小分子靶向肽修饰壳聚糖作为基因载体的研究  

Study on RGD peptide coupled chitosan as gene vector

作　　者：徐苏颖[1] 王伟[1] 丁杨[1] 周建平[1] 

机构地区：[1]中国药科大学药剂学教研室,天然药物活性组分与药效国家重点实验室,江苏南京210009

出　　处：《中国医院药学杂志》2012年第10期737-740,共4页Chinese Journal of Hospital Pharmacy

基　　金：国家"重大新药创制"科技重大专项资助项目(编号:2009ZX09310-004);国家重点基础研究发展计划(973计划)资助项目(编号:2009CB903300);国家自然科学基金项目(编号:81102398);中央高校基本科研业务费专项资金项目(编号:JKQ2009017)

摘　　要：目的:将小分子靶向肽RGD(Arg-Gly-Asp)偶联到壳聚糖(CS)上,并包载质粒DNA(pDNA),制成一种具有靶向性的壳聚糖载基因纳米粒。方法:将RGD肽上的羧基和CS上的氨基通过酰化反应发生偶联,运用红外(FT-IR)和元素分析对RGD偶联壳聚糖(CS-RGD)的化学结构进行确证;采用复凝聚法制备CS-RGD/pDNA纳米粒(CS-RGD/pDNA);应用凝胶阻滞实验和DNA酶(DNase I)降解实验考察CS-RGD对pDNA的复合和保护能力;通过激光粒度仪和原子力显微镜对纳米粒的粒径分布和形态进行考察。结果:CS和RGD肽通过酰胺键偶联;CS-RGD/pDNA在N/P≥2时完全复合,在N/P≥4时具有抗DNase I酶降解能力,N/P=2～30的CS-RGD/pDNA复合物粒径在90～260 nm之间,Zeta电位在4～39 mV之间,原子力显微镜结果证明复合物为类球形且分布良好。具有良好的稳定性和易于进入细胞的性质。结论:CS-RGD是一种制备工艺简单,具有应用前景的非病毒基因载体。OBJECTIVE To prepare the CS-RGD by the reaction of the activated RGD peptide with the amine group on the chitosan （CS）, to prepare CS-RGD loading with plasmid DNA （pDNA） nanoparticles as a targeted gene carrier. METHODS RGD peptide-coupled chitosan（CS-RGD） was prepared by the reaction of the activated RGD with the amine group on the chitosan （CS）. The structure of CS-RGD was confirmed with FT-IR and element analysis. Nanoparticles of CS-RGD with plasmid DNA （CSRGD/pDNA） were prepared using a complex coacervation process. The condensation ability and the resistance to DNase I of CS-RGD/pDNA were evaluated by agarose gel electrophoresis. And CS-RGD/pDNA was characterized for their size, zeta potential and morphologies. RESULTS RGD was coupled with CS. The results of gel electrophoresis demonstrated full binding of CS-RGD with the pDNA at N/P≥2 by electrostatic interaction. CS-RGD could protect the pDNA from degradation by DNase I at N/P≥4. The particle size and zeta potential were evaluated by Zeta sizer, which indicated complex sizes ranging from 90 - 260 nm and zeta potential ranging from 4 - 39 mV. The morphology of the CS-RGD nanoparticles was mostly spherical and well distributed. The results showed that the CS-RGD/pDNA complex was well stable and could easily enter into cells. CONCLUSION CS-RGD is easy to prepare and a promising non-viral target gene vector.

关 键 词：基因载体RGD偶联壳聚糖 凝胶电泳阻滞 CS-RGD/pDNA 

分 类 号：R944[医药卫生—药剂学]