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microRNA-21靶向调控Wnt2基因对肝癌细胞HepG2增殖与迁移的影响
Effect of targeted regulation of the Wnt2 gene by microRNA(microRNA-21) on the proliferation and migration of HepG2 hepatoma cells
文章发布日期:2020年04月10日  来源:  作者:王泽鑫,关利君,李建明,等  点击次数:272次  下载次数:48次

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【摘要】:目的 探讨分析microRNA-21(miR-21)靶向调控Wnt2基因对肝癌细胞HepG2增殖与迁移的影响。方法 采用实时荧光定量PCR法检测肝癌细胞HepG2及正常肝细胞株LO2 miR-21的表达情况。对HepG2细胞转染miR-21抑制剂,分析转染后抑制剂组与对照组miR-21的表达情况、细胞增殖、迁移及凋亡情况,检测两组细胞Wnt2蛋白表达水平,并采用双荧光素酶报告基因实验验证miR-21与Wnt2基因的关系。计量资料两组间比较采用t检验。结果 HepG2细胞miR-21相对表达水平明显高于LO2细胞(1.978±0.035 vs 1.586±0.022,t=16.424, P<0.05)。转染miR-21抑制剂后,抑制剂组miR-21相对表达水平较对照组显著降低(0.857±0.017 vs 1.684±0.039,t=33.669, P<0.05)。转染miR-21抑制剂24、48、72 h后,抑制剂组HepG2细胞增殖能力均较对照组显著降低(P值均<0.05);抑制剂组穿过Tranwell小室的细胞数显著低于对照组(83.72±15.06 vs 147.85±20.64,t=4.347, P<0.05);抑制剂组细胞凋亡率明显高于对照组(25.67%±3.95% vs 10.27%±2.14%,t=5.937, P<0.05)。抑制剂组HepG2细胞的Wnt2相对表达水平明显低于对照组(0.862±0.127 vs 1.306±0.218,t=3.048, P<0.05)。TargetScan软件显示,miR-21抑制剂可显著抑制野生型Wnt2-3′ UTR质粒转染细胞的荧光素酶活性(0.972±0.102 vs 0.612±0.092,t=4.219,P<0.05),而对突变型Wnt2-3′UTR质粒转染细胞的荧光素酶活性并无明显影响(0.982±0.093 vs 0.911±0.128,t=0.972,P>0.05)。结论 抑制miR-21表达可有效抑制HepG2细胞增殖和迁移,促进HepG2细胞凋亡,并抑制Wnt信号通路的过度激活,可能成为肝癌治疗的潜在靶基因之一。
【Abstract】:Objective To investigate the effect of targeted regulation of the Wnt2 gene by microRNA(miR-21) on the proliferation and migration of HepG2 hepatoma cells. Methods Quantitative real-time PCR was used to measure the mRNA expression of miR-21 in HepG2 hepatoma cells and normal liver cell line LO2. HepG2 cells were transfected with miR-21 inhibitor, and then the expression of miR-21 and cell proliferation, migration, and apoptosis were analyzed for the inhibitor group and the control group. The protein expression of Wnt2 was measured for the two groups, and dual-luciferase reporter assay was used to verify the association between miR-21 and the Wnt2 gene. The t-test was used for comparison of continuous data between groups. Results The relative expression of miR-21 in HepG2 cells was significantly higher than that in LO2 cells (1.978±0.035 vs 1.586±0.022, t=16.424, P<0.05). After the transfection of miR-21 inhibitor, the inhibitor group had significantly lower expression of miR-21 than the control group (0.857±0.017 vs 1.684±0.039, t=33.669, P<0.05). Compared with the control group after the transfection of miR-21 inhibitor, the inhibitor group had a significant reduction in the proliferation ability of HepG2 cells (P<0.05), a significantly lower number of cells passing through the Transwell chamber (83.72±15.06 vs 147.85±20.64, t=4.347, P<0.05), and a significantly higher cell apoptosis rate (25.67%±3.95% vs 10.27%±2.14%, t=5.937, P<0.05). The inhibitor group had significantly lower relative expression of Wnt2 in HepG2 cells than the control group (0.862±0.127 vs 1.306±0.218, t=3.048, P<0.05). TargetScan software showed that miR-21 inhibitor significantly inhibited the luciferase activity of the cells transfected with wild-type Wnt2-3′UTR plasmid (0.972±0.102 vs 0.612±0.092, t=4.219, P<0.05), while there was no effect on the luciferase activity of the cells transfected with mutant Wnt2-3′UTR plasmid (0.982±0.093 vs 0.911±0.128, t=0.972, P>0.05). Conclusion Inhibition of miR-21 expression can effectively inhibit the proliferation and migration of HepG2 cells, promote the apoptosis of HepG2 cells, and inhibit the over-activation of the Wnt signaling pathway, and therefore, it may become one of the potential target genes for liver cancer treatment.
【关键字】:微RNAs; Wnt2蛋白质; 癌,肝细胞; 细胞增殖; 细胞运动
【Key words】:microRNAs; Wnt2 protein; carcinoma,hepatocellular; cell proliferation; cell movement
【引证本文】:WANG ZX, GUAN LJ, LI JM, et al. Effect of targeted regulation of the Wnt2 gene by microRNA-21 on the proliferation and migration of HepG2 hepatoma cells[J]. J Clin Hepatol, 2020, 36(4): 803-807. (in Chinese)
王泽鑫, 关利君, 李建明, 等. microRNA-21靶向调控Wnt2基因对肝癌细胞HepG2增殖与迁移的影响[J]. 临床肝胆病杂志, 2020, 36(4): 803-807.

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