巨细胞病毒临床分离株感染血管内皮细胞伴肾素基因表达  

Renin gene expression in the vascular endothelial cells infected by the clinical isolate of cytomegalovirus

作　　者：李多多[1,2] 程远[3] 程计林[1,4] 程新 刘宝玲 王海滨[3] 金壮[3] 张杰林[3] Crumpacker Clyde 

机构地区：[1]上海市复旦大学附属公共卫生临床中心,上海201508 [2]河南省新乡医学院第一附属医院小儿内一科,河南新乡453000 [3]河南省新乡医学院生理学与神经生物学教研室,河南新乡 [4]The Division of Infectious Disease,Beth Israel Deaconess Medical Center, Harvard Medical School, Harvard University, USA [5]Worcest Polytechnic Institute in USA [6]Massachusetts General Hospital, Harvard Medical School, Harvard University, USA.

出　　处：《公共卫生与临床医学》2009年第3期164-169,共6页Public health and dinical medicine

基　　金：复旦大学医学院基础-临床交叉研究基金（JC08-7）、复旦大学生物医学研究院（IBS）2009年度开放课题基金（2008B100）和上海市（复旦大学附属）公共卫生临床中心引进人才科研启动基金（RCJJ7）资助

摘　　要：目的探讨巨细胞病毒（Cytomegalovirus，CMV）是否感染血管内皮细胞伴肾素表达。方法①用10^7pfu（空斑形成单位）／mLCMV临床分离株BI-5和实验室型CMVA D169分别与10^6腹主动脉内度细胞和人脐静脉内皮细胞共同孵育，在23h后、d3、d7、d10和d14分别收集培养上清200μL，d14用PBS缓冲液洗细胞3次，收获细胞。每组实验均设培养液代替病毒液的无感染对照；②COBAS定量PCR检测培养上清中CMV DNA拷贝数；③PCR检测感染细胞中CMVpol基因；@RT-PCR、Real time RT-PCR和Western blot检测肾素在感染细胞内的表达。结果①BI-5和AD169感染静脉和动脉细胞后，其形态学变化相似，无细胞裂解病理效应；②AD169感染细胞不同时间培养上清中CMVDNA拷贝数无明显增加，BI-5呈增殖趋势；③BI-5感染动脉细胞CMVDNA拷贝数和肾素表达量均大于静脉细胞。结论临床分离株CMV以非裂解形式在血管内皮细胞持续存在并诱导肾素基因表达，血管内皮细胞分泌肾素可能是CMV感染引起心血管疾病的新机制。Objective To explore whether cytomegalovirus （CMV） is able to infect cardiovascular endothelial cells and cause renin gene expression. Methods ①Umbilical vein cell line and abdominal aorta cell line were respectively infected with the clinical isolate of CMV BI-5 and the laboratory strain of CMV AD169 at multiplicities of infection （MOI） of 10 （107 pfu/ml CMV/106 cells） or with mock control of media alone. 200μL supematant from each culture was collected at the 23rd hour, day 3, 7, 10, and 14 post-infection. At the 14th day of post-infection, the cells were rinsed with PBS three times and then harvested. ②CMV DNA copies in the supernatant were quantified by COBAS PCR. ③CMV DNA polymerase （pol） gene in the infected cells was determined by PCR. ④The renin expression in the viral infected cells was measured with RT-PCR, real time quantity RT-PCT and western blot. Results ①Lytic replication of CMV was not observed in both CMV-infected venous and arterial cells during the 14 day culture.②The clinical isolates BI-5 persistently replicated in both venous and arterial endothelial cells but not the AD169, determined by viral pol gene copies. ③CMV DNA and renin expression in CMV-infected arterial cells were higher than those in venous cells. Conclusions CMV was non-lytic and continuously released from the infected vascular ceils and induced renin expression in CMV-infected cells. The local renin from CMV- infected cells may play an important role in pathogenesis of atherosclerosis.

关 键 词：巨细胞病毒 血管内皮细胞 肾素 

分 类 号：R543.5[医药卫生—心血管疾病] R373.9[医药卫生—内科学]