白细胞介素-1受体相关激酶1蛋白过表达对细菌脂蛋白耐受的影响

目的 探讨细菌脂蛋白(BLP)信号转导分子Toll样受体2(TLR2)和白细胞介素-1受体相关激酶1(IRAK-1)与BLP耐受发生的关系.方法 在人胚肾293(HEK293)细胞中过表达TLR2以及IRAK-1蛋白,利用蛋白质免疫印迹法和双荧光素酶报告基因检测实验观察其对BLP耐受的影响.结果 以BLP刺激稳定转染并表达TLR2的HEK293细胞可以剂量依赖性地诱导NF-κB活化,而且BLP预处理HEK-TLR2细胞可诱导BLP耐受的发生.在HEK-TLR2细胞中过表达IRAK-1可以剂量依赖性地增加NF-κB的活化并逆转BLP耐受:转染前BLP活化组与耐受组的NF-κB活化程度分别为0.329±0.010和0.168±0.010;转染0.02μg IRAK-1质粒后活化组与耐受组的NF-κB活化程度分别为0.493±0.010和0.427±0.035,均比转染前明显升高,差异均有统计学意义(均P＜0.01).结论 HEK293细胞中过表达TLR2不能阻止BLP耐受的诱导,但是在HEK-TLR2细胞中过表达IRAK-1可部分逆转BLP耐受,提示IRAK-1蛋白表达水平的变化在BLP耐受发生中起关键作用;IRAK-1可作为细菌感染和脓毒症治疗的重要靶点.

The influence of over expression of interleukin-1 receptor-associated kinase 1 on bacterial lipoprotein-induced tolerance

Objective To investigate Toll-like receptor 2(TLR2)and interleukin-1 receptorassociated kinase 1(IRAK-1)in bacterial lipoprotein(BLP)tolerance.Methods Western blotting was used to confirm the over expression of TLR2 and IRAK-1 in human embryo kidney 293(HEK293)cells.Plasmids for dual luciferase reporter gene with nuclear factor-κB promoter(pNF-κB-Luc)or CMV promoter (phRL-CMV internal control vector)were used to detect the NF-κB activation and the induction of BLP tolerance in HEK-TLR2 cells.Results BLP stimulation resulted in dose-dependent NF-κB activation in HEK293 cells stably expressing TLR2.And BLP pretreatment could reduce NF-κB activation and induce BLP tolerance in HEK-TLR2 cells.The NF-κB activation was 0.329±0.010 and 0.168±0.010 in BLP-activated and BLP-tolerant HEK-TLR2 cells,respectively.After transfection with 0.02μg IRAK-1 plasmid,NF-κB activation in the two groups was 0.493±0.010 and 0.427±0.035,respectively(both P＜0.01).So over expression of IRAK-1 could increase NF-κB activation in a dose-dependent manner.Conclusion These results demonstrated that over expression of IRAK-1 could reverse BLP tolerance,whereas over expression of TLR2 failed to prevent the induction of BLP tolerance.Therefore reduced IRAK-1 protein expression is an important mechanism in the development of BLP-induced tolerance,suggesting that it could be a potentially important target for future therapeutic strategies in bacterial infection and sepsis.