Functional in vivo gene transfer into the myofibers of adult skeletal muscle
Functional in vivo gene transfer into the myofibers of adult skeletal muscle
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DOI：
10.1016/j.bbrc.2003.11.032
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发表时间：
2003-12-26
影响因子：
3.1
通讯作者：
Carraro, U
中科院分区：
文献类型：
--
作者：
Donà, M;Sandri, M;Carraro, U
骨骼肌纤维有丝分裂后的性质和寿命允许任何转染基因的稳定表达。在成人和再生肌肉中直接注射质粒DNA是一种安全、廉价和简单的方法。在这里，我们提出了一种优化的电穿孔方案，该方案基于使用刮刀电极将cDNA在体内转移到解剖学定义的肌肉的成人肌纤维中，可以对其进行功能表征。在我们的研究中，大约80%的成人肌纤维在体内被不同的质粒转染，用于GFP融合蛋白或β -半乳糖苷酶。与标准DNA传递相比，荧光素酶活性增加了几个数量级。在解剖上确定的肌肉中，广泛的基因转移与逆转录病毒的传递相当或更好，最近在人类临床研究中已被证明容易产生严重的副作用。此外，我们的方法大大减少了组织损伤。因此，目前的工作描述了成人骨骼肌纤维中基因的体内功能电转移，这一方案在基因治疗和基础研究方面具有很大的潜力。(C) 2003 Elsevier Inc.版权所有。
The postmitotic nature and longevity of skeletal muscle fibers permit stable expression of any transfected gene. Direct in vivo injection of plasmid DNA, in both adult and regenerating muscles, is a safe, inexpensive, and easy approach. Here we present an optimized electroporation protocol based on the use of spatula electrodes to transfer cDNA in vivo into the adult myofibers of an anatomically defined muscle, which could be functionally characterized. In our hands, about 80% of adult myofibers were transfected in vivo by different plasmids for GFP fusion proteins or for beta-galactosidase. The luciferase activity increased several orders of magnitude when compared to standard DNA delivery. In an anatomical defined muscle, the wide gene transfer was comparable to or better than that of retrovirus delivery, that recently has been shown to be prone to severe side-effects in human clinical studies. Furthermore, with our method the tissue damage was greatly decreased. Thus, the present work describes in vivo functional electrotransfer of genes in adult skeletal muscle fibers by a protocol that is of great potential for gene therapy, as well as for basic research. (C) 2003 Elsevier Inc. All rights reserved.