慢病毒载体在大鼠颈动脉球囊损伤模型中的应用及观察  

Application and observation of lentiviral vector in rat carotid artery balloon injury

作　　者：杨简[1] 江洪[1] 李万强[2] 陈思思[1] 陈静[1] 徐盛开[1] 王继春[1] 

机构地区：[1]武汉大学人民医院心内科-武汉大学心血管病研究所,武汉430060 [2]武汉大学人民医院泌尿外科-武汉大学心血管病研究所,武汉430060

出　　处：《心肺血管病杂志》2010年第2期145-147,156,共4页Journal of Cardiovascular and Pulmonary Diseases

基　　金：国家自然科学基金(30770849)

摘　　要：目的:研究携带绿色荧光蛋白(Green fluorescent protein,GFP)的慢病毒转染球囊损伤大鼠颈动脉的效率和可行性,为利用慢病毒载体介导的基因治疗预防血管再狭窄奠定基础。方法:将携带GFP的慢病毒载体(pGC-LV-GFP载体)和慢病毒包装质粒供转染293T细胞,完成慢病毒颗粒包装及滴度测定。包装产生的慢病毒Lenti-GFP转染至球囊损伤的大鼠颈动脉。术后28d,损伤及病毒转染血管段标本分别行荧光显微镜、光镜检查,评估慢病毒的转染效率及内膜增生情况,并与假手术组(Sham组)、PBS对照组进行比较。结果:经包装产生的Lenti-GFP滴度为2×109TU/mL;术后28d,Sham组与PBS组血管中膜弹力层仅见微弱的自发性绿色荧光,而Lenti-GFP组动脉壁全层可见较强的绿色荧光分布;PBS组与Lenti-GFP转染组大鼠损伤的颈动脉均可见明显内膜增生,内膜和中膜面积之比差异无统计学意义。结论:Lenti-GFP成功转染至球囊损伤的大鼠颈动脉,并能维持目的基因稳定长期的表达,是血管再狭窄基因治疗的理想载体。Objective：To explore the feasibility of lentivirus transfection to deliver green fluorescent protein（GFP） into a balloon-damaged rat carotid artery,which can provide a basis for the treatment of restenosis after percutaneous coronary intervention（PCI）.Methods：pGC-LV-GFP vector was cotransfected with packaging plasmids into 293T cells by Lipofectamine 2000.The titer of virus was detected according to the expression of GFP.The carotid arteries of rats were injured by balloon catheter and followed by incubated with 100 μl Lenti-GFP or PBS solution for 30 minutes.The rats were euthanized for immunofluorescence and morphometric analysis at 28 day after operation.Results：The titer of concentrated lentivirus was 2 × 109 TU /ml.Compared with Sham and PBS control groups,the strong green fluorescence was observed in the all layers of Lenti-GFP transfection carotid arteries.Meanwhile,obvious neointimal formation was also found in the Lenti-GFP and PBS groups;However,there was no statistical difference between the two groups.Conclusion：Lenti-GFP can steadily infect balloon-injured carotid arteries and the transfection efficiency is considerable high,which makes the lentivirus become a promising vector for future gene therapy on restenosis after percutaneous coronary intervention.

关 键 词：慢病毒载体 颈动脉损伤 基因治疗 再狭窄 大鼠 

分 类 号：R543.4[医药卫生—心血管疾病]