RNA interference in vitro and in vivo using a chitosan/siRNA nanoparticle system
RNA interference in vitro and in vivo using a chitosan/siRNA nanoparticle system
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DOI：
10.1016/j.ymthe.2006.04.010
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发表时间：
2006-10-01
影响因子：
12.4
通讯作者：
Kjems, Jorgen
中科院分区：
文献类型：
--
作者：
Howard, Kenneth A.;Rahbek, Ulrik L.;Kjems, Jorgen
本工作介绍了一种新型的基于壳聚糖的siRNA纳米颗粒递送系统，用于体外和体内的RNA干扰。使用原子力显微镜和光子相关光谱法显示了siRNA双链体（21-mer）和壳聚糖聚合物之间形成纳米颗粒（范围从40至600 nm）的插入复合物。使用荧光显微镜观察Cy 5标记的纳米颗粒快速摄取（1小时）到NIH 3 T3细胞中，随后在24小时内积累。在H1299人肺癌细胞和小鼠腹腔巨噬细胞中证实了纳米颗粒介导的内源性增强型绿色荧光蛋白（EGFP）的敲低（EGFP荧光分别降低77.9%和89.3%）。此外，Western分析显示，在使用含有对BCR/ABL-1连接序列特异性的siRNA的纳米颗粒转染后，K562（Ph+）细胞中BCR/ABL-1白血病融合蛋白的表达降低了约90%，而BCR表达未受影响。在鼻内施用壳聚糖/siRNA制剂后，在转基因EGFP小鼠的细支气管上皮细胞中实现了有效的体内RNA干扰（与错配和未处理的对照相比，分别减少37%和43%）。这些发现突出了这种新型的基于壳聚糖的系统在RNA介导的全身性和粘膜疾病的治疗中的潜在应用。
This work introduces a novel chitosan-based siRNA nanoparticle delivery system for RNA interference in vitro and in vivo. The formation of interpolyelectrolyte complexes between siRNA duplexes (21-mers) and chitosan polymer into nanoparticles, ranging from 40 to 600 nm, was shown using atomic force microscopy and photon correlation spectroscopy. Rapid uptake (I h) of Cy5-labeled nanoparticles into NIH 3T3 cells, followed by accumulation over a 24 h period, was visualized using fluorescence microscopy. Nanoparticle-mediated knockdown of endogenous enhanced green fluorescent protein (EGFP) was demonstrated in both H1299 human lung carcinoma cells and murine peritoneal macrophages (77.9% and 89.3% reduction in EGFP fluorescence, respectively). in addition, Western analysis showed similar to 90% reduced expression of BCR/ABL-1 leukemia fusion protein while BCR expression was unaffected in K562 (Ph+) cells after transfection using nanoparticles containing siRNA specific to the BCR/ABL-1 junction sequence. Effective in vivo RNA interference was achieved in bronchiole epithelial cells of transgenic EGFP mice after nasal administration of chitosan/siRNA formulations (37% and 43% reduction compared to mismatch and untreated control, respectively). These findings highlight the potential application of this novel chitosan-based system in RNA-mediated therapy of systemic and mucosal disease.