载鱼精蛋白-pDNA复合物固体脂质纳米粒的初步研究

 目的制备非病毒基因载体载鱼精蛋白-pDNA复合物的固体脂质纳米粒(PD-SLN),PD-SLN由多聚阳离子缩合DNA内核与一个脂质外壳组成,从而构成多功能信封式纳米载体(multifunctional envelope-type nano device,MEND)结构,研究其特征,对DNA的保护作用以及DNA的体外释放性质。方法分别采用溶剂扩散法和复乳法制备纳米粒;用透射电镜观察形态;用Zeta电位测定仪测定粒径、多分散指数和Zeta电位;用荧光分光光度法测定基因包封率;分别用琼脂糖凝胶电泳观察复合物及PD-SLN保护pDNA抵抗剧烈外力和核酸酶的降解情况;采用双室扩散法对PD-SLN进行体外释放研究。结果用2种方法制备的SLN呈球形和类球形,平均粒径分别为(231±13.7)和(627±22.9)nm,Zeta电位分别为(-17.8±3.2)和(-25.2±2.7)mV,包封率分别为(41.5±3.62)%和(56.5±5.28)%。pDNA保护性试验表明,PD-SLN对pDNA有保护作用。体外释放实验结果表明PD-SLN缓释能力强。结论PD-SLN是一种制备工艺简单,体外缓释能力好,对pDNA保护性强,具有一定应用前景的非病毒基因载体。

Preliminary Studies on Protamine-pDNA Complex Loaded Solid Lipid Nanoparticles

OBJECTIVE To prepare nonviral vector of solid lipid nanoparticles carrying protamine-pDNA complex(PD-SLN),to develop a Multifunctional Envelope-type Nano Device(MEND) structure,in which the core of a plasmid DNA,condensed by a polycation,is encapsulated by a lipid envelope,then the nanoparticles' characteristics,the protection ability and the DNA release property in vitro were evaluated.METHODS PD-SLN was prepared by emulsion solvent diffusion method and double emulsion solvent evaporation method,respectively.The morphology of PD-SLN was observed by transmission electron microscopy.The particle sizes,polydispersity,Zeta potential were measured by nanoparticle size analyser.Encapsulating efficiency of pDNA was determined by fluorescence spectrometer.The protection of complex and PD-SLN from intensive force and nuclease degradation were evaluate by agarose gel electrophoresis,respectively.The method of two-compartment diffusion was employed to investigate the releasing character of DNA from PD-SLN.RESULTS The morphology of PD-SLN were approximately spherical.The average partic1e sizes of PD-SLN obtained by the two different methods were(231±13.7) and(627±22.9)nm,respectively.The Zeta potentials were(-17.8±3.2) and(-25.2±2.7)mV,respectively.Encapsulating efficiencies were(41.5±3.62)% and(56.5±5.28)%,respectively.The intensive agitation and nuclease degradation test results confirmed that the pDNA was protected considerably.PD-SLN Maintained sustained-release of pDNA for several days in vitro.CONCLUSION PD-SLN could be prepared easily with small particle sizes,excellent sustained-release activity,and protection of pDNA.In vitro studies have showed that PD-SLN could be a promising device,which has the potential to make in vivo cancer gene therapy achievable.