Research Institute of Natural Sciences and Technology, Vytautas Magnus University, Universiteto str. 10, Akademija, Kaunas district LT-53361, Lithuania.
Department of Bionanoscience, Delft University of Technology, Van der Maasweg 9 2629 HZ Delft, Netherlands; Kavli Institute of Nanoscience, Delft University of Technology, Van der Maasweg 9 2629 HZ Delft, Netherlands.
Bioelectrochemistry. 2024 Aug;158:108696. doi: 10.1016/j.bioelechem.2024.108696. Epub 2024 Apr 4.
RNA interference (RNAi) is a powerful and rapidly developing technology that enables precise silencing of genes of interest. However, the clinical development of RNAi is hampered by the limited cellular uptake and stability of the transferred molecules. Electroporation (EP) is an efficient and versatile technique for the transfer of both RNA and DNA. Although the mechanism of electrotransfer of small nucleic acids has been studied previously, too little is known about the potential effects of significantly larger pDNA on this process. Here we present a fundamental study of the mechanism of electrotransfer of oligonucleotides and siRNA that occur independently and simultaneously with pDNA by employing confocal fluorescence microscopy. In contrast to the conditional understanding of the mechanism, we have shown that the electrotransfer of oligonucleotides and siRNA is driven by both electrophoretic forces and diffusion after EP, followed by subsequent entry into the nucleus within 5 min after treatment. The study also revealed that the efficiency of siRNA electrotransfer decreases in response to an increase in pDNA concentration. Overall, the study provides new insights into the mechanism of electrotransfer of small nucleic acids which may have broader implications for the future application of RNAi-based strategies.
RNA 干扰 (RNAi) 是一种强大且快速发展的技术，可实现对感兴趣基因的精确沉默。然而，RNAi 的临床发展受到转导分子细胞摄取和稳定性的限制。电穿孔 (EP) 是一种高效且多功能的技术，可用于转导 RNA 和 DNA。尽管先前已经研究了小核酸的电转移机制，但对于明显更大的 pDNA 对该过程的潜在影响知之甚少。在这里，我们通过使用共聚焦荧光显微镜对寡核苷酸和 siRNA 与 pDNA 独立和同时发生的电转移机制进行了基础研究。与对机制的有条件理解相反，我们已经表明，寡核苷酸和 siRNA 的电转移是由 EP 后的电泳力和扩散驱动的，随后在治疗后 5 分钟内进入细胞核。该研究还表明，siRNA 电转移的效率会随着 pDNA 浓度的增加而降低。总的来说，该研究为小核酸的电转移机制提供了新的见解，这可能对未来基于 RNAi 的策略的应用具有更广泛的意义。
