A transfection method for short interfering RNA with the lipid-like self-assembling nanotube, A6K.
A transfection method for short interfering RNA with the lipid-like self-assembling nanotube, A6K.
复制标题
一种用类脂质自组装纳米管 A6K 转染短干扰 RNA 的方法。
DOI：
10.1007/s00795-013-0011-6
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发表时间：
2013
期刊：
影响因子：
--
通讯作者：
Teramoto A.
中科院分区：
文献类型：
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作者：
Yoshida D;Kim K;Takumi I;Yamaguchi F;Adachi K;Teramoto A.
本研究的目的是建立一种新的短干扰RNA（siRNA）转染方法。具有表面活性剂活性的纳米管，A6 K，由6个丙氨酸残基和亲水性头，赖氨酸，进行了比较，以常规的阳离子转染剂试剂siFECTOR和Lipofectamine 2000。采用MTS试验检查了人胶质母细胞瘤细胞系U87 MG、A172和T98 G的细胞毒性。荧光显微镜下观察基质金属蛋白酶（MMP）-2siRNA转染效率。电镜观察A6 K细胞的超微结构。U87 MG细胞中与A6 K相关的细胞毒性水平显著低于siFECTOR和Lipofectamine 2000。siRNA的转染效率以剂量和时间依赖性方式增加。实时定量RT-PCR分析显示，MMP-2 mRNA相对于β-actin的表达量呈剂量依赖性降低。当与siRNA混合时，A6 K的超微结构转变为胶束形成。脂质样自组装肽A6 K在与亲水性尾部相关的胶束中具有基因。这种转染方法是一种新的、稳定的技术，其细胞毒性低于目前的标准方法。
The aim of the present study was to develop a novel transfection method for short interfering RNA (siRNA). A nanotube with surfactant activity, A6K, consisting of six alanine residues and a hydrophilic head, lysine, was compared to the conventional cationic transfectant reagents siFECTOR and Lipofectamine 2000. Cytotoxicity for the human glioblastoma cell lines U87MG, A172, and T98G was examined with the MTS assay. Transfection efficiency was analyzed with FITC-labeled siRNA targeting matrix metalloproteinase (MMP)-2 mRNA by fluorescent activity on microscopy. The ultrastructure of A6K was evaluated by electron microscopy. The level of cytotoxicity associated with A6K in the U87MG cells was significantly lower than with siFECTOR and Lipofectamine 2000. Transfection efficiency for siRNA was increased in a dose- and time-dependent fashion. The relative expression of MMP-2 mRNA to β-actin was reduced in a dose-dependent manner by real-time RT-PCR analysis. The ultrastructure of the A6K was transformed to micelle formation when mixed with the siRNA. The lipid-like self-assembling peptide, A6K, has genes in the micelle associated with the hydrophilic tail. This transfection method is a novel and stable technique with lower cytotoxicity than the current standard methods.