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目的探讨mi R-203对胞质多聚腺苷酸化元件结合蛋白4(cytoplasmic polyadenylation element binding protein4,CPEB4)基因的靶向作用及对人脑胶质瘤U87细胞增殖的影响。方法采用免疫组化法检测CPEB4在人脑胶质瘤U87细胞中的表达,生物信息学方法预测mi R-203与CPEB4基因的靶向配对关系,并应用Dual-Luciferase誖报告系统鉴定;将mi R-203 mimics及si RNA转染U87细胞,Real-time PCR法检测mi R-203和CPEB4 m RNA水平,Western blot法检测CPEB4蛋白的表达,平板克隆形成试验检测癌细胞的增殖情况。结果光镜下可见U87细胞均匀分布,但细胞形态多样性明显,胞质内有CPEB4蛋白高表达,呈棕褐色;mi R-203与CPEB4基因靶向配对良好,mi R-203能够抑制CPEB4 m RNA水平。过表达mi R-203能够降低CPEB4 m RNA和蛋白的表达,抑制U87细胞的增殖。结论mi R-203通过负性调控人脑胶质瘤U87细胞中CPEB4基因的表达,进而抑制癌细胞的增殖。
Objective To investigate the effect of mi R-203 on the cytoplasmic polyadenylation element binding protein 4 (CPEB4) gene and its effect on the proliferation of human glioma U87 cells. Methods The expression of CPEB4 in human glioma U87 cells was detected by immunohistochemistry. The bioinformatics method was used to predict the target-pairing relationship between mi R-203 and CPEB4 genes and identified by Dual-Luciferase reporter system. R-203 mimics and si RNA were transfected into U87 cells. The mi R-203 and CPEB4 m RNA levels were detected by Real-time PCR, the expression of CPEB4 protein by Western blot, and the proliferation of cancer cells by plate clone formation assay. Results The U87 cells were evenly distributed under light microscope, but the morphological diversity of the cells was obvious. The expression of CPEB4 protein in the cytoplasm was high and tan. The targeting of mi R-203 and CPEB4 gene was good. Mi R-203 could inhibit the expression of CPEB4 m RNA level. Overexpression of mi R-203 reduced the expression of CPEB4 mRNA and protein and inhibited the proliferation of U87 cells. Conclusion mi R-203 negatively regulates the expression of CPEB4 gene in human glioma U87 cells and further inhibits the proliferation of cancer cells.