Nanoscale Science and Engineering Center for Affordable Nanoengineering of Polymeric, Biomedical Devices, The Ohio State University, 174 West 18th Avenue, Columbus, OH, 43210, USA; Department of Chemical Engineering, Delft University of Technology, Julianalaan 136, 2628, BL, Delft, The Netherlands.
Adv Healthc Mater. 2014 May;3(5):682-9. doi: 10.1002/adhm.201300213. Epub 2013 Aug 29.
The delivery of biomolecules, including siRNAs (≈21 bp) and large plasmids (≈10 kbp), into living cells holds a great promise for therapeutic and research applications. Lipoplex nanoparticles are popular nanocarriers for gene delivery. In conventional transfection methods, the cellular uptake of lipoplex nanoparticels occurs through the endocytosis process. The entrapment of lipoplex nanoparticles into endocytic vesicle is a major barrier in achieving efficient gene silencing and expression. Here, a novel nanochannel electroporation (NEP) method is employed to facilitate the cellular uptake and release of siRNAs/DNAs from lipoplexes. First, it is demonstrated that in a NEP device, lipoplex nanoparticles can be injected directly into the cell cytoplasm within several seconds. Specifically, it is found that lipoplexes containing MCL-1 siRNA delivered by NEP can more efficiently down-regulate the expression of MCL-1 mRNA in A549 cancer cells than conventional transfection. Quantum dot-mediated Förster resonance energy transfer (QD-FRET) reveals that lipoplexes delivered via NEP can directly release siRNA in the cytoplasm without going through the endocytosis route, which unravels the responsible mechanism for efficient gene delivery. Furthermore, the advantage of combining NEP with lipoplex nanoparticles by the successful delivery of large plasmids (pCAG2LMKOSimO, 13 kbp) into CHO cells is demonstrated.
将生物分子（包括 siRNA（约 21 个碱基）和大质粒（约 10 kbp））递送到活细胞中，在治疗和研究应用方面具有很大的前景。脂质体纳米颗粒是用于基因传递的流行纳米载体。在传统的转染方法中，脂质体纳米颗粒通过内吞作用进入细胞。脂质体纳米颗粒被内吞小泡捕获是实现有效基因沉默和表达的主要障碍。在这里，采用了一种新的纳米通道电穿孔（NEP）方法来促进 siRNA/DNA 从脂质体中的细胞摄取和释放。首先，证明在 NEP 装置中，脂质体纳米颗粒可以在几秒钟内直接注入细胞质。具体而言，发现通过 NEP 递送的含有 MCL-1 siRNA 的脂质体可以更有效地下调 A549 癌细胞中 MCL-1 mRNA 的表达，优于传统转染。量子点介导的Förster 共振能量转移（QD-FRET）揭示了通过 NEP 递送的脂质体可以直接在细胞质中释放 siRNA，而无需经过内吞作用途径，这揭示了有效基因传递的负责机制。此外，通过将 NEP 与脂质体纳米颗粒结合成功地将大质粒（pCAG2LMKOSimO，13 kbp）递送到 CHO 细胞中，证明了这种方法的优势。
