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PEI增强Pluronic介导的BALBc小鼠骨骼肌原位基因传递和表达效率
PEI enhances in situ gene delivery and expression in skeletal muscles of BALB/c mice in the Pluronic-mediated system
何雨桐,孙哲,王刚
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作者单位:四川大学生物材料工程研究中心
中文关键字:BALB/c小鼠,骨骼肌基因传递,质粒,PEI,Pluronic L64
英文关键字:BALB/c mice, skeletal muscle gene delivery, plasmid, PEI, Pluronic L64
中文摘要:目的 在Pluronic L64介导的小鼠骨骼肌基因传递体系中正确引入阳离子PEI以增强基因表达水平。方法 本实验以裸质粒DNA(pDNAs)和Pluronic L64/pDNA(L/D)为对照组,各比例PEI/pDNA与Pluronic L64混合物(L/P/D-0.5,L/P/D-3,L/P/D-10)为实验组,采用肌肉注射的方式将各组样品注射到小鼠胫骨前肌中。7天后取组织,检测报告基因表达和肌肉组织病变情况。另外,通过尾静脉注射的方式,考察L/P/D-0.5组的体内急性毒性。最后,通过粒径分析和原子力显微镜测定了L/P/D-0.5体系中PEI/pDNA复合物的粒径、电位及形貌。结果 ①报告基因表达评估:L/P/D-0.5组外源基因表达效率最高;随着N/P增加,几乎检测不到外源基因的表达。②生物相容性评估:无论原位注射还是尾静脉注射,L/P/D-0.5组均没有显示出组织病变,而其他比例的L/P/D组出现了不同程度的组织学病变。③物理特征表征:虽然在L/P/D-0.5组中DNA未被完全压缩,但PEI/pDNA复合物已拥有良好纳米粒径和形态结构,且表面电位为负。结论 L/P/D-0.5体系能安全高效地增强外源基因在骨骼肌内的表达。
英文摘要:Objective Cationic PEI is correctly introduced into Pluronic L64-mediated gene delivery system of mice skeletal muscle to enhance the efficiency of gene expression. Methods Different samples were transferred into mice tibialis anterior muscles via intramuscular injection in this experiment, among the naked plasmid DNA(pDNAs) and Pluronic L64/pDNA(L/D) were treated as the control groups, the mixtures of PEI/pDNA with different ratios and Pluronic L64 were treated as the experimental groups (L/P/D-0.5, L/P/D-3, L/P/D-10). Seven days after injection, these tissues were taken out, the gene expression and the potential pathological changes of muscle tissue were detected. In addition, the in vivo acute toxicity experiment was further executed via tail vein injection of L/P/D-0.5 group. Finally, the particle size, zeta potential and morphology of PEI/pDNA complexes in L/P/D-0.5 system were detected by DLS measurement and Atomic Force Microscope (AFM). Results ① The evaluation of report gene expression: the L/P/D-0.5 showed the highest gene expression; as N/P ratios increased, the gene expression could hardly be detected. ② The assessment of biocompatibility: no tissue lesion was observed in L/P/D-0.5 groups, regardless of muscular injection or intravenous injection. While other proportions of L/P/D showed histologic lesions with varying degrees. ③ Characterization analysis: PEI/pDNA complexes were spherical shape with nanometer size, which could contribute to gene delivery in skeletal muscle-based system, although pDNA was not fully compressed in L/P/D-0.5 group. Conclusion The L/P/D-0.5 system can safely and effectively enhance the expression of exogenous genes in skeletal muscle. Keywords: BALB/c mice, skeletal muscle gene delivery, plasmid, PEI, Pluronic L64
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国内统一连续出版物号:51-1193/Q |国际标准出版物号:1000-7083
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