微小RNA-148a对胃癌细胞MKN45增殖能力的影响及机制研究

Impact of miR-148a on the proliferation of gastric carcinoma MKN45 cells and its mechanism

目的：探讨微小RNA（miR）-148a对人胃癌细胞增殖的影响及可能机制。方法采用实时定量PCR技术检测60例胃癌组织及癌旁组织中miR-148a的表达水平。利用慢病毒包装建立稳定表达miR-148a的MKN45细胞系为转染组，未转染的MKN45细胞系为对照组。 CCK8法检测MKN45细胞的增殖情况，通过targetscan预测miR-148a的靶基因，Western-blot检测靶基因的表达水平。结果54例（90．0％，54/60）胃癌组织中miR-148a的表达量较癌旁组织下调，其中37例（61．7％，37/60）下降2倍以上；淋巴结转移、有无癌旁血管浸润及TNM分期Ⅲ～Ⅴ期者其表达量明显降低（P＜0．05）。转染miR-148a后3 d，MKN45细胞生长明显受抑，转染组吸光度值（0．978±0．091）明显低于对照组（1．182±0．074，P＝0．000）。转染组CDC25B（通过targetscan发现的miR-148a靶基因）蛋白表达较对照组明显降低。结论 miR-148a在胃癌组织中的表达明显低于癌旁组织；其具有抑制胃癌细胞增殖的作用，CDC25B可能是miR-148a发挥抑癌作用的靶基因。

Objective To investigate the influence of miR-148a on cell proliferation of human gastric cancer cell lines MKN45. Methods Expression level of miR-148a was detected by qRT-PCR in the carcinoma tissues and the tissues adjacent to carcinoma of 60 patients with gastric cancer. Lentivirus packaging was used to establish MKN45 gastric cancer cell line expressing stable miR-148a as transfection group,and the untransfected MKN45 cell line as the control group. Gastric cancer MKN45 cell proliferation was detected by CCK8 method . The target gene of miR-148a was predicted by targetscan. Expression of target gene was examined by Western blotting. Result Expression of miR-148a in gastric cancer tissues of 54 cases （90.0%, 54/60） decreased, and among them, 37 tissue samples （61.7%, 37/60） decreased by 2 times. mir-148a expression in cancer tissues of patients with lymph node metastasis , vascular invasion and TNM stage Ⅲ-Ⅳ was down-regulated more obviously（P〈0.05）. From the third day after transfection, the growth of MKN45 cells was significantly inhibited （P〈0.01）. Gene CDC25B might be the target gene of miR-148a according to the results of targetscan. Western blot showed that CDC25B expression in transfection group was lower as compared to control group. Conclusion MiR-148a expression is down-regulated in gastric cancer tissues and inhibits gastric cancer cell proliferation. CDC25B may be the target gene of miR-148a that plays a role in tumor suppressor.