School of Pharmacy, Tokyo University of Pharmacy and Life Sciences.
School of Pharmaceutical Sciences, University of Shizuoka.
Biol Pharm Bull. 2021;44(12):1866-1871. doi: 10.1248/bpb.b21-00559.
In this study, we focused on a nuclear localization signal (NLS)-based versatile peptide vector, designed by us, combined with electroporation (EP) to establish an efficient gene delivery system to non-dividing or slow growing dendritic cells. We determined the intranuclear transport, gene expression, and cell viability in JAWS II mouse dendritic cells transfected with the green fluorescent protein (GFP) expression plasmid DNA alone (naked pEGFP); positive charged complex of NLS derivative STR-CH2SV40H2C, and pEGFP (binary complex); or negative charged complex of the binary complex with a biocompatible polyanion, γ-polyglutamic acid (ternary complex) combined with or without EP application. Although the binary complex showed higher nuclear transport and GFP expression in the absence of EP than those for naked pEGFP, the combination of EP significantly decreased the cell viability and did not improve the efficiency of compared gene expression. However, in the ternary complex, the intranuclear transport and GFP expression efficiency were significantly higher than those of naked pEGFP and the binary complex when combined with EP, and there was no decrease in cell viability. The results suggest that polyanion-coated ternary complex with EP is useful for non-viral gene delivery system into non-dividing cells for ex vivo gene/cell therapy.
在这项研究中，我们专注于一种基于核定位信号（NLS）的多功能肽载体，由我们设计，与电穿孔（EP）相结合，建立一种有效的基因传递系统，用于非分裂或生长缓慢的树突状细胞。我们确定了 JAWS II 小鼠树突状细胞中绿色荧光蛋白（GFP）表达质粒 DNA （裸 pEGFP）；NLS 衍生物 STR-CH2SV40H2C 和 pEGFP 的正电荷复合物（二元复合物）；或与生物相容的聚阴离子γ-聚谷氨酸（三元复合物）形成的二元复合物的带负电荷的复合物，与或不与 EP 应用结合时的核内转运、基因表达和细胞活力。虽然二元复合物在没有 EP 的情况下显示出比裸 pEGFP 更高的核内转运和 GFP 表达，但 EP 的组合显着降低了细胞活力，并且没有提高基因表达的效率。然而，在三元复合物中，与 EP 结合时，核内转运和 GFP 表达效率均明显高于裸 pEGFP 和二元复合物，且细胞活力没有下降。结果表明，与 EP 结合的带负电荷的聚阴离子包覆的三元复合物对于非分裂细胞的非病毒基因传递系统用于体外基因/细胞治疗是有用的。
