The State Key Laboratory of Power Transmission Equipment & System Security and New Technology, School of Electrical Engineering, Chongqing University, Chongqing, China.
PLoS One. 2018 May 24;13(5):e0197167. doi: 10.1371/journal.pone.0197167. eCollection 2018.
Traditionally, microsecond pulsed electric field was widely used in cell electrofusion technology. However, it was difficult to fuse the cells with different sizes. Because the effect of electroporation based on microsecond pulses was greatly influenced by cell sizes. It had been reported that the differences between cell sizes can be ignored when cells were exposed to nanosecond pulses. However, pores induced by those short nanosecond pulses tended to be very small (0.9 nm) and the pores were more easy to recover. In this work, a finite element method was used to simulate the distribution, radius and density of the pores. The innovative idea of "cell electrofusion based on nanosecond/microsecond pulses" was proposed in order to combine the advantages of nanosecond pulses and microsecond pulses. The model consisted of two contact cells with different sizes. Three kinds of pulsed electric fields were made up of two 100-ns, 10-kV/cm pulses; two 10-μs, 1-kV/cm pulses; and a sequence of a 100-ns, 10-kV/cm pulse, followed by a 10-μs, 1-kV/cm pulse. Some obvious advantageous can be found when nanosecond/microsecond pulses were considered. The pore radius was large enough (70nm) and density was high (5×1013m-2) in the cell junction area. Moreover, pores in the non-contact area of the cell membrane were small (1-10 nm) and sparse (109-1012m-2). Areas where the transmembrane voltage was higher than 1V were only concentrated in the cell junction. The transmembrane voltage of other areas were at most 0.6V when we tested the rest of the cell membrane. Cell fusion efficiency can be improved remarkably because electroporation was concentrated in the cell contact area.
传统上，微秒脉冲电场被广泛应用于细胞电融合技术中。然而，对于不同大小的细胞融合却存在一定的困难。这是因为基于微秒脉冲的电穿孔效果受到细胞大小的极大影响。据报道，当细胞暴露在纳秒脉冲下时，可以忽略细胞大小的差异。然而，这些短纳秒脉冲诱导的孔通常很小（0.9nm），并且更容易恢复。在这项工作中，使用有限元方法模拟了孔的分布、半径和密度。为了结合纳秒脉冲和微秒脉冲的优势，提出了“基于纳秒/微秒脉冲的细胞电融合”的创新思想。该模型由两个具有不同大小的接触细胞组成。三种脉冲电场由两个 100ns、10kV/cm 的脉冲和两个 10μs、1kV/cm 的脉冲组成；或者由一个 100ns、10kV/cm 的脉冲，后面跟着一个 10μs、1kV/cm 的脉冲序列组成。当考虑纳秒/微秒脉冲时，可以发现一些明显的优势。在细胞连接区域，孔半径足够大（70nm）且密度较高（5×1013m-2）。此外，细胞膜非接触区域的孔较小（1-10nm）且稀疏（109-1012m-2）。跨膜电压高于 1V 的区域仅集中在细胞连接区。当测试细胞膜的其余部分时，其他区域的跨膜电压最高可达 0.6V。由于电穿孔集中在细胞接触区域，因此可以显著提高细胞融合效率。
