miR-124通过靶向调控DNMT3B抑制胃癌细胞增殖

MiR-124 Suppresses Cell Proliferation by Targeting DNMT3B in Gastric Carcinoma

目的明确miR-124是否通过靶向调控DNA甲基化转移酶3B(DNA methyltransferase 3B,DNMT3B)表达而抑制胃癌细胞增殖能力,从而揭示miR-124的抑瘤分子机制。方法采用MTT检测miR-124对人胃癌MKN-45细胞的增殖能力;构建DNMT3B 3'UTR-荧光素酶报告载体,通过荧光素酶报告检测观察miR-124对DNMT3B3'UTR-荧光素酶活性的影响;将miR-124 mimics转染胃癌细胞MKN-45,采用Western blot检测DNMT3B表达水平。结果 MTT结果显示,在转染miR-124 mimics 24、48和72 h后OD值(0.264±0.023、0.377±0.041、0.524±0.029)分别与对照组(0.414±0.051、0.619±0.065、0.898±0.072)比较,差异均有显著性(均P<0.05),且具有时间依赖性;荧光素报告载体系统证实DNMT3B是miR-124直接调控的靶基因。Western blot结果显示,miR-124可抑制DNMT3B蛋白的表达。结论 miR-124通过靶向调控DNMT3B的表达而抑制胃癌细胞增殖能力。

Objective To explicit whether miR-124 suppresses cell proliferation by targeting DNMT3B, thus to re- veal molecular mechanism of miR-124 function as a tumorsuppressor in gastric carcinoma. Methods DNMT3B 3＇UTR- lueiferase vector was constructed and luciferase reporter gene assay was employed to examine the effect of miR-124 on lucif- erase activity. MKN-45 cells were transfeeted with miR-124 mimics, and next Western blotting was performed to detect the expressions of DNMT3B protein. Results After transfection of miR-124 mimics for 24 h, 48 h ,72 h respectively, MTT assay showed that the OD values were （0.264 ±0. 023）, （0. 377 ±0.041 ） and （0. 524 ±0. 029） ,compared with the con- trol group （0. 414 ±0. 051）, （0. 619 ±0. 065） and （0. 898 ±0. 072） ,the groups were significantly different （ all P 〈 0. 05 ） ,and it is on a time-dependent manner. Luciferase reporter vector system confirmed that DNMT3B was a target gene of miR-124. Western blot showed that the expressions of DNMT3B protein were inhibited by miR-124. Conclusion miR- 124 suppresses cell proliferation by targeting DNMT3B in gastric carcinoma.