人偏肺病毒感染人肺上皮细胞后Toll样受体的表达及其功能  

Toll-like receptors expression and function of human metapneumovirus infected human lung epithelial cells

作　　者：窦颖[1] 赵晓东[1] 赵耀[1] 

机构地区：[1]重庆医科大学附属儿童医院P2实验室,400014

出　　处：《中华微生物学和免疫学杂志》2009年第12期1122-1127,共6页Chinese Journal of Microbiology and Immunology

基　　金：国家自然科学基金重点项目（30730098）;教育部新世纪优秀人才支持计划（NCET-05-0774）;重庆市杰出青年科学基金（CSTC,2008BA5040）

摘　　要：目的观察人偏肺病毒（human metapneumovirus）感染人肺上皮细胞后Toll样受体（TLR）表达变化及其信号通路的功能，探讨hMPV诱导气道炎症的部分机制。方法hMPV感染体外培养的人肺上皮细胞株A549，检测病毒在A549细胞中的生长曲线，并通过RT—PCR，real—time RT-PCR方法检测细胞TLR mRNA的表达，ELISA检测细胞培养上清IFN-α及TNF-α的表达。结果（1）hMPV可在A549细胞中复制，感染后3d达高峰10^5.2TCID50／ml。（2）RT—PCR结果提示：hMPV感染A549细胞6h后大部分TLR的表达均上调。（3）定量PCR结果提示：hMPV感染A549细胞后TLR3-4、TLR7-9的表达增高，且有时间依赖性，而紫外灭活的hMPV刺激细胞后TLR表达无明显变化。（4）ELISA结果提示hMPV感染后24h，IFN—α及TNF—α的表达均明显升高。结论人偏肺病毒感染A549细胞后可上调TLR表达，其诱导的炎性反应与部分TLR介导的信号转导途径有关。Objective To investigate the expression changes of Toll-like receptors（TLR） and the signaling pathway function on airway epithelial cells infected with human metapneumovirus（ hMPV）, and to explore the mechanisms of hMPV-induced airway inflammation. Methods A549 cells were infected with hMPV live or co-cultured with UV-inactivated hMPV in vitro. Growth curve of the virus in A549 cells was established and mRNA expression of TLR were detected by RT-PCR assay and real-time PCR assay at 3,6, 9, 12, 24 h post infection. IFN-α and TNF-α in the culture supernatants were detected by ELISA. Results hMPV was able to replicate in A549 cell, growing to peak titer of 10^5.2TCID50/ml at 3 d after infection. Most of TLR mRNA levels were up-regulated by hMPV infection but not UV-inactivated viruses via RT-PCR assay （P 〈0.05 ） after 6 h of stimulation. Real-time PCR assay showed that TLR3-4, TLR7-9 mRNA increased after being infected by hMPV in a time-dependent manner. The expression of IFN-α and TNF-α in the culture supernatants were significantly up-regulated after 24 h infected by hMPV. Conclusion hMPV infection up-regulates the expression of TLR in lung epithelial cells. The inflammatory response to hMPV may associate with part of TLR signal transduction pathway.

关 键 词：人偏肺病毒 TOLL样受体 A549细胞 IFN-Α TNF—α 

分 类 号：R686[医药卫生—骨科学]