单纯疱疹病毒Ⅱ型ICP27对Vero细胞凋亡作用的研究  

Investigation of Vero cell apoptosis caused by the herpes simplex virus type 2 ICP27

作　　者：龙朝钦[1] 杨慧兰[2] 张发洲[2] 薛礼长[2] 杜华[1] 

机构地区：[1]兰州兰州军区兰州总医院皮肤科,730050 [2]广州广州军区广州总医院皮肤科,510010

出　　处：《国际病毒学杂志》2013年第4期160-165,共6页International Journal of Virology

摘　　要：目的探讨单纯疱疹病毒Ⅱ型转录前蛋白ICP27对Vero细胞凋亡作用及其参与细胞凋亡的可能途径。方法用前期构建成功的pcDNA3．0-ICP27质粒瞬时转染Vero细胞，MTr法检测pcDNA3．0-ICP27转染Vero细胞的活性，Giemsa染色及流式细胞术检测转染后ICP27在细胞凋亡中的作用，实时荧光定量PCR检测Bax、Bcl-2mRNAs的变化和对凋亡执行蛋白Caspase-3的活性的检测以探讨ICP27参与细胞凋亡的途径。结果pcDNA3．0-ICP27转染Vero细胞后，M1Tr法检测发现其活性明显低于转染pcDNA3．0及空白转染的Vero细胞活性；Giemsa染色后发现转染pcDNA3．0-ICP27的Vero细胞变圆，细胞核破裂，流式细胞检测发现pcDNA3．0-ICP27转染的Veto细胞发生的凋亡率更高。凋亡执行蛋白Caspase-3的活性明显高于转染pcDNA3．0及空白对照；荧光定量PCR检测Bax、Bcl-2mRNAs的变化，转染重组质粒的Vero细胞中BaxmRNAs表达量上调，Bcl-2mRNAs表达降低，Bax／Bcl-2明显高于正常培养的Vero细胞及转染pcDNA3．0质粒的Vero细胞。结论ICP27对Vero细胞有促进细胞凋亡的作用。而这种作用可能是使Bax及Bcl-2在细胞中的比例发生变化而导致的。Objective To investigate the Veto cell apoptosis caused by the protein ICP27 of herpes simplex virus type II and its possible way involved in apoptosis. Methods The successfully constructed pcDNA3.0-ICP27 were transiently transfected into Vero cells. The viability of Veto cells transfected with pcDNA3.0-ICP27 was detected by MTT assay. The effect of ICP27 on Vero apoptosis was detected by Gime- sa stain and flow cytometry. Real time fluorescent quantitative PCR was carried out to detect the change of Bax and Bcl-2 mRNAs, and the Caspase-3 was detect by Caspase-3 kits to investigate the pathway of apopto- sis induced by ICP27. Results MTF assay showed that the viability of Vero cells transfected with peD- NA3.0-1CP27 was obviously lower than the Veto cell transfected with no plasmid or transfected with pcD- NA3. 0. Vero ceils transfected with pcDNA3. 0-ICP27 became round and the disrupted nucleus was observed by Giemsa stain. Flow cytometry showed that Vero cells transfected with pcDNA3.0-ICP27 had higher apop- tosis rate. The expression of Bax increased, while the expression of Bcl-2 decreased in Vero cells transfected with pcDNA3.0-ICP27. Compared with the normal Vero cells and the Vero cells transfected with pcD- NA3. 0, Bax/Bcl-2 significantly increased in the Vero cells transfected with pcDNA3. 0-ICP27. Conclusions HSV-2 ICP27 might induce apoptosis by changing Bax/Bcl-2 ratio in the course of reactivation from latent.

关 键 词：HSV-2 ICP27 真核表达 凋亡 

分 类 号：R752.12[医药卫生—皮肤病学与性病学]