呼吸道合胞病毒诱导COX-2的表达

The investigation of COX-2 expression induced by respiratory syncytial virus in vivo and in vitro

目的研究呼吸道合胞病毒(RSV)感染Balb/c鼠及A549细胞(人肺腺癌上皮细胞)COX-2表达的变化,初步探讨COX-2在RSV感染所致炎性反应中的作用。方法以1×106PFU/mlRSV感染体外培养的A549细胞,在感染后8、16和24h,采用免疫细胞化学法检测COX-2蛋白表达;以1×106PFU/mlRSV滴鼻感染Balb/c鼠,在感染后24、48和96h,用免疫组化法检测肺组织COX-2蛋白表达,并对肺组织切片行HE染色;以未感染病毒的Balb/c鼠及A549细胞作正常对照。采用生物图像分析系统对免疫组化和免疫细胞化学结果进行COX-2定量检测。用TRIzol提取肺组织和A549细胞总RNA,逆转录聚合酶链反应(RT-PCR)检测COX-2 mRNA表达的变化。结果正常细胞对照组中,A549细胞COX-2 mRNA和蛋白有微弱表达,经由RSV感染后,A549细胞COX-2 mRNA和蛋白有高水平表达,差异有显著性(P<0.05),且随着感染时间的延长,其表达量逐渐增加。正常鼠肺组织中COX-2 mRNA和蛋白仅有微弱表达,RSV感染后,COX-2 mRNA和蛋白表达水平随感染时间延长而增加,差异具有显著性(P<0.05);HE染色显示肺组织炎症随感染时间延长而加重,且COX-2的表达水平与肺组织炎症严重程度相一致。结论RSV感染可诱导肺组织及A549细胞高水平表达COX-2 mRNA和蛋白,并可能参与RSV感染引起的肺组织炎性反应。

Objective To investigate the expression changes of COX-2 in human lung epithelial cells （A549 cells） and the lung tissue of Balb/e mice infected with respiratory syncytial virus （RSV） , and to explore the inflammatory response to RSV. Methods A549 cells were infected with 1 ×10^6 PFU/ml RSV. After infection of 8,16 and 24 h, the expression of COX-2 protein of A549 cells was measured with immunoeytoehemical method. Balb/e mice were infected with 1×10^6 PFU/ml RSV intranasally. After infection of 24 -48 and 96 h, the expression of lung COX-2 protein was measured with immunohistochemieal method （SP assay）. HE stain was performed to detect inflammatory level of lung tissues. COX-2 protein in mouse lung and A549 cells were assessed quantitatively by biological image analysis system. Additionally, Trizol was used to extract total RNA of lung tissues and cells, and RT-PCR was used to evaluate the expression of COX-2 mRNA. Results The expression of COX-2 mRNA and protein in normal A549 cells was very weak, whereas the amounts of COX-2 mRNA and protein in A549 cells infected with RSV increased in a time-dependent manner. RSV infection promoted COX-2 mRNA and protein expression, which was higher than that of uninfected group （ P 〈 0. 05 ）. COX-2 mRNA and protein in lung tissues of RSV-infeeted mice increased in a time-dependent manner compared with that of normal group （ P 〈 0. 05 ）. HE staining showed pulmonary inflammation in RSV-infected mice was heavier along with the infective process. Furthermore, the levels of COX-2 mRNA and protein were consistent with the degree of mouse pulmonary inflammation. Conclusion RSV infection can induce Balb/c mouse lung tissues and A549 cells to express high - level of COX-2 mRNA and protein, which may be involved in the lung inflammatory process caused by RSV infection.