miR-29b的作用靶点及其对Huh7细胞增殖及凋亡的干预作用

Targets of miR-29b and Its Effect on Proliferation and Apoptosis of Huh7 Cells

目的探讨mi R-29b(micro RNA-29b)的作用靶点及其对肝肿瘤细胞系Huh7细胞增殖及凋亡的干预作用。方法荧光定量PCR检测正常肝细胞系LO2及肝癌细胞系Huh7的mi R-29b和Mcl-1表达水平。通过生物信息学分析预测Mcl-1基因是否受micro RNA调控。将mi R-29b模拟物通过Lipofectamine 2000顺时转染Huh7细胞,检测mi R-29b对Mcl-1表达水平的影响。MTT法检测mi R-29b模拟物转染对Huh7细胞增殖的影响。Annexin V/PI染色法检测mi R-29b对Huh7细胞凋亡的影响。结果肝肿瘤细胞相比于正常肝细胞高表达Mcl-1(3.42±0.15比1.00±0.04,P<0.05)低表达mi R-29b(0.43±0.04比1.00±0.06,P<0.05),在肝癌细胞中转染mi R-29b可使Mcl-1的表达水平下降(0.37±0.03比1.03±0.07,P<0.05),转染mi R-29b可抑制Huh7细胞的增殖(0.85±0.11比1.68±0.17,P<0.05)并诱导其发生凋亡[(28.4±2.10)%比(3.6±0.06)%,P<0.05]。结论 Mi R-29b下调Huh7细胞Mcl-1蛋白的表达并诱导其发生凋亡。

Objective To investigate the therapeutic targets of miR-29b and its effect on proliferation and apoptosis of hepatocarcinoma cell line Huh7. Methods The expression of Mcl-1 and miR-29b in normal liver cell line LO2 and hepatocellular carcinoma cell line Huh7 was detected by qPCR. The putative binding sites of Mcl-1 gene was analyzed by using bioinformatic method. MiR-29b was transfected into Huh7 cells by Lipofectamine 2000, then the expression of Mcl-1 was detected. The cell growth was assessed by MTT after Huh7 cells transfected with miR-29b mimics. The apoptosis of cells was measured by Annexin V/PI staining. Results Compared with normal liver cells, Huh7 cells had higher expression of Mcl-1 （3.42±0.15 vs 1.00±0.04, P〈0.05） and lower expression of miR-29b（0.43±0.04 vs 1.00±0.06, P〈0.05）. Transfection of miR-29b mimics suppressed Mcl-1 expression in Huh7 cells （0.37±0.03 vs 1.03±0.07, P〈0.05） and inhibited the proliferation of Huh7 cells （0.85±0.11 vs 1.68±0.17, P〈0.05） while inducing apoptosis in Huh7 cells [（28.4±2.1）% vs （3.6±0.06）%, P〈0.05]. Conclusion MiR-29b down-regulates the expression of Mcl-1 protein inducing apoaptosis in Huh7 cells.