类鼻疽伯克霍尔德氏菌感染THP-1细胞模型的评价及其感染机制的初步探讨  

Evaluation of a Burkholderia pseudomallei invaded THP-1 cell model and primary investigation on mechanism of its infection to host cells

作　　者：黎元莉 方瑶[2] 陈海[2] 毛旭虎[2] 

机构地区：[1]海南省三亚市人民医院检验科,海南三亚572000 [2]第三军医大学医学检验系临床微生物学及免疫学教研室,重庆400038

出　　处：《第三军医大学学报》2015年第23期2348-2352,共5页Journal of Third Military Medical University

基　　金：海南省重点科技计划项目资助(ZDXM2014143);三亚市病原微生物学重点实验室(L1216)~~

摘　　要：目的建立类鼻疽伯克霍尔德氏菌(Burkholderia pseudomallei,B.pseudomallei,BP for short)感染人急性单核白血病细胞THP-1的细胞模型,探讨聚乙二醇3350(PEG 3350)对细菌侵入细胞及胞内生存的影响,对比类鼻疽伯克霍尔德氏菌与其减毒株(也称为泰国株,即Burkholderia thailandensis,B.thailandensis,BT for short)的感染特性。方法通过绘制细菌生长曲线,探讨类鼻疽伯克霍尔德氏菌及其减毒株的增殖规律;通过透射电镜观察细菌胞内感染状态和宿主细胞的超微病理变化;分析不同时相的细菌入胞率和胞内生存状态,探讨细菌与宿主细胞的相互作用;通过检测受感染细胞培养液的上清和患者血清中的细胞因子,评价宿主细胞的反应性和PEG对于细菌侵入细胞及胞内生存的影响,以及对比细胞受类鼻疽伯克霍尔德氏菌及其减毒株感染后的炎症反应性,从而评价该模型的稳定性。结果确定了B.pseudomallei和B.thailandensis的生长规律和感染时相特征及超微病理损伤特点。炎症因子测定中,两种细菌感染细胞后分泌的TNF-α出现感染中期的高峰,受BP感染的细胞的IL-6分泌则呈双峰改变,受BT感染的细胞IL-6的分泌和PEG对细胞分泌炎症因子的影响更为复杂。同时,患者血清中细胞因子的检测则没有显示出一定的规律,与其临床转归关系也不明确。结论成功构建类鼻疽伯克霍尔德氏菌感染THP-1细胞的模型,初步探讨了类鼻疽伯克霍尔德氏菌感染宿主细胞的机制。Objective To establish a cell model of human acute mononuclear leukemia cells THP-1 are infected by Burkholderia pseudomallei （BP） and then determine the effect of polyethylene glycol 3350 （PEG3350） on bacteria-affected cells and intracellular survival. To compare the infection characteristics of BP and its attenuated strain, Burkholderia thailandensis （BT） to lay the foundation of the infection mechanism of the bacterium. Methods The growth curves of BP and BT were drawn to probe their proliferation rule. Transmission electron microscopy was used to observe the ultra-structure pathological changes of the number of intracellular bacteria and the infected host cells for the reaction of the bacteria and host cells. Plate counting was tested for bacterial colonies at different hours after invasion to analyze the intracellular growth of bacteria. The cytokines in the supernatant of the infected cells and patients＇ serum samples were detected to evaluate the reactivity of host cells and the effect of PEG3350 on the bacteria-affected cells and intracellular survival. The inflammatory responses of infected cells to BP and BT strains were contrasted and analyzed. Thereby the stability of this model was evaluated. Results The growth curves, phase-infected characteristics, ultra-structurepathological impairments of BP and BT strains were revealed in this study. During the detection of cytokines, TNF-α contents in the supernatant of the cells infected by 2 strains of bacteria reached the peak in the infected metaphase, the exudation of IL-6 contents in the supernatant infected by BP presented 2 peaks, while, the exudation of IL-6 contents of the cells infected by BT and the effect of cytokines by PEG3350 were more complex. In addition, no definite rule was found in the detection of cytokines in patients＇ serum. There was no obvious relationship of the cytokines with clinical outcomes. Conclusion The model of BP-infected THP-1 cells is constructed successfully in this study. The mechanism of BP infecting the

关 键 词：类鼻疽伯克霍尔德氏菌 类鼻疽伯克霍尔德氏菌减毒株 人急性单核白血病细胞 聚乙二醇 

分 类 号：R378.99[医药卫生—病原生物学] R392.11[医药卫生—基础医学]