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Improvement of muscle function by introducing genes by electroporation following intramuscular injection

肌肉注射后通过电穿孔导入基因改善肌肉功能

基本信息

批准号：
13670063
负责人：
MIYAZAKI Jun-ichi
金额：
$ 0.77万
依托单位：
Osaka University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2001
资助国家：
日本
起止时间：
2001 至 2002
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-13670063/
关键词：
electroporation gene therapy gene transfer expression vector Intramuscular injection

项目摘要

The proportion of aged people in the whole population is increasing rapidly, so that maintaining skeletal muscle function is becoming more important for the healthy life of people. Recently, various kinds of methods for in vivo gene transfer have been developed. For maintaining muscle function, direct plasmid DNA injection into muscle might be useful for improvement of muscle function. Previously, I demonstrated that gene transfer into muscle by electroporation in vivo was far more efficient than simple intramuscular DNA injection. In this study, I speculated that this method might be used to allow rat muscle to express some genes related to muscle function, insulin-like growth factor 1 (IGF-1) for proliferation of muscle cells, vascular endothelial growth factor (VEGF) for angiogenesis, and growth hormone(GH). And some indices, concentration of DNA, DNA solution, structure of electrodes and frequency of pulse, were analyzed for improvement of efficiency of this method. Vectors inserted with CDNA of these genes were introduced into cultured cells, showing that products of these genes were secreted into the culture media. As a control study, a plasmid containing erythropoietin cDNA was introduced into rat muscle. The results showed that the best condition for gene transfer was as follows: injection of 400 μg of DNA and electroporation with 5mm-electrode needle, 8 times of 50 msec-pulse at 100V.Intramuscular injection following by electroporation in vivo is simple and efficient for gene transfer, and may provide a potential approach toward systemic delivery of cytokines , growth factors, and other serum proteins for human gene therapy. More improvement of this method could contribute for human hearth.

老年人口在总人口中的比例迅速增加，维持骨骼肌功能对于人们的健康生活变得越来越重要。最近，已经开发了多种体内基因转移方法。为了维持肌肉功能，将质粒 DNA 直接注射到肌肉中可能有助于改善肌肉功能。此前，我证明通过体内电穿孔将基因转移到肌肉中比简单的肌肉内注射 DNA 更有效。在这项研究中，我推测这种方法可能用于让大鼠肌肉表达一些与肌肉功能相关的基因，用于肌肉细胞增殖的胰岛素样生长因子1（IGF-1）、用于血管生成的血管内皮生长因子（VEGF）和生长激素（GH）。并对DNA浓度、DNA溶液、电极结构、脉冲频率等指标进行了分析，以提高该方法的效率。将插入这些基因的cDNA的载体引入培养细胞中，显示这些基因的产物被分泌到培养基中。作为对照研究，将含有促红细胞生成素 cDNA 的质粒引入大鼠肌肉中。结果表明，基因转移的最佳条件为：注射400 μg DNA，用5mm电极针电穿孔，100V 50 msec脉冲8次。体内肌肉注射后电穿孔是简单而有效的基因转移方法，可能为人类基因治疗提供细胞因子、生长因子和其他血清蛋白的全身递送的潜在方法。这种方法的进一步改进可以为人类的壁炉做出贡献。