Transport, resealing, and re-poration dynamics of two-pulse electroporation-mediated molecular delivery
Transport, resealing, and re-poration dynamics of two-pulse electroporation-mediated molecular delivery
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DOI：
10.1016/j.bbamem.2015.04.007
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发表时间：
2015-08-01
影响因子：
3.4
通讯作者：
Shan, Jerry W.
中科院分区：
文献类型：
--
作者：
Demiryurek, Yasir;Nickaeen, Masoud;Shan, Jerry W.
电穿孔是许多药物传递和基因治疗应用的兴趣。先前的研究表明，双脉冲电穿孔方案由短时间的高压第一脉冲和较长的低压第二脉冲组成，可以提高输送效率并保持生存能力。在这项工作中，研究了第一脉冲和第二脉冲的场强和脉冲间延迟时间对两种不同大小的荧光素-葡聚糖（FD）偶联物递送的影响。在3t3小鼠成纤维细胞上进行了一系列双脉冲电穿孔实验，第一脉冲为交流电，第二脉冲为直流电。合理设计方案，以最好地分离透性和电泳转运机制。结果表明，FD的传递随第一脉冲的强度和靶分子的大小有很大的变化。传递的FD浓度也随脉冲间延迟的对数线性下降。数据表明，电渗透后膜的重新密封发生得很快，但不可忽略的一部分孔隙可以在第二次脉冲时重新打开，延迟时间约为数百秒。假设第二个脉冲的作用不仅仅是电泳，它需要一个最小阈值场强来重新打开第一脉冲留下的纳米级孔或缺陷。这些结果表明，膜电穿孔、密封和再穿孔是一个复杂的过程，既有短期成分，也有长期成分，这可能部分解释了文献中报道的膜再密封时间的广泛差异。(C) 2015 Elsevier B.V.版权所有
Electroporation is of interest for many drug-delivery and gene-therapy applications. Prior studies have shown that a two-pulse-electroporation protocol consisting of a short-duration, high-voltage first pulse followed by a longer, low-voltage second pulse can increase delivery efficiency and preserve viability. In this work the effects of the field strength of the first and second pulses and the inter-pulse delay time on the delivery of two different-sized Fluorescein-Dextran (FD) conjugates are investigated. A series of two-pulse-electroporation experiments were performed on 3T3-mouse fibroblast cells, with an alternating-current first pulse to permeabilize the cell, followed by a direct-current second pulse. The protocols were rationally designed to best separate the mechanisms of permeabilization and electrophoretic transport. The results showed that the delivery of FD varied strongly with the strength of the first pulse and the size of the target molecule. The delivered FD concentration also decreased linearly with the logarithm of the inter-pulse delay. The data indicate that membrane resealing after electropermeabilization occurs rapidly, but that a non-negligible fraction of the pores can be reopened by the second pulse for delay times on the order of hundreds of seconds. The role of the second pulse is hypothesized to be more than just electrophoresis, with a minimum threshold field strength required to reopen nano-sized pores or defects remaining from the first pulse. These results suggest that membrane electroporation, sealing, and re-poration is a complex process that has both short-term and long-term components, which may in part explain the wide variation in membrane-resealing times reported in the literature. (C) 2015 Elsevier B.V. All rights reserved.