摘要
人巨细胞病毒(human cytomegalovirus,HCMV)在神经胶质瘤细胞中的复制水平不一,其机制尚不清楚。本研究通过下调转录激活因子5(ATF5)在神经胶质瘤细胞中的表达,检测HCMV感染神经胶质瘤细胞后病毒复制水平的变化。首先用HCMV AD169(MOI=5)分别感染U87、SY5Y及A172细胞,观察细胞形态变化,分别在24、48、72、96、120 h取各时间点上清液检测病毒滴度;Real-time PCR检测HCMV即刻早期基因(IE2)、早期基因(UL44)、晚期基因(UL99)及ATF5的表达情况;Western-blot检测病毒基因编码蛋白及ATF5表达的情况。结果显示HCMV在U87、SY5Y细胞中复制水平与病毒在A172细胞中复制水平相比,U87、SY5Y细胞组明显高于A172细胞组(P<0.05),ATF5表达在U87、SY5Y细胞组与A172细胞组相比,U87、SY5Y细胞组ATF5表达明显高于A172组(P<0.05);利用慢病毒介导的RNA干扰技术下调ATF5在U87、SY5Y细胞的表达,用HCMV感染细胞检测病毒基因及蛋白的表达,结果ATF5表达下调可抑制HCMV的复制(P<0.05)。以上结果表明,在胶质瘤细胞中下调ATF5表达水平可以抑制HCMV的复制水平。
The replication levels of human cytomegalovirus（HCMV） in gliomata are different,and the mechanism is still unknown.In this study the changes of replication levels of the virus after its infection of gliomata were detected through down-regulation of the expression of activation transcription factor 5（ATF5） in gliomata.Firstly,cells U87,SY5 Y,and A172 were infected with HCMV respectively,and observe the changes of their cellular morphology,and collect each of their supernatants at the time points of 24,48,72,96,and 120 h to test their viral titers,and detect the expression situation of incontinent incipient gene IE2,incipient gene UL44,terminal gene UL99,and ATF5 with real-time PCR,and detect the expression situation of the HCMV coded protein and ATF5 with Western-blot.The results showed that as compared the replication level of HCMV in cells A172 with the ones in cells U87 and SY5 Y,the group of U87 and SY5 Y cells were significantly higher than that of the replication level in the group of A172 cells（P〈0.05）; and as compared the expression of ATF5 in the groups of U87 cell and SY5 Y cell were significantly higher than the group of A172（P〈0.05）; Using lentivirus mediation RNA interfering technique to down-regulate the expression of ATF5 in U87 and SY5 Y,using HCMV infected cells to detect the expression of viral gene and protein,the results showed that the down-regulation of the expression of ATF5 could inhibit the replication of HCMV（P〈0.05）.The above results showed that down-regulate the expression level of ATF5 in glioma cell could inhibit the replication level of HCMV.
出处
《微生物学杂志》
CAS
CSCD
2016年第2期25-32,共8页
Journal of Microbiology
基金
国家自然科学基金项目(81471958)
青岛大学创新团队青年教师培育项目
青岛市博士后应用研究项目资助(2015160)
关键词
人巨细胞病毒
转录激活因子5
复制
人胶质瘤细胞
病毒基因
human cytomegalovirus（HCMV）
activation transcription factor 5（ATF5）
replication
human glioma cell
viral gene
