TRANSGENE S.A., 11 rue de Molsheim, 67082, Strasbourg Cedex, France.
Cytotechnology. 1999 Jul;30(1-3):181-9. doi: 10.1023/A:1008026913715.
We have developed and characterized cultures of healthy and dystrophic canine myoblasts for the evaluation of various gene transfer protocols. The number of desmin-positive myoblasts was elevated (>>80%) in cultures of myoblasts obtained from different muscle territories, the diaphragm muscle giving rise to the purest cultures. Myoblasts from dogs turned out to be a very convenient source of well transfectable and transducible cells. Transfection with plasmid DNA allowed efficient transgene expression (50% of beta-galactosidase positive cells and about 375 ng luciferase/mg protein after transfection with a calcium phosphate-precipitated plasmid). Infection with high concentrations of adenoviral and retroviral vectors allowed transgene (beta-galactosidase or mini-dystrophin) detection in about 75 to 90% of the canine cells. Therefore, primary dog myoblast cultures represent a useful in vitro model for viral and non-viral gene delivery, as well as for functional evaluation and cell grafting with applications in genetic diseases, vaccination or production of circulating therapeutic proteins.
我们已经建立并鉴定了健康和萎缩性犬肌母细胞的培养物，用于评估各种基因转移方案。从不同肌肉区域获得的肌母细胞培养物中，结蛋白阳性的肌母细胞数量增加（>>80%），膈肌肌肉产生最纯净的培养物。犬的肌母细胞是一种非常方便的、易于转染和转导的细胞来源。用质粒 DNA 转染可实现高效的转基因表达（用磷酸钙沉淀的质粒转染后，β-半乳糖苷酶阳性细胞的比例为 50%，约 375ng 荧光素酶/mg 蛋白）。用高浓度的腺病毒和逆转录病毒载体感染可使约 75%至 90%的犬细胞中检测到转基因（β-半乳糖苷酶或微型肌营养不良蛋白）。因此，原代犬肌母细胞培养物是一种用于病毒和非病毒基因传递的有用的体外模型，也可用于功能评估和细胞移植，应用于遗传疾病、疫苗接种或生产循环治疗性蛋白。
