Aránega A, Marchal J A, Melguizo C, Prados J, Aránega A E, Vélez C, Fernández J E, Arena N, Alvarez L
Department of Morphological Sciences, School of Medicine, University of Granada, Spain.
Cell Mol Biol (Noisy-le-grand). 1996 Dec;42(8):1219-27.
Gene transfection has been accomplished with a variety of techniques such as DEAE dextran, calcium phosphate coprecipitation, protoplast fusion, liposomes, microinjection and recombinant bacteriophages. However, transfection by electroporation, consisting of the reversible permeabilization of cell membranes after exposure to a pulsed electric field, has been shown to be the most rapid, simple and efficient method for the stable incorporation of genes in different cell lines. We studied rhabdomyosarcoma cells subjected to electroporation in two different vol. [400 microliters (group 1) and 150 microliters (group 2] of 140 mM NaCl/15 mM Hepes buffer, pH 7.2) and evaluated the effects of electroporation volume on growth and differentiation. Low sample volumes induced a terminal process of morphological and ultrastructural myogenic differentiation in rhabdomyosarcoma cells, which concluded with cell death. Our results suggest that in electroporation low sample vol. of rhabdomyosarcoma cells induced morphological and phenotypic differentiation, with increased expression of desmin, alpha-actinin and tropomyosin.
基因转染已通过多种技术实现，如DEAE葡聚糖法、磷酸钙共沉淀法、原生质体融合法、脂质体法、显微注射法和重组噬菌体法。然而，电穿孔转染，即在暴露于脉冲电场后使细胞膜可逆性通透，已被证明是将基因稳定导入不同细胞系的最快速、简单且有效的方法。我们研究了横纹肌肉瘤细胞在两种不同体积[400微升（第1组）和150微升（第2组）]的140 mM NaCl/15 mM Hepes缓冲液（pH 7.2）中进行电穿孔的情况，并评估了电穿孔体积对生长和分化的影响。低样本体积诱导横纹肌肉瘤细胞发生形态学和超微结构的肌源性终末分化过程，最终导致细胞死亡。我们的结果表明，在电穿孔过程中，横纹肌肉瘤细胞的低样本体积诱导了形态学和表型分化，同时结蛋白、α-辅肌动蛋白和原肌球蛋白的表达增加。
