呼吸道合胞病毒感染人肺上皮细胞诱导氧化应激对TLR7基因及其下游因子表达的影响  

Effect of oxidative stress on the expression of Toll-like receptor 7 and signaling molecules induced by respiratory syncytial virus infection in human lung epithelial cells

作　　者：朱童娜[1] 吴璇[1] 邱欢[1] 于莉[1] 黄升海[1] 

机构地区：[1]安徽医科大学基础医学院微生物学教研室,合肥230032

出　　处：《安徽医科大学学报》2014年第11期1539-1543,共5页Acta Universitatis Medicinalis Anhui

基　　金：安徽省自然科学基金(编号:1308085MH129)

摘　　要：目的观察呼吸道合胞病毒(RSV)感染人肺上皮细胞(A549细胞)引起的氧化应激对Toll样受体7(TLR7)活化及其主要信号分子的作用,探讨其所介导的抗炎作用和可能的调控机制,为临床治疗和预防RSV感染提供新的思路。方法 RSV体外感染A549细胞为感染组,以抗氧化剂BHA进行预处理作为抗氧化剂组,于感染后的4、8、12、24 h收集细胞,以未感染细胞作为正常组。1化学法检测活性氧指标·OH含量的变化;2硝酸还原酶法检测NO含量的变化;3用TRIzol提取细胞总RNA,通过半定量RT-PCR法检测TLR7、白介素(IL-6)的mRNA转录水平的变化;4 ELISA法检测细胞上清液中IL-6的表达变化;5 MTT法检测各组细胞增殖情况。结果 1感染组里·OH的含量升高并呈时间依赖性,在抗氧化剂组,·OH的含量与感染组相比是降低的;2感染组各时间段NO含量升高并呈时间依赖性,抗氧化剂组的NO含量与感染组同一时间点相比降低;3感染组中不同时间点的TLR7基因转录水平表达上调,抗氧化剂组TLR7的表达虽有上调,表达量都比感染组同时间点下降;4感染组里IL-6的表达量与感染之间有时间依赖性关系,抗氧化剂组IL-6的表达虽有上调,但比感染组同时间点下降;5感染组随时间点的延长,细胞抑制率增加,抗氧化剂组的细胞抑制率与感染组相比下调明显。结论 RSV感染可诱导氧化应激的产生,在加入抗氧化剂BHA预处理后,可明显减轻TLR7及其下游主要信号分子的表达变化和细胞抑制率,表明RSV感染活化TLR7及引起细胞损伤可能是通过氧化应激途径来调节的。Objective To investigate the effect of oxidative stress, which is induced by respiratory syncytial virus （RSV） ＇s infection in human lung epithelial cells （A549 cells）, on the expression of Toll-like receptor 7 （TLR7） and its major signaling molecules, explore the anti-inflammatory effects mediated by TLR7 and its possible regulato- ry mechanism, and provide new ways on clinical treatment and RSV＇s prevention. Methods We took RSV-infec- ted A549 cells in vitro as infection group, pretreatment of antioxidants BHA on A549 cells as antioxidant group （ cell harvest after infection of 4, 8, 12, and 24 h） , and uninfected A549 cells as reference group. First of all, we de- tected changes in reactive oxygen · OH＇s content by chemical method. Secondly, We used nitrate reductase assay to detect NOg content. Thirdly, we used TRIzol to extract cell＇s total RNA and measured changes of mRNA tran- scription levels of TLR7, IL-6 by semi-quantitative RT-PCR assay. Then, we used ELISA to detect changes of IL-6 g expression in cell supernatant. At last, we used MTT assay to detect cell proliferation in each group. Results Firstly, the content of · OH in the infection group increased in a time-dependent manner and the content of · OH was lower in the anti-oxidant group as to the infection group. Secondly, the NO＇s content in the infection group in- creased in a time-dependent manner. Besides, the NO＇s content was lower in the anti-oxidant group as to the infec- tion group at the same time point. Thirdly, the gene transcription levels of TLR7 increased with time extension both in the infection group and the antioxidant group, as the latter one was lower at the same time point. Fourthly, the gene transcription levels of IL-6 increased with time extension both in the infection group and the antioxidant group, however the latter one was lower at the same time point. At last, cell inhibition rate increased with time extension in the infection group. However, it dropped significantly in the anti-oxidant gro

关 键 词：呼吸道合胞病毒 TOLL样受体7 氧化应激 A549细胞 

分 类 号：R373.14[医药卫生—病原生物学]