慢病毒介导shRNA载体定向敲除ZNF217基因表达对人神经母细胞瘤KELLY细胞侵袭和凋亡的影响  

Lentivirus-mediated shRNA Vector Targeted Knockout of ZNF217 Gene Expression on Invasion and Apoptosis of Human Neuroblastoma KELLY Cells

作　　者：唐湘莲[1] 文佳冰[1] 黎明[1] 肖雅玲[1] 李勇[1] TANG Xiang-lian;WEN Jia-bing;LI Ming(Hunan children's Hospital,Changsha 410007,Hunan)

机构地区：[1]湖南省儿童医院普外二科,湖南长沙410007

出　　处：《医学临床研究》2019年第12期2295-2298,共4页Journal of Clinical Research

基　　金：湖南省自然科学基金面上项目(2018JJ2210)。

摘　　要：【目的】探究shRNA载体定向敲除ZNF217基因表达对神经母细胞瘤KELLY细胞侵袭和凋亡的影响。【方法】构建人ZNF217基因的shRNA慢病毒载体,以神经母细胞瘤细胞系KELLY细胞为研究对象,根据实验目的,设空白对照组(A组),空白载体病毒组(B组)和ZNF217基因沉默表达载体慢病毒组(C组)。RT-PCR和Western blot检测ZNF217、Caspase-3和Caspase-9的表达水平。Transwell试剂盒检测细胞侵袭能力,AV-PI试剂盒检测细胞凋亡水平,CCK-8试剂盒检测细胞增殖能力。【结果】慢病毒转染滴度为3×10^7 TU/mL,转染率为92%以上;RT-PCR和Western blot检测结果显示,C组ZNF217的mRNA和蛋白的相对表达量要明显低于于A组和B组(P<0.05),但是Caspase-3和Caspase-9的mRNA和蛋白的相对表达量要明显高于A组和B组(P<0.05);C组细胞增殖率要明显低于B组和A组,有显著性差异(P<0.05),C组细胞侵袭能力要明显低于B组和A组,有显著性差异(P<0.05);C组细胞的凋亡率要明显高于B组和A组,有显著性差异(P<0.05)。【结论】shRNA载体定向敲除ZNF217基因表达可以抑制人神经母细胞瘤细胞系KELLY细胞的增殖和侵袭,抑制凋亡,可以作为神经母细胞瘤的潜在治疗靶点。【Objective】To investigate the effect of targeted knockout of ZNF217 gene expression by shRNA vector on invasion and apoptosis of human neuroblastoma Kelly.【Methods】The shRNA hairpin structure of ZNF217 gene was constructed by siRNA technology.The human neuroblastoma cell line Kelly cells were divided into blank control group(group A),blank vector virus group(group B)and ZNF217 gene silencing expression vector lentivirus group(Group C).The expression levels of ZNF217,caspase-3 and caspase-9 were detected by RT-PCR and Western blot.Transwell kit was used to detect cell invasion,AV-PI kit to detect apoptosis,CCK-8 kit to detect cell proliferation.【Results】The transfection titer of lentivirus was 3×10^7 TU/mL,and the transfection efficiency was over 92%.RT-PCR and Western blot showed that the relative expression of ZNF217 mRNA and protein in the group C was lower than that in the group A and B(P<0.05).However,the relative expression of Caspase-3 and caspase-9 in the group C was higher than that in the group A and B(P<0.05).The cell proliferation rate of the group C was significantly lower than that of the group B and the group A(P<0.05),and the cell invasion ability of the group C was significantly lower than that of the group B and the group A(P<0.05).The apoptosis rate of the group C was significantly higher than that of the group B and the group A(P<0.05).【Conclusion】Targeted knockout of ZNF217 gene expression by shRNA vector can inhibit the proliferation,invasion and apoptosis of human neuroblastoma cell line Kelly cells,so ZNF217 gene can be used as a potential therapeutic target for neuroblastoma.

关 键 词：神经母细胞瘤/遗传学 神经母细胞瘤/病理学 细胞增殖 

分 类 号：R730.264.4[医药卫生—肿瘤]