Tahara Kohei, Sakai Takeshi, Yamamoto Hiromitsu, Takeuchi Hirofumi, Hirashima Naohide, Kawashima Yoshiaki
Laboratory of Pharmaceutical Engineering, School of Pharmacy, Aichi Gakuin University, Chikusa, Nagoya, Japan.
Chem Pharm Bull (Tokyo). 2011;59(3):298-301. doi: 10.1248/cpb.59.298.
This study sought to evaluate the in vitro transfection efficiency of plasmid DNA (pDNA)-loaded chitosan-modified poly(DL-lactide-co-glycolide) nanospheres (CS-PLGA NS) in a gene-delivery system. Using the emulsion solvent diffusion (ESD) method, pDNA-loaded PLGA NS was prepared and the surface of the PLGA NS was modified by binding to CS. Gene transfection ability of CS-PLGA NS was examined in A549 cells. The luciferase gene was used as a reporter gene. The pattern of luciferase activity by pDNA-loaded CS-PLGA NS was initially weak, but gradually grew stronger before decreasing activity. These phenomena should be in accordance with the sustained-release profile of pDNA from PLGA NS in the cytosol and the pDNA protection against DNase. Positively charged CS-PLGA NS was found, by 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium, inner salt (MTS) assay, not to exhibit cytotoxicity on A549 cells. These results suggest that CS-PLGA NS are potential contributors to efficient pDNA delivery due to their increased interactions with cells and lack of cytotoxic effects.
本研究旨在评估载有质粒DNA（pDNA）的壳聚糖修饰聚（DL-丙交酯-共-乙交酯）纳米球（CS-PLGA NS）在基因递送系统中的体外转染效率。采用乳液溶剂扩散（ESD）法制备了载有pDNA的PLGA NS，并通过与壳聚糖结合对PLGA NS的表面进行修饰。在A549细胞中检测了CS-PLGA NS的基因转染能力。使用荧光素酶基因作为报告基因。载有pDNA的CS-PLGA NS的荧光素酶活性模式最初较弱，但在活性降低之前逐渐增强。这些现象应与pDNA在细胞质中从PLGA NS的缓释特性以及pDNA对脱氧核糖核酸酶的保护作用一致。通过3-（4,5-二甲基噻唑-2-基）-5-（3-羧甲氧基苯基）-2-（4-磺基苯基）-2H-四唑内盐（MTS）测定发现，带正电荷的CS-PLGA NS对A549细胞不表现出细胞毒性。这些结果表明，CS-PLGA NS由于其与细胞的相互作用增加且缺乏细胞毒性作用，是高效pDNA递送的潜在贡献者。
