Purkinje-cell-preferential transduction by lentiviral vectors with the murine stem cell virus promoter
Purkinje-cell-preferential transduction by lentiviral vectors with the murine stem cell virus promoter
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DOI：
10.1016/j.neulet.2008.07.058
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发表时间：
2008-09
影响因子：
2.5
通讯作者：
Kiyohiko Takayama;T. Torashima;Hajime Horiuchi;H. Hirai
中科院分区：
文献类型：
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作者：
Kiyohiko Takayama;T. Torashima;Hajime Horiuchi;H. Hirai
病毒载体介导的基因导入Purkinje细胞是探索小脑病理生理学的一种有前途的方法；然而，由于转录活性的弱点，使用带有细胞类型特异性启动子的病毒载体在Purkinje细胞中实现足够高的基因表达通常是困难的。在本研究中，我们制备了在小鼠干细胞病毒、巨细胞病毒、巨细胞病毒早期增强子/鸡β肌动蛋白和劳斯肉瘤病毒启动子的控制下表达绿色荧光蛋白的慢病毒载体，并比较了它们对浦肯野细胞的转导能力。慢病毒注射后7d处死小鼠，测定GFP(+)浦肯野细胞占全部转导细胞的比例。转导效率以含MSCV启动子的慢载体为最高：∼70%的绿色荧光蛋白(+)细胞为浦肯野细胞，其次是CMV启动子(∼40%)，其次是CAg启动子(∼35%)，最低为∼启动子(10%)。此外，最高水平的GFP表达也是由MSCV启动子引起的。因此，在所研究的普遍存在的启动子中，MSCV启动子是在体内浦肯野细胞中表达外源基因的最佳启动子。
Viral-vector-mediated gene delivery into Purkinje cells is a promising method for exploring the pathophysiology of the cerebellum; however, it is generally difficult to achieve sufficiently high levels of gene expression in Purkinje cells using viral vectors with a cell-type-specific promoter because of the weakness of transcriptional activity. In this study, we prepared lentiviral vectors that express GFP under the control of various ubiquitous promoters derived from murine stem cell virus (MSCV), cytomegalovirus (CMV), CMV early enhancer/chicken β actin (CAG), and Rous sarcoma virus (RSV) and compared their potential to transduce Purkinje cells. Mice were sacrificed 7 days after lentiviral injection and the ratios of GFP(+) Purkinje cells to all transduced cells were determined. The highest transduction ratio was observed when we used lentivectors containing the MSCV promoter: ∼70% of GFP(+) cells were Purkinje cells, the next highest ratio was for the CMV promoter (∼40%), then the CAG promoter (∼35%), and the lowest ratio was for the RSV promoter (∼10%). Moreover, the highest levels of GFP expression were also caused by the MSCV promoter. Thus, among the ubiquitous promoters examined, the MSCV promoter was the best for the expression of a foreign gene in Purkinje cells in vivo.