重组逆转录病毒介导的人载脂蛋白AI基因在肌源细胞中的表达

The Expression of Human Apolipoprotein AI in Myogenic Cells by Retrovirus Vector

目的 探索一种安全有效的载脂蛋白 AI基因表达系统。方法 构建含人载脂蛋白 AI c DNA的双顺反子逆转录病毒载体 p L AEN,用其转化小鼠原代肌母细胞及肌源性细胞 C2 C12 ,EL ISA法检测细胞液中人载脂蛋白 AI的含量。结果 转化后的小鼠原代肌母细胞及 C2 C12细胞在分化为肌管细胞后均获得异源表达人载脂蛋白 AI的能力 ;而且经 G418筛选也获得稳定转化的 C2 C12细胞株 ,30天后仍保持有效表达人载脂蛋白 AI的能力。结论 提示肌源性细胞在体外 ,以含人载脂蛋白 AI c DNA的逆转录病毒载体进行基因修饰后再移植回骨骼肌 。

Objective To observe the expression of apolipoprotein AI in transduced cultured muscle cells.Methods Bicistronic retrovirus vector containing human apolipoprotein AI cDNA were constructed and packaged it into GP+E 86 and AM12 cell lines which was selected by G418. G418 resistant clone with high virus producing titer was isolated. Mouse primary myoblasts and C2C12 mouse myoblasts were infected with recombinant virus supernatant. The content of apolipoprotein AI in cell culture medium was detected by ELISA.Results All of transduced mouse primary myoblasts and C2C12 cells gained and remained the ability for heterologous expression of human apolipoprotein AI even after differentiation into myotubes. Moreover, stable transduced C2C12 cell clones were generated by G418 selection, which effectively expressed human apolipoprotein AI up to 30 days.Conclusion The use of retrovirus vectors to genetically modify myoblasts and then implantation back to skeletal muscle might be a safe and feasible strategy for gene therapy of atherosclerosis.