交联型聚乙烯亚胺智能基因载体的制备及PEG化影响  

SYNTHESIS AND CHARACTERIZATION OF A CROSSLINKING POLYETHYLENIMINE AS SMART GENE CARRIER AND EFFECTS OF PEGYLATION DEGREE

作　　者：陈磊[1] 田华雨[1] 陈学思[1] 朴泰宽 丸山厚[3] 景遐斌[1] 

机构地区：[1]中国科学院长春应用化学研究所高分子物理与化学国家重点实验室,长春130022 [2]韩国科学技术院生物科学系,大田305-701 [3]日本九州大学材料化学与工程学院,福冈812-8581

出　　处：《高分子学报》2009年第6期499-505,共7页Acta Polymerica Sinica

基　　金：国家自然科学基金-亚洲三国前瞻项目(项目号20621140369);国家自然科学基金(基金号20604028,50873102,50733003);科技部国际合作重点项目(项目号20071314)资助

摘　　要：使用胱胺双丙烯酰胺(CBA)对低分子量聚乙烯亚胺(PEI)进行交联反应制备智能降解型聚阳离子基因载体.通过与聚乙二醇(PEG)反应得到不同程度PEG化的聚阳离子载体.利用核磁、黏度测试、粒度仪、zeta电位仪和凝胶电泳对聚阳离子载体及其与DNA的复合物进行了表征.研究表明随着PEG含量的增加,聚阳离子载体/DNA复合物颗粒粒径变小、表面正电荷降低,PEG具有明显的屏蔽作用,但过多的PEG也使载体与DNA复合能力下降.通过MTT细胞毒性测试和荧光素酶质粒转染实验得出,含二硫键的交联型阳离子聚合物在测试范围内显示了非常低的细胞毒性,最佳转染效率是PEI25k的4倍,PEG化后其细胞毒性得到进一步改善,转染效率却明显降低.A smart biodegradable cationic polymer （CBA-PEI） based on the disulfide bond-containing cross-linker cystamine bisacrylamide （CBA） and low molecular weight branched polyethylenimine （1800-Da, PEI1800） was successfully synthesized by Michael addition reaction in our recent study. Furthermore, a series of copolymers （ CBA- PEI-PEG） with different PEGylation degree were obtained by the mPEG-SPA （5000-Da） reacting with CBA-PEI at various weight ratios directly. The molecular structures of the resulting polymers CBA-PEI and CBA-PEI-PEG were evaluated by nuclear magnetic resonance spectroscopy （^1H-NMR） and capillary viscosity measurements, all of which had successfully verified formation of the copolymers. The polymer/DNA complexes based on CBA-PEI and CBA- PEI-PEG were measured by dynamic light scattering and gel retardation assay. The results showed that the particle size and zeta potential of complexes were reduced with increasing amount of PEG grafting, even no particle formation. The particle size of CBA-PEI/DNA complexes was in range of 103.1 to 129.1 nm, and the zeta potential was in range of 14.2 to 24.3 mV above the 2 ： 1 weight ratio. In the same measure condition, the particle size of CBA-PEI-PEG complexes was reduced to a range of 32.2 to 55 nm, and the zeta potential was in range of 9.3 to 13.8 mV at the 2：1 weight ratio. In the gel retardation assay, CBA-PEI showed a strong DNA-binding capability. Plasmid DNA was completely retarded at the 0.1 ： 1 weight ratio. By contrast, CBA-PEI-PEG which consists of PEG/ CBA-PEI （ W/ W,38.9 % ） or PEG/CBA-PEI （ W/ W,95.8 % ） displayed a poor retardation effect even at the high weight ratio. PEG-grafted showed a shield effect in polymer/DNA complexes. However, it showed the reduced DNA binding ability if the cationic polymers were modified with too much PEGs. Moreover, the CBA-PEI and CBA-PEI- PEG were evaluated in CHO and KB cells for their cytotoxicity and transfection efficiency. MTT assay revealed that both CBA-PEI and CBA-PEI

关 键 词：基因传递 聚乙烯亚胺 二硫键 生物降解 PEG化 

分 类 号：R346[医药卫生—基础医学]