Interaction of DNA/nuclear protein/polycation and the terplexes for gene delivery
Interaction of DNA/nuclear protein/polycation and the terplexes for gene delivery
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DNA/核蛋白/聚阳离子与基因传递三联体的相互作用
DOI：
10.1088/0957-4484/21/4/045102
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发表时间：
2010-01-29
期刊：
影响因子：
3.5
通讯作者：
Wu, Chuanbin
中科院分区：
文献类型：
--
作者：
Shen, Yuan;Peng, Hui;Wu, Chuanbin
外源DNA的核转运是非病毒基因递送的主要障碍，需要在新载体的设计中加以解决。在本研究中，我们制备了pDNA/HMGB 1/PEG-PEI三元复合物以促进核输入。三聚体中的HMGB 1用于辅助pDNA转运到细胞核中，因为它含有核定位信号（NLS）;引入PEG链以稳定pDNA/载体三聚体并降低细胞毒性。HMGB 1/PEG-PEI组合载体的结构相互作用已经通过原子力显微镜和圆二色性光谱进行了专门研究。结果表明，HMGB 1分子能与pDNA链结合，但不能很好地缩合pDNA。PEG-PEI进一步将pDNA/HMGB 1复合物压缩成纳米级球形三聚体。激光共聚焦显微镜观察发现，HMGB 1/PEG-PEI复合载体转染的pDNA在细胞核内有明显的聚集。HMGB 1/PEG-PEI诱导的GFP转染细胞百分比和VEGF蛋白表达水平分别是普通阳离子聚合物PEI 25 kDa的2.6-4.9倍和1.4-2.8倍。因此，HMGB 1/PEG-PEI组合载体可作为一种多功能载体，用于促进外源DNA核定位，从而提高其表达。
Nuclear transport of exogenous DNA is a major barrier to nonviral gene delivery that needs to be addressed in the design of new vectors. In this study, we prepared pDNA/HMGB1/PEG-PEI terplexes to promote nuclear import. HMGB1 in the terplexes was used to assist the transportation of pDNA into the nucleus of cells, since it contained nuclear localization signal (NLS); PEG chains were introduced to stabilize pDNA/vector terplexes and reduce the cytotoxicity. HMGB1/PEG-PEI combined vectors have been investigated specifically for their structure interaction by atomic force microscopy and circular dichroic spectroscopy. The results demonstrated that the HMGB1 molecule could bind with the pDNA chains, but not condense pDNA well. The PEG-PEI further compacted pDNA/HMGB1 complexes into nanosized spherical terplexes. The pDNA delivered by HMGB1/PEG-PEI combined vectors was significantly accumulated in the nucleus of cells, as observed by confocal laser scanning microscopy. The percentage of GFP-transfected cells and VEGF protein expression level induced by HMGB1/PEG-PEI were 2.6-4.9-fold and 1.4-2.8-fold higher, respectively, than that of a common cationic polymer PEI 25 kDa. Therefore, the HMGB1/PEG-PEI combined vector could be used as a versatile vector for promoting exogenous DNA nuclear localization, thereby enhancing its expression.