Feasibility of ex vivo gene therapy for neurological disorders using the new retroviral vector GCDNsap packaged in the vesicular stomatitis virus G protein
Feasibility of ex vivo gene therapy for neurological disorders using the new retroviral vector GCDNsap packaged in the vesicular stomatitis virus G protein
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DOI：
10.1046/j.1471-4159.2002.01048.x
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发表时间：
2002-08-01
影响因子：
4.7
通讯作者：
Mochizuki, H
中科院分区：
文献类型：
--
作者：
Suzuki, A;Obi, K;Mochizuki, H
神经祖细胞（NPC）特别适合作为遗传和细胞治疗神经系统疾病（如帕金森病或中风）的目标群体。然而，利用逆转录病毒载体对这些细胞进行遗传修饰仍然是一个巨大的挑战，因为转导率低，并且在诱导细胞分化为成熟神经元的转导过程中需要胎牛血清（FCS）。为了克服这些问题，我们开发了一种新的逆转录病毒生产系统，该系统将经过改造的抗脱氧甲基化的简化逆转录病毒载体GCDNsap包装在水泡性口炎病毒G蛋白（VSV-G）中，离心浓缩，然后在无血清培养基（StemPro-34 SFM）中重悬。在以增强的绿色荧光蛋白（EGFP）为标记物的转导实验中，浓缩的不含fcs的病毒上清液在体外保持NPC细胞自我更新和分化的能力的同时，具有较高的感染率。将这些细胞移植到小鼠大脑中，在移植后8周，在注射部位周围区域检测到表达egfp的NPC。这些发现表明，本文描述的基因转移系统可能为基因修饰NPC治疗神经系统疾病提供有用的工具。
Neuronal progenitor cells (NPC) are particularly suited as the target population for genetic and cellular therapy of neurological disorders such as Parkinson's disease or stroke. However, genetic modification of these cells using retroviral vectors remains a great challenge because of the low transduction rate and the need for fetal calf serum (FCS) during the transduction process that induces the cell differentiation to mature neurons. To overcome these problems, we developed a new retrovirus production system in which the simplified retroviral vector GCDNsap engineered to be resistant to denovo methylation was packaged in the vesicular stomatitis virus G protein (VSV-G), concentrated by centrifugation, and resuspended in serum-free medium (StemPro-34 SFM). In transduction experiments using enhanced green fluorescent protein (EGFP) as a marker, the concentrated FCS-free virus supernatant infected NPC at a high rate, while maintaining the ability of these cells to self-renew and differentiate in vitro . When such cells were grafted into mouse brains, EGFP-expressing NPC were detected in the region around the injection site at 8 weeks post transplantation. These findings suggest that the gene transfer system described here may provide a useful tool to genetically modify NPC for treatments of neurological disorders.