慢病毒载体感染人主动脉血管平滑肌细胞的效率

Infection efficiency of human aorta vascular smooth muscle cells infected by lentivirus vectors

为探讨慢病毒载体感染人主动脉血管平滑肌细胞(VSMCs)株效率的最佳条件,培养人主动脉VSMCs,携带UbiEGFP-MCS-IRES-Puromycin报告基因表达元件慢病毒载体按0、1、10、50和100等不同病毒感染复数(MOI)或完全培养液、完全培养液加polybrene、感染增强液、感染增强液加polybrene等不同感染试剂进行细胞感染,48 h后荧光显微拍照,分析感染效率;MTT法测定不同MOI慢病毒载体感染48 h后细胞活性;荧光显微镜观察病毒感染后第2代和第10代细胞中荧光强度。结果显示,随MOI的增加,感染效率逐渐增加,细胞活力逐渐降低,MOI为100时,对VSMCs株具有接近90%的感染效率,添加polybrene后感染效率显著升高,完全培养液与感染增强液间无显著性差异。第10代细胞与第2代细胞相比荧光信号无差异。

To explore the infection efficiency of human aorta vascular smooth muscle cells（ VSMCs） infected by lentivirus vectors,human aorta VSMCs were cultured in vitro and infected with lentivirus vectors containing Ubi-EGFP-MCS-IRES-Puromycin. Cells were infected by lentivirus based on the multiplicity of infection（ MOI） values（0,1,10,50 and 100） or different infection solutions（ complete medium,complete medium containing polybrene,enhanced infection solution and enhanced infection solution containing polybrene）. Fluorescence microscopy was applied for determining the expression of EGFP protein after infection for 48 h. The infection efficiency was assessed by EGFP positive expression. Cell viability was determined with MTT after lentivirus infection. EGFP stable expression was detected in F2 and F10 VSMCs. The results showed that EGFP expression was increased with MOI increasing after infected with lentivirus vectors for 48 h in VSMCs;The infection efficiency was up to 90 percent after infection with lentivirus（ MOI = 100） for48 h;Polybrene in infection solution could increase the infection efficiency obviously;The cell viability decreased with MOI values increased;The EGFP expression appeared no difference between F10 cells and F2 cells.