candida albicans .