microRNA-451对人脑胶质瘤细胞株A172增殖和凋亡的影响

Suppressive effect of microRNA-451 on proliferation and apoptosis of glioma cell line A172 in vitro

目的 观察microRNA-451（miR-451）对胶质瘤细胞株A172增殖和凋亡的影响.方法 合成寡核苷酸miR-451拟似物（miR-451 mimics）转染A172细胞,实时聚合酶链反应（Real-time PCR）检测转染后miR-451的表达,Western blot法检测蛋白表达,应用流式细胞术、噻唑蓝（MTT）比色法和Annexin Ⅴ法评价miR-451对A172细胞生长、增殖和凋亡的影响.结果 与对照组比较,miR-451 mimics转染组细胞miR-451表达升高＞6倍 Western blot法显示癌基因AKT1表达下降到（43.81±5.20）%、Cyclin D1（56.09±3.40）%和bcl-2（63.49±3.70）%,抑癌基因p27表达升高到（145.51±6.70）% 细胞周期G0/G1期细胞增多达17.4%,出现G0/G1期阻滞 MTT法分析显示细胞生长受抑＞35% Annexin Ⅴ法检测显示细胞凋亡明显升高＞15%.结论 miR-451可抑制人脑胶质瘤细胞生长和增殖,诱导细胞凋亡,具有抑瘤作用.

Objective To investigate the effect of microRNA-451 （miR-451） on proliferation and apoptosis of glioma cell line A172. Methods The oligonucleotide was transfected to human glioma cell line A172. The expression of miRNA-451 was identified by Real-time polymerase chain reaction （PCR） after transfection, and targeting protein expression was evaluated by Western blotting. The cell proliferation was determined by flow cytometry and methylthiazol tetrazolium （MTT） assay. Moreover, cell proliferation and apoptosis molecules were detected by Western blotting. The number of apoptotic cells was determined by Annexin Ⅴ assay. Results After transfection of miRNA-451 mimics, as compared with control group,the miR-451 was increased larger than 6-fold in miRNA-451 mimics group, meanwhile the protein expression of AKT1 was down-regulated to （43.81 ±5.20）%, Cyclin D1 （56.09 ±3.40）% and bcl-2 （63.49± 3.70） %, conversely, p27 was up-regulated to （ 145.51 ±6.70） %. And then the cell cycle was significantly arrested in G0/G1 phase and G0/G1 phase cells were increased larger than 17.4% by flow cytometry assay. Cell proliferation was decreased larger than 35% by the MTT assay and cell apoptosis was significantly increased larger than 15% by Annexin V assay. Conclusion miR-451 could inhibit proliferation and induce apoptosis acivity of A172 glioma cells, suggesting that miR-451 was a tumor suppressor gene in glioma.