摘要
目的探讨微小RNA-101(miR-101)对胃癌化疗耐药的影响及其机制。方法选择人正常胃黏膜上皮细胞GES1(以下称GES1细胞)、胃癌细胞SGC-7901(以下称SGC-7901细胞)、顺铂耐药胃癌细胞SGC-7901(以下称SGC-7901/DDP细胞),采用实时荧光定量PCR法检测3种细胞miR-101、c-met、MDR1 mRNA表达,采用Western blotting法检测3种细胞c-met、P-gp蛋白表达。选择SGC-7901/DDP细胞,随机分为miR-101 mimic组及阴性对照组,分别给予miR-101 mimic(miR-101模拟物)和mimic control(阴性对照)干预,采用实时荧光定量PCR法、Western blotting法检测MDR1 mRNA和P-gp蛋白表达。采用双荧光素酶报告基因实验检测miR-101 mimic组和阴性对照组下游靶基因c-met荧光素酶活性。结果与GES1细胞比较,SGC-7901细胞、SGC-7901/DDP细胞miR-101mRNA相对表达量明显降低,c-met mRNA和蛋白相对表达量明显升高,且以SGC-7901/DDP细胞变化更明显(P均﹤0.05)。与GES1细胞比较,SGC-7901细胞、SGC-7901/DDP细胞MDR1 mRNA、P-gp蛋白相对表达量明显升高,且以SGC-7901/DDP细胞变化更明显(P均﹤0.05)。miR-101 mimic组MDR1 mRNA、P-gp蛋白相对表达量明显低于阴性对照组(P均﹤0.05)。双荧光素酶报告基因实验结果显示,miR-101能与c-met 3'-UTR结合,并且miR-101mimic组荧光素酶活性明显低于阴性对照组(P﹤0.05)。结论胃癌细胞miR-101低表达;miR-101可能通过抑制c-met表达逆转胃癌细胞的化疗耐药。
Objective To investigated the effect of microRNA-101（ miR-101） on the chemoresistance of gastric cancer（ GC） cells and its mechanism. Methods Human normal gastric mucosal epithelial cells GES1,gastric cancer cells SGC-7901,and cisplatin-resistant gastric cancer cells SGC-7901/DDP were collected in this study. The real-time fluorescent quantitative PCR was used to detect the expression of miR-101,c-met and MDR1 mRNA,and the expression of c-met and P-gp protein was detected by Western blotting. The SGC-7901/DDP cells were randomly assigned to the miR-101 mimic group and negative control group,which were treated with miR-101 mimic and mimic control,respectively. The expression of MDR1 mRNA and P-gp protein was measured by qRT-PCR and Western blotting. Meanwhile,Luciferase reporter gene assay were applied to detect the downstream target gene c-met expression in the miR-101 mimic and negative group.Results As compared to the GES1 cells,the expression of miR-101 significantly decreased and the mRNA and protein expression of c-met increased in the SGC-7901 cells and SGC-7901/DDP cells,in addition,the changes in the SGC-7901/DDP cells were more significantly（ all P〈0. 05）. The expression of MDR1 mRNA and P-gp protein significantly increased in the SGC-7901 cells and SGC-7901/DDP cells as compared with that of the GES1 cells,and SGC-7901/DDP cell displayed higher expression of MDR1 mRNA and P-gp protein（ all P〈0. 05）. The relative expression of MDR1 mRNA and P-gp protein in the miR-101 mimic group was significantly lower than that in the negative control group（ P〈0. 05）. Luciferase reporter assay demonstrated that miR-101 bound to the 3＇-UTR of c-met,and luciferase activity was significantly lower in the miR-101 mimic group as compared with that of the control group（ P〈0. 05）. Conclusion The miR-101 is low expressed in the GC cells and miR-101 may reverse chemoresistance of gastric cancer cells by inhibiting c-met expression.
作者
王新平
王彩云
张建武
WANG Xinping;WANG Cai;ZHANG Jianwu(Honghu Second People＇ s Hospital,Honghu 433202,China)
出处
《山东医药》
CAS
2018年第28期44-48,共5页
Shandong Medical Journal
