非病毒基因载体材料聚谷氨酸-聚丙烯亚胺的合成及与DNA的结合稳定性

Synthesis of Nonviral Gene Carrier Poly( L-glutamic acid)-graft-Polypropylenimine and Its Complexation Stability with DNA

目的合成非病毒基因载体材料聚谷氨酸-聚丙烯亚胺,并考查其与DNA的结合能力及稳定性。方法以G2.0聚丙烯亚胺(PPI)为引发剂,通过开环聚合的方法,合成了聚丙烯亚胺-聚谷氨酸苄脂多臂接枝聚合物,再以G1.0聚丙烯亚胺或G2.0聚丙烯亚胺作为氨解试剂进行氨解合成了新型树枝状的聚谷氨酸-聚丙烯亚胺聚合物(MP-g-PPI)。采用红外光谱(IR)、1HNMR对聚合产物进行结构表征,通过琼脂糖凝胶电泳实验考察聚合物与DNA的结合能力,通过离心法考察了聚合物与DNA形成复合物的沉淀稳定性,通过MTT法考察聚合物在Hela细胞中的毒性,并通过测定发荧光细胞百分数获得复合物在Hek-293细胞中的转染率。结果成功合成了聚谷氨酸-聚丙烯亚胺聚合物,此聚合物可有效的与DNA结合成纳米复合物,可通过一定的空间斥力阻止复合物的聚集,且毒性较PEI 25K低,可有效介导基因的转染。结论聚谷氨酸-聚丙烯亚胺聚合物可与DNA形成稳定的复合物,此聚合物有望成为新型的非病毒基因载体材料。

OBJECTIVE To synthesize nonviral gene carrier poly （L-glutamic acid） -graft-polypropylenimine （MP-g-PPI） and investigate the capability of the polymers to condense DNA and the stability of MP-g-PPI DNA complexes. METHODS A series of new multi-armed MP-g-PPI polymers were synthesized by ring-opening polymerization using G2. 0PPI as macromolecular initiator and subsequent aminolysis with G1.0PPI or G2.0PPI. The new synthetic polymers were characterized by IR and 1H-NMR. The capability of the polymers to condense DNA was evaluated by agarose gel electrophoresis, and the sedimentation stability of the complexes was also in- vestigated by centrifugal method. The transfection efficiency in Hek-293 ceils was evaluated by measuring the percentage of cells expressing green fluorecensce protein. RESULTS The MP-g-PPI polymers were successfully synthesized. The polymers could effectively condense DNA to form nanoparticles, and provide efficient steric repulsion to prevent the complexes from aggregation. Meanwhile, the polymers could effectively transfer the gene vectors. CONCLUSION The MP-g-PPI polymers can form stable complexes with DNA, and thus may be potential candidates as nonviral gene carriers.