9型重组腺相关病毒转染microRNA-21前体在大鼠心肌的转染效率及安全性  

The transfection efficiency and safety of recombinant adeno-associated virus serotype 9 transducing microRNA-21 precursor in rat myocardium

作　　者：杨琼[1] 杨侃[1] 李安莹[1] 谭文鹏[2] 

机构地区：[1]中南大学湘雅三医院心内科,湖南长沙410013 [2]广州医学院第三附属医院心内科,广东广州510150

出　　处：《中国现代医学杂志》2013年第9期13-17,共5页China Journal of Modern Medicine

摘　　要：目的研究含荧光标记ZsGreen的9型重组腺相关病毒(rAAV9)转染目的基因microRNA-21前体(pre-miR-21)至大鼠心肌的转染效率、表达时间及安全性。方法用冠脉注射方法转染PBS及rAAV9-Zs-Green-pre-miR-21 1.0×1010vg/只、1.0×1011vg/只、1.0×1012vg/只至SD大鼠心肌,倒置荧光显微镜观察心肌组织荧光表达并计算转染效率,Realtime PCR方法测定miR-21表达,超声心动图观察大鼠心功能。结果冠脉注射后7 d PBS组未见绿色荧光,余各组大鼠心肌均可观察到少量绿色荧光表达,第14天达高峰,之后呈下降趋势。14 d高病毒量组较其他中、低病毒量组转染效率明显高([73.7±8.4)%vs(39.6±4.8)%,(P<0.001)];([73.7±8.4)%vs(24.1±4.2)%,(P<0.001)]。14 d miR-21表达在低、中、高病毒量组较PBS组分别增加1.51倍、2.89倍及4.43倍。rAAV9转染后对大鼠心功能无影响(P>0.05)。结论 rAAV9-Zs-Green-pre-miR-21可以有效、持久、安全地在大鼠心肌表达。【Objective】 To evaluate the transfection efficiency and safety using recombinant adeno-associated virus Serotype 9 mediated microrna-21 precursor（pre-miR-21） to rat myocardium.【Methods】 rAAV9-ZsGreen-pre-miR-21 was transfected into rat myocardium by coronary injection at different vector dose（1×1010vg,1×1011vg,1×1012vg）.ZsGreen expression in myocardium was observed under inverted fluorescence microscope.Realtime-PCR was used to assess the expression level of miR-21,Echocardiography was used to assess cardiac function.【Results】 Myocardium with rAAV9-ZsGreen-pre-miR-21 transfection at three groups began to exhibit ZsGreen expression 7 days after transfection with exception of the PBS group.The fluorescence intensity increased with the vector dose used for transfection.ZsGreen expression reached the maximum on day 14,at the point of which the transfection efficiency of rAAV9-ZsGreen-pre-miR-21 in myocardium was（24.1±4.2）%,（39.6±4.8）% and（73.7±8.4）%.Corresponding to vector dose 1×1010vg,1×1011vg,1×1012vg,respectively.MiR-21 were 1.51,2.89,4.43 times than the PBS group on day 14,corresponding to vector dose 1×1010vg,1×1011vg,1×1012vg,respectively.Echocardiography did not reveal significant difference in the cardiac function among four groups.【Conclusion】 rAAV9-ZsGreen-pre-miR-21 gene can be stably and efficiently expressed in rat myocardium without affecting cardiac function.

关 键 词：腺相关病毒 9型重组腺相关病毒 MICRORNA-21 转染效率 

分 类 号：R346[医药卫生—基础医学]