miR-19a通过Fas基因调节胃癌细胞增殖水平的实验研究

Experimental study on miR-19a regulating gastric cancer cells proliferation via Fas gene

目的研究miR⁃19a通过Fas基因调节胃癌细胞增殖水平的作用。方法培养胃癌SGC⁃7901细胞株后转染阴性对照(NC)模拟物、miR⁃19a模拟物、Fas的siRNA、NC的siRNA、过表达Fas的pcDNA3.1质粒、空白的pcDNA3.1质粒,检测细胞增殖活力、细胞凋亡率及细胞中miR⁃19a、Fas的表达量,采用双荧光素酶报告基因验证miR⁃19a与Fas基因mRNA 3’UTR的靶向结合;皮下注射胃癌SGC⁃7901细胞株后建立胃癌荷瘤小鼠模型,瘤周注射NC模拟物或miR⁃19a模拟物,测定移植瘤质量及移植瘤中Fas的表达量。结果miR⁃19a模拟物组的细胞增殖活力及细胞中miR⁃19a的表达量明显均高于NC组、NC模拟物组,细胞凋亡率、细胞中Fas的表达量、双荧光素酶报告基因的荧光值均明显低于NC组、NC模拟物组(P<0.05);siRNA⁃Fas组的细胞增殖活力明显高于siRNA⁃NC组,细胞凋亡率及细胞中Fas的表达量均能明显低于siR⁃NA⁃NC组(P<0.05);转染过表达Fas的pcDNA3.1质粒能够削弱miR⁃19a模拟物促进细胞增殖、抑制细胞凋亡的作用;在移植瘤小鼠中,miR⁃19a模拟物组的移植瘤质量明显高于NC组、NC模拟物组,移植瘤中Fas的表达量明显低于NC组、NC模拟物组(P<0.05)。结论miR⁃19a能够促进胃癌细胞增殖且靶向抑制Fas基因是介导该作用的分子机制之一。

Objective The aim of this study is to study the regulatory effect of miR⁃19a on the proliferation of gastric cancer cells through Fas gene.Methods Gastric cancer SGC⁃7901 cells were cultured and transfected with negative control(NC)mimics,miR⁃19a mimics,Fas siRNA,NC siRNA,over⁃expressed Fas pcDNA3.1 plasmids and blank pcDNA3.1 plasmids.Cell proliferation activity,apoptotic rate and the expression of miR⁃19a and Fas were detected.Double luciferase reporter gene was used to validate the binding between miR⁃19a and Fas gene 3'UTR.After establishing mouse model of gastric cancer by subcutaneous injection of SGC⁃7901 cell line,NC or miR⁃19a mimics was peritumorally injected.The quality of transplanted tumors and the expression of Fas in trans⁃planted tumors were determined.Results The cell proliferation viability and the expression of miR⁃19a in cells of miR⁃19a mimic group were significantly higher than those in NC group and NC mimic group.The apoptosis rate,expression of Fas and fluorescence value of double luciferase reporter gene in cells were significantly lower than NC group and NC mimic group(P<0.05).Moreover,the proliferation viability of siRNA⁃Fas group was significantly higher than NC group.The apoptosis rate and the expression of Fas were remarkably lower in siRNA⁃Fas group than siRNA⁃NC group(P<0.05).Transfection of Fas⁃expressing pcDNA3.1 plasmid could weak the effects of miR⁃19a mimic in promoting cell proliferation and inhibiting cell apoptosis.In addition,the quality of transplanted tumor of miR⁃19a mimic group was significantly higher than that in NC group and NC mimic group in transplanted tumor mice.Expression of Fas in transplanted tumors was significantly lower than that in NC group and NC mimic group(P<0.05).Conclusion miR⁃19a could promote the proliferation of gastric cancer cells by targeting Fas gene.