肺炎克雷伯杆菌对小鼠中性粒细胞表达IL-1β和TNF-α的影响

Effects of Klebsiella pneumoniae on the expression of cytokines by polymorphonuclear neutrophil granulocytes in mice

目的观察肺炎克雷伯杆菌(Klebsiellapneumoniae,Kle.p)感染对小鼠中性粒细胞细胞因子表达的影响。方法以热灭活的Kle.p感染小鼠肺部建立急性肺炎模型,分离并纯化中性粒细胞(PMN),用RT-PCR、Westernblotting、ELISA等方法检测PMN中TNF-α和IL-1β的表达改变。结果Kle.p感染促进了早期小鼠中性粒细胞肿瘤坏死因子-α(TNF-α)和白介素-1β(IL-1β)的mRNA表达及分泌,TNF-αmRNA表达在感染6h最高,12h后降低;IL-1β的mRNA表达在感染12h最强;24h后两者表达均减弱,但仍显著强于对照组。Westernblotting结果显示,感染6h后TNF-α、IL-1β的蛋白表达量最高,24h后表达明显减少。ELISA结果显示,TNF-α分泌量在感染后6h最高,24h后明显降低,但仍较对照组高;IL-1β分泌量峰值在感染12h后出现。结论TNF-α可能主要参与小鼠感染热灭活Kle.p早期的PMN免疫防御功能;而IL-1β则主要参与感染中后期的PMN免疫防御功能。

Objective To observe the effect of Klebsiella pneumoniae (Kle.p) on the expression of cytokines in mouse polymorphonuclear neutrophil granulocytes (PMNs).Methods Mouse models of acute pneumonia were induced by intranasal inoculation of 30 μl PBS containing 1×107 Kle.p (heat-inactivated at 60 ℃ for 1.5 h).The mRNA and protein expressions of the cytokines in the isolated and purified PMNs were assayed by reverse transcriptional (RT)-PCR,Western blotting and enzyme- linked immunosorbent assay (ELISA),respectively.Results Kle.p infection induced the mRNA expressions of tumor necrosis factor (TNF)-α and interleukin (IL)-1β in the early stage,and TNF-α mRNA reached its peak at 6 h and IL-1β mRNA at 12 h after infection,both of which decreased 24 h after infection.Western blot analysis showed that the protein expressions of TNF-α and IL-1β were the highest at 6 h,and decreased obviously at 24 h after infection.ELISA showed that the secretion of TNF-α peaked at 6 h and decreased obviously at 24 h after infection,while the peak of IL-1β production occurred at 12 h and maintained the high level till 24 h after the infection.Conclusion TNF-α may participate in the immune defense of PMN against Kle.p infection mainly in the early stage of infection while IL-1β mainly in the later stage.