摘要
目的研究纳米粒子PLGA携带特异性HIF-1α基因转染的可行性及其效率。方法构建HIF-1α基因的真核表达质粒pcDNA3.1(+)的重组体。应用复乳溶剂挥发法制备pcDNA3.1(+)-HIF-1α的PLGA纳米粒,用正交设计法对纳米粒子的制备工艺进行优化,检测其包埋率、体外释放情况及粒度。观察纳米粒的形态、大小和粒度,研究体外释药特性、抗核酸酶降解性能。用PLGA-pcDNA3.1(+)-HIF-1α纳米粒转染U251细胞,Western blot检测PLGA纳米粒子中HIF-1α蛋白表达。结果用正交设计法优化制备工艺成功构建pcDNA3.1(+)-HIF-1α的PLGA纳米粒,形态圆整,大小均匀,平均粒径102 nm,含药量0.83%,包封率可达72%,有对抗核酸酶降解能力,体外释药缓慢。PLGA-pcDNA3.1(+)-HIF-1α纳米粒可持续高效表达HIF-1α蛋白。结论 PLGA-pcDNA3.1(+)-HIF-1α纳米粒可用于特异性基因的转染。
Objective To study the possibility and efficiency of PLGA nanoparticle as a vector of Hif-1α in specific gene transfection.Methods pcDNA3.1（＋）-HIF-1α plasmid and PLGA nanoparticles carrying pcDNA3.1（＋）-HIF1α plasmid were constructed.HIF-1α cDNA was subcloned into pcDNA3.1（＋） plasmid.PLGA nanoparticles with water/ oil/ water double emulsion-solvent evaporation were prepared.The orthogonal design was taken for improving and perfecting nanoparticle preparation process,observing and assessing the shape,size and diameter distribution of PLGA nanoparticles.The in vitro release nature of carried drug and its anti-nuclease degradation features were investigated.PLGA nanoparticles carrying pcDNA3.1（＋）-HIF-1α plasmid were used to transfect U251 cells.HIF-1α protein expression was detected by Western blot.Results PLGA nanoparticles carrying pcDNA3.1（＋）-HIF-1α plasmid were constructed,which were uniform and round with an average diameter of about 102nm.The pcDNA3.1（＋）-HIF-1α loading in the nanoparticles was 0.83% and the encapsulation ratio reached 72%,which had anti-nuclease degradation properties.The in vitro drug release was slow.The PLGA nanoparticles carrying pcDNA3.1（＋）-HIF-1α plasmid could continuously over-express HIF-1α protein.Conclusion PLGA-pcDNA3.1（＋）-HIF-1α plasmid can be used as a vector of HIF-1α to transfect specific gene.
出处
《江苏医药》
CAS
CSCD
北大核心
2010年第22期2669-2672,共4页
Jiangsu Medical Journal
基金
江苏省科委自然基金科研课题(bk2003025)
