An efficient nonviral gene-delivery vector based on hyperbranched cationic glycogen derivatives.
An efficient nonviral gene-delivery vector based on hyperbranched cationic glycogen derivatives.
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DOI：
10.2147/ijn.s51919
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发表时间：
2014
影响因子：
8
通讯作者：
Yang L
中科院分区：
文献类型：
--
作者：
Liang X;Ren X;Liu Z;Liu Y;Wang J;Wang J;Zhang LM;Deng DY;Quan D;Yang L
本研究的目的是合成超支化阳离子糖原衍生物，并评估其作为一种有效的非病毒基因传递载体。合成了一系列超支化的阳离子糖原衍生物（DMAPA-Glyp和AEPZ-Glyp），并通过傅立叶变换红外光谱和氢核磁共振对其结构进行了表征。通过在NaCl水溶液中的酸碱滴定来评估它们的缓冲能力。使用琼脂糖凝胶电泳评估质粒脱氧核糖核酸（pDNA）的缩合能力和对糖原衍生物的DNA酶I降解的保护。测定了糖原衍生物/pDNA复合物的zeta电位和粒径，并用原子力显微镜观察了复合物的形貌。分别通过溶血试验和MTT（3-[4，5-二甲基噻唑-2-基]-2，5-二苯基四氮唑溴盐）试验评价材料的血液相容性和细胞毒性。通过流式细胞术和荧光显微镜在293 T（人胚肾）和CNE 2（人鼻咽癌）细胞系中评估阳离子糖原衍生物介导的pDNA转染效率。评估pDNA在模型动物（Sprague道利大鼠）中的体内递送以鉴定安全性和转染效率。合成了含DMAPA和AEPZ的超支化阳离子糖原衍生物。与支链聚乙烯亚胺（bPEI）相比，它们表现出更好的血液相容性和更低的细胞毒性。它们能够结合并浓缩pDNA以形成大小为100-250 nm的复合物。DMAPA-Glyp/pDNA复合物在293 T和CNE 2细胞中的转染效率均高于AEPZ-Glyp/pDNA复合物，与bPEI的转染效率几乎相等。此外，与对照组相比，DMAPA-Glyp衍生物可以更安全地将pDNA递送到Sprague道利大鼠脑组织中的血管中，然后表达为绿色荧光蛋白。超支化阳离子糖原衍生物，特别是DMAPA-Glyp衍生物，在体内外转染时表现出高的基因转染效率、良好的血液相容性和低的细胞毒性，是一种潜在的新型非病毒基因载体。
The purpose of this study was to synthesize and evaluate hyperbranched cationic glycogen derivatives as an efficient nonviral gene-delivery vector. A series of hyperbranched cationic glycogen derivatives conjugated with 3-(dimethylamino)-1-propylamine (DMAPA-Glyp) and 1-(2-aminoethyl) piperazine (AEPZ-Glyp) residues were synthesized and characterized by Fourier-transform infrared and hydrogen-1 nuclear magnetic resonance spectroscopy. Their buffer capacity was assessed by acid–base titration in aqueous NaCl solution. Plasmid deoxyribonucleic acid (pDNA) condensation ability and protection against DNase I degradation of the glycogen derivatives were assessed using agarose gel electrophoresis. The zeta potentials and particle sizes of the glycogen derivative/pDNA complexes were measured, and the images of the complexes were observed using atomic force microscopy. Blood compatibility and cytotoxicity were evaluated by hemolysis assay and MTT (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide) assay, respectively. pDNA transfection efficiency mediated by the cationic glycogen derivatives was evaluated by flow cytometry and fluorescence microscopy in the 293T (human embryonic kidney) and the CNE2 (human nasopharyngeal carcinoma) cell lines. In vivo delivery of pDNA in model animals (Sprague Dawley rats) was evaluated to identify the safety and transfection efficiency. The hyperbranched cationic glycogen derivatives conjugated with DMAPA and AEPZ residues were synthesized. They exhibited better blood compatibility and lower cytotoxicity when compared to branched polyethyleneimine (bPEI). They were able to bind and condense pDNA to form the complexes of 100–250 nm in size. The transfection efficiency of the DMAPA-Glyp/pDNA complexes was higher than those of the AEPZ-Glyp/pDNA complexes in both the 293T and CNE2 cells, and almost equal to those of bPEI. Furthermore, pDNA could be more safely delivered to the blood vessels in brain tissue of Sprague Dawley rats by the DMAPA-Glyp derivatives, and then expressed as green fluorescence protein, compared with the control group. The hyperbranched cationic glycogen derivatives, especially the DMAPA-Glyp derivatives, showed high gene-transfection efficiency, good blood compatibility, and low cyto toxicity when transfected in vitro and in vivo, which are novel potential nonviral gene vectors.