【摘要】 目的观察库普弗细胞(KCs)内毒素耐受形成后白细胞介素-1受体相关激酶-4(IRAK-4)表达的变化,探讨KCs内毒素耐受形成的相关机制。方法分离培养Balb／c小鼠KCs,分为非内毒素耐受组与内毒素耐受组[给予脂多糖(LPS)10ng／ml预处理24h]。用LPS 100ng／ml刺激后,蛋白免疫印迹法及逆转录聚合酶链反应法测定两组细胞IRAK-4蛋白及mRNA的表达水平；酶联免疫吸附法检测两组细胞核因子-kB(NF-kB)活性及培养上清液肿瘤坏死因子α(TNFα)含量。结果 LPS刺激在两组细胞均引起IRAK-4 mRNA表达及NF-kB活性增强、TNFα释放增加,但内毒素耐受组3种指标的高峰值明显低于非内毒素耐受组(t值分别为12.4、17.4、138.9,P值均＜0.01)。结论 LPS预处理可诱导KCs形成内毒素耐受状态,IRAK-4表达受到抑制可能是KCs内毒素耐受形成的机制之一。更多还原

【Abstract】 Objective To explore the mechanism of endotoxin tolerance (ET) through observing the expression of interleukin 1 receptor associated kinase-4 (IRAK-4) during endotoxin tolerance development in Kupffer cells (KCs). Methods Isolated KCs of Balb/c mouse were divided into two groups: the non-endotoxin tolerance (NET) group and the endotoxin tolerance (ET) group, which were pretreated with 10 ng/ml lipopolysaccharide (LPS) for 24 h. Then, the two groups were treated with 100 ng/ml LPS. The expressions of IRAK-4 gene and protein level were determined by RT-PCR and Western blot. The activities of NF-kB of KCs and the TNFα level were estimated by ELISA at 0 h, 1 h, 3 h, 6 h and 12 h after LPS stimulation. Results The ultimate level of IRAK-4, the activities of NF-KB and the TNFαlevel were evidently lower in the ET group than those in the NET group (t=12.4, 17.4 and 138.9 respectively, P＜0.01). Conclusions Pretreatment with LPS on KCs could induce endotoxin tolerance of KCs and inhibitation of IRAK-4 expression may be one of the reasons for its development.更多还原