MiR-125a-5P 通过靶向生长因子受体结合蛋白-10调控髓母细胞瘤表皮生长因子受体信号通路  

miR -125a -5P regulated epidermal growth factor receptor signal pathway by targeting growth factor recep-tor-bound protein 10 in medulloblastoma

作　　者：楚冬梅[1] 刘晓智[2] 高心静 刘翠苹[1] 张岩[1] 李艳霞[2] 姜忠敏[3] 姚玲[1] 

机构地区：[1]天津市第五中心医院儿科,300450 [2]天津市第五中心医院中心实验室,300450 [3]天津市第五中心医院病理科,300450

出　　处：《中华实用儿科临床杂志》2015年第20期1585-1588,共4页Chinese Journal of Applied Clinical Pediatrics

基　　金：天津市滨海新区医药卫生科研基金（2012BWKZ003）

摘　　要：目的：探索 miR-125a-5P 在髓母细胞瘤表皮生长因子受体（EGFR）信号通路中的调控机制。方法利用 TargetScan 和 Sanger 软件在 EGFR 信号通路中预测 miR-125a-5P 的潜在靶点，脂质体介导细胞基因转染，共分3组：对照组、无义组和 miR-125a-5P 组。荧光素酶实验验证 miR-125a-5P 与细胞周期蛋白依赖性激酶抑制剂-2B（CDKN2B）、E2F 转录因子3（E2F3）、丝裂原活化蛋白激酶-14（MAPK14）和生长因子受体结合蛋白-10（GRB10）间的靶矢关系；转染髓母细胞瘤 D341细胞 miR-125a-5p 寡核苷酸，反转录聚合酶链反应检测D341细胞中 miR-125a-5p 的表述水平，噻唑蓝法绘制细胞生长曲线，Transwell 实验检测瘤细胞迁移能力， Western blot 法检测 GRB10、EGFR、磷脂酰肌醇-3-羟基激酶（PI3K）和 Ras 的蛋白表达水平。结果荧光素酶实验结果显示，GRB10是所检测基因中唯一的 miR-125a-5P 靶基因；获得 miR-125a-5P 转染后 D341细胞增殖速度减慢。miR-125a-5P 组的细胞迁移率与对照组[（38.16±7.47）％]和无义组[（36.79±8.94）％]比较， miR-125a-5P 组最低[（13.59±4.41）％]，3组间比较有统计学差异（χ2＝11.495，P 约0.05）。miR-125a-5P 转染可致 EGFR 蛋白表达水平升高1.67倍，GRB10、PI3K 和 Ras 的水平分别降低至23％、61％和42％。结论miR-125a-5P 通过靶向沉默GRB10的表达抑制髓母细胞瘤EGFR 下游信号通路，发挥肿瘤生长抑制作用。Objective To explore the regulation mechanism for miR - 125a - 5P in epidermal growth factor receptor（EGFR）signaling pathway in medulloblastoma. Methods The potential targets of miR - 125a - 5P in the EGFR signaling pathway were predicted by TargetScan and Sanger software,there were 3 groups：control group,non -sense group and miR - 125a - 5P group. Their relationship,between miR - 125a - 5P and cyclin - dependent kinase in-hibitor 2B（ CDKN2B）,E2F transcription factor 3（ E2F3）,mitogen - activated protein kinase 14（ MAPK14）and growth factor receptor - bound protein 10（GRB10）,were tested by luciferase experiments. After miR - 125a - 5P oligo-nucleotide was transfected to D341 cells,miR - 125a - 5P level was detected by reverse transcription polymerase chain reaction. Then the thiazolyl blue tetrazolium bromide assay was used to draw the cell growth curves,and Transwell assay was used to detect cell migration ability. The expression levels of GRB10,EGFR,phosphatidylinositol 3 - kinase（PI3K） and Ras were tested by Western blot method. Results The results of luciferase experimental results showed that GRB10 was the only target gene of miR - 125a - 5P. After miR - 125a - 5P being transfected,the D341 cell prolifera-tion obviously declined markedly. Compared with control group[（38. 16 ± 7. 47）% ]and the non - sense group [（36. 79 ± 8. 94）% ],cell migration rate in the miR - 125a - 5P group was lowest[（13. 59 ± 4. 41）% ],and there was a significant difference among 3 groups（χ2 = 11. 495,P ＆lt; 0. 05）;in the miR - 125a - 5P group,the expression level of EGFR increased 1. 67 times,GRB10,PI3K and Ras levels were reduced to 23% ,61% and 42% . Conclusion miR - 125a - 5P can inhibit tumor growth by silenced GRB10 expression targeting EGFR downstream signaling pathways in medulloblastoma.

关 键 词：髓母细胞瘤 增殖 转移 微小RNA 

分 类 号：R739.4[医药卫生—肿瘤]