摘要
目的探讨miR-214对Hep G2细胞增殖及细胞周期的影响。方法 Realtime-PCR法检测肝癌细胞株SMMC-7721、Hep G2、SK-Hep-1、Huh 7、Hep3B及正常肝细胞L-02中miR-214表达量,并利用脂质体转染miR-214NC及miR-214 mimics,采用Realtime-PCR法检测转染效果;采用MTT法、流式细胞术分别检测miR-214对肝癌细胞活力、凋亡及细胞周期影响;蛋白印迹(WB)检测miR-214对肝癌细胞中细胞周期蛋白D1(cyclin D1),细胞周期蛋白依赖性激酶4(CDK4),生存素(survivin)及增值细胞核抗原(PCNA)表达影响。结果 miR-214在肝癌细胞SMMC-7721、SK-Hep-1、Huh 7、Hep3B、Hep G2中表达量[分别为(1.25±0.10)、(1.43±0.10)、(0.95±0.09)、(0.98±0.01)、(0.82±0.08)]明显低于正常肝细胞L-02(2.52±0.23),其中Hep G2中miR-214表达量最低。miR-214 mimics组Hep G2细胞中miR-214表达量(2.38±0.23)明显高于miR-214 NC组(0.83±0.08),miR-214mimics组Hep G2细胞活力(0.37±0.03)明显低于miR-214 NC组(0.78±0.07);与miR-214 NC组比较,miR-214mimics组Hep G2细胞凋亡率明显升高,细胞周期阻滞于G1期,cyclin D1、CDK4、survivin及PCNA表达量下调,差异均有统计学意义(P<0.01)。结论上调miR-214表达可抑制肝癌细胞增殖,其机制可能与调控细胞周期相关蛋白表达水平有关。
Objective To explore inhibitive effect of miR-214 on proliferation and cell cycle in hepatocellular carci- noma cells. Methods The expression of rniR-214 in hepatoceUular carcinoma cells of SMMC-7721 ,HepG2,SK-Hep-1, Huh 7, and Hep3B and normal liver cell L-02 was detected with realtime-PCR. Hepatocellular carcinoma cells were trans- fected with miR-214 NC and miR-214 mimics by liposome and the outcome of the transfection was determined with real-tirne-PCR. The cell viability, cell apoptosis and cell cycle was assessed with （3-（ 4,5-dimethyl-2-thiazolyl）-2,5-diphen- yl-2-H-tetrazolium bromide （MTT） assay and flow cytometry, respectively. The expression of cyclin D1, cyclin-depend- ent protein kinases （CDK4） ,survivin,and proliferating cell nuclear antigen （PCNA） were detected with Western blot. Results The expression of miR-214 in the cells of SMMC-7721 （ 1.25±0. 10 ）, SK-Hep-1 （1. 43 ±0. 10 ）, Huh 7 （0. 95±0. 09） ,Hep3B （0. 98 ±0. 0） ,and HepG2 （0. 82 ±0. 08） were all lower than that in L-02 cells （2. 52±0. 23） and the expression of miR-214 was the lowest in HepG2 cells. The expression of miR-214 in rniR-214 mimics HepG2 cells （2. 38±0. 23 ） was higher than that in miR-214 NC HepG2 cells （ 0. 83±0. 08 ）. The cell viability of rniR-214 mimics HepG2 cells （0. 37 ±0. 03） was lower than that of rniR-214 NC cells （0. 78±0. 07）. Compared with those of the rniR-214 NC HepG2 cells ,the cell apoptotic rate was increased;cell cycle was arrested at G1 ;and expressions of cyclin D1, CDK4, survivin, and PCNA were down-regulated in miR-214 mimics HepG2 cells, with significant differences be- tween the two groups （ P 〈0. 01 for all）. Conclusion miR-214 mimics inhibits HepG2 cell proliferation;the mechanism of the inhibitive effect may be related to the expression of cell cycle regulation related protein.
出处
《中国公共卫生》
CAS
CSCD
北大核心
2017年第4期614-617,共4页
Chinese Journal of Public Health
