Golzio M, Mora M P, Raynaud C, Delteil C, Teissié J, Rols M P
Institut de Pharmacologie et de Biologie Structurale du CNRS UPR 9062, Toulouse, France.
Biophys J. 1998 Jun;74(6):3015-22. doi: 10.1016/S0006-3495(98)78009-9.
Cells can be transiently permeabilized by a membrane potential difference increase induced by the application of high electric pulses. This was shown to be under the control of the pulsing buffer osmotic pressure, when short pulses were applied. In this paper, the effects of buffer osmotic pressure during electric treatment and during the following 10 min were investigated in Chinese hamster ovary cells subjected to long (ms) square wave pulses, a condition needed to mediate gene transfer. No effect on cell permeabilization for a small molecule such as propidium iodide was observed. The use of a hypoosmolar buffer during pulsation allows more efficient loading of cells with beta-galactosidase, a tetrameric protein, but no effect of the postpulse buffer osmolarity was observed. The resulting expression of plasmid coding for beta-galactosidase was strongly controlled by buffer osmolarity during as well as after the pulse. The results, tentatively explained in terms of the effect of osmotic pressure on cell swelling, membrane organization, and interaction between molecules and membrane, support the existence of key steps in plasmid-membrane interaction in the mechanism of cell electrically mediated gene transfer.
通过施加高电脉冲诱导的膜电位差增加，细胞可被短暂通透化。当施加短脉冲时，这显示受脉冲缓冲液渗透压的控制。在本文中，研究了在接受长（毫秒）方波脉冲的中国仓鼠卵巢细胞中，电处理期间及随后10分钟内缓冲液渗透压的影响，这是介导基因转移所需的条件。对于小分子如碘化丙啶，未观察到对细胞通透化的影响。脉动期间使用低渗缓冲液可使细胞更有效地加载β-半乳糖苷酶（一种四聚体蛋白），但未观察到脉冲后缓冲液渗透压的影响。编码β-半乳糖苷酶的质粒的表达在脉冲期间及之后均受缓冲液渗透压的强烈控制。这些结果初步根据渗透压对细胞肿胀、膜组织以及分子与膜之间相互作用的影响进行了解释，支持了细胞电介导基因转移机制中质粒-膜相互作用关键步骤的存在。
