Survivin启动子驱动LRIG1基因表达的重组腺病毒对人膀胱癌细胞BIU87生长的影响

Effect of Recombinant Adenoviral Vector Carrying LRIG1 Gene Driven by Survivin Promoter(Ad-Surp-LRIGl) on Growth of Human Bladder Cancer Cell Line BIU87

目的:构建Survivin启动子驱动LRIG1基因表达的重组腺病毒Ad-Surp-LRIGl,并检测其对人膀胱癌细胞BIU87生长的影响。方法:将经同源重组产生的重组腺病毒Ad-Surp-LRIGl在293细胞中大量扩增并通过氯化铯密度梯度离心法纯化,测其滴度。用Ad-Surp-LRIGl和Ad-LRIGl分别感染人膀胱癌细胞BIU87及人膀胱上皮永生化细胞SV-HUC-1,MTT法评估Ad-Surp-LRIGl和Ad-LRIGl对BIU87细胞生长的抑制作用。结果:当感染复数(MOI)=25时,Ad-Surp-LRIGl在BIU87细胞中的转染效率为74.56%,在SV-HUC-1细胞中转染率为0,差异有统计学意义(χ2=58.64,P<0.05);Ad-LRIGl在BIU87细胞中的转染效率为68.27%,在SV-HUC-1细胞中转染率为72.52%,差异无统计学意义(χ2=0.075,P>0.05)。Ad-Surp-LRIGl和Ad-LRIGl在BIU87细胞中的转染效率相比,差异无统计学意义(χ2=0.016,P>0.05)。与PBS相比,Ad-Surp-LRIGl和Ad-LRIGl均能明显抑制BIU87细胞的生长,转染后第4天这一差异显著(F=15.96,P<0.05),而Ad-Surp-LRIGl组和Ad-LRIGl组细胞生长速度无明显差异。结论:成功构建了含有Survivin启动子驱动LRIG1基因的重组复制腺病毒Ad-Surp-LRIGl并鉴定正确,Ad-Surp-LRIGl能选择性转染人膀胱癌细胞BIU87,在体外能明显抑制膀胱癌细胞BIU87的生长。

Objective To construct the recombinant adenoviral vector Ad-Surp-LRIG1 carrying LRIG1 gene driven by Survivin promoter, and to explore the effect of the vector on the growth of human bladder cancer cell line BIU87. Methods Ad-Surp-LRIG1 was propagated in 293 cells and purified by cesium chloride gra- dient centrifugation. Human bladder cancer cell line BIU87 and immortalized human bladder epithelial cells SV-HUC-1 were infected with Ad-Surp-LRIG1 and Ad-LRIG, respectively. The selective infection efficiency of Ad-Surp-LRIG1 and Ad-LRIG were evaluated by checking the expression of EGFP. The MTT method was used to test cell growth. Results Identified by restriction endonuclease analysis and PCR, recombinant adenovirus with Survivin promoter and LRIG1 gene was constructed successfully. The transfection efficiency of Ad-Surp-LRIG1 was 74.56% in BIU87 cells and 0 in SV-HUC-1 cells （X^2=58.64, P 〈 0.05）,while the transfec- tion efficiency of Ad-LRIG was 68.27% in BIU87 cells and 72.52% in SV-HUC-1 cells （X^2=0.075,P〉 0.05）. The transfection efficiency difference of Ad-Surp-LRIG1 and Ad-LRIG in BIU87 cells wasn＇t statistically signif- icant （X^2=0.016, P 〉 0.05）.Compared with PBS, Ad-Surp-LRIG1 and Ad-LRIG1 could inhibit BIU87 cell growth, the difference was significant 4 days after transfection（F=15.96, P 〈 0.05）. There wasn＇t significant dif- ference in cell growth rates of Ad-Surp-LRIG1 group and Ad-LRIG1 group. Conclusion The recombinant ad- enovirus carrying LRIG1 gene driven by Survivin promoter has been constructed successfully, which could inhib- it significantly the growth of bladder cancer cell line BIU87 in vitro.