载体因素对非病毒基因治疗体内实验方法的影响

Effect of vector factors on hGH gene therapy in vivo

目的 探讨非病毒基因治疗方法质粒载体和质粒包装载体对表达效率的影响 ,以获得一种较理想的高表达基因治疗方法。方法 构建多种人生长激素基因真核表达载体 ,分别和脂质体LipofectAMINETM 、多聚物聚乙烯吡咯烷酮 (PVP)结合 ,大鼠胫前肌肌肉注射 ,1周时放射免疫法检测血清中hGH ,注射部位肌肉匀浆后多聚酶链反应 (PCR)、逆转录 (RT) PCR检测。结果 质粒载体与LipofectAMINETM 、PVP结合后 ,1周时hGH血清均得到表达 ,0 .7～ 1.4μg/L之间 ;PCR、RT PCR均为阳性。hGH表达均显著高于裸露质粒组 (P <0 .0 5 )。结论 非病毒基因治疗骨骼肌注射方法受不同质粒载体的影响 ,与脂质体或人工多聚物结合后能显著提高表达率。

Objective To study the effect of liposome and polyplexes on the expression of human growth hormone (hGH) gene vectors in vivo.Methods Each of the three different eukaryotic expression vectors was combined with LipofectAMINE TM or 5%PVP respectively,and then was injected into anterior tibialis muscle.The gene dose of each was 100 μg.One week later,PCR and RT-PCR were performed in homogenates of the muscle,and serum hGH was detected by using radioimmunoassay.Results At first week,serum hGH could be detected in groups of the eukaryotic expression vectors in combination with LipofectAMINE TM or 5% PVP with the level ranging 0.7～1.4 μg/L averagely.PCR and RT-PCR of each group were all positive.hGH expression of naked plasmid group was obviously statistically lower (P<0.05) than that of LipofectAMINETM or PVP group.Conclusion hGH gene in eukaryotic expression vectors combined with Liposome or Polyplexes could enhance the gene expression efficiency with a method of intramuscular injection.