miRNA-221和miRNA-222对前列腺癌细胞PC-3的增殖、迁移及SIRT1表达的调控

doi:10.3969/j.issn.1673-8640.2014.05.005

摘要/Abstract

摘要：

目的观察前列腺癌PC-3细胞中微小RNA(microRNA, miRNA)-221和miRNA-222对细胞增殖和迁移的作用, 以及对沉默信息调节因子1(SIRT1)表达的影响。方法设立无处理细胞组(简称control组)、转染空载质粒pEX-5组(简称pEX-5组)、转染miRNA-221 反义抑制质粒组(简称miRNA-221抑制组)、转染miRNA-222 反义抑制质粒组(简称miRNA-222抑制组), 应用细胞计数试剂盒(CCK-8法)检测miRNA-221和miRNA-222对前列腺癌细胞增殖的影响;采用细胞划痕修复实验检测miRNA-221和miRNA-222对细胞迁移能力的影响;采用免疫印迹法和荧光定量聚合酶链反应(PCR)检测SIRT1在蛋白水平和mRNA水平的变化;采用miRNAanda靶标进行预测分析。结果在成功抑制了PC-3细胞中miRNA-221或miRNA-222的表达后, 连续7 d检测细胞活性, miRNA-221抑制组和miRNA-222 抑制组的细胞活性(A_{450 nm}值)较pEX-5组降低1.5~2.0倍, 自第3天起两组之间即有明显差异(t=6.7, P<0.01; t=5.3, P<0.01); 划痕实验显示, miRNA-221抑制组和miRNA-222抑制组的迁移能力明显低于pEX-5组。miRNA-221抑制组、miRNA-222抑制组中SIRT1蛋白的相对表达量分别为(0.26±0.021)、(0.21±0.005 8), 与pEX-5组(0.14±0.017)比较差异均有统计学意义(t值分别为6.3、6.4, P均<0.05);而SIRT1 mRNA的表达水平3组之间差异无统计学意义(P>0.05)。结论 miRNA-221和miRNA-222能促进前列腺癌PC-3细胞的增殖与迁移能力, 影响SIRT1蛋白的表达。

关键词: 微小RNA-221, 微小RNA-222, 沉默信息调节因子, 前列腺癌

Abstract:

Objective To investigate the roles of micro RNA(miRNA)-221 and miRNA-222 on the proliferation and migration of prostate cancer cell PC-3 and the influence on silent information regulator 1(SIRT1). Methods The groups were arranged as cells without treatment (control group), cells transfected with pEX-5 empty plasmid (pEX-5 group), cells transfected with miRNA-221 inhibitor (miRNA-221 inhibition group) and cells transfected with miRNA-222 inhibitor (miRNA-222 inhibition group). The influence of miRNA-221 and miRNA-222 on prostate cancer cell proliferation was determined by CCK-8, and the migration ability was determined by wound healing test. The protein and mRNA levels of SIRT1 were determined by Western blotting and fluorescence quantitation polymerase chain reaction (PCR), respectively. The prediction analysis was performed by miRNAanda target. Results After the expression inhibition of miRNA-221 or miRNA-222 in cell PC-3, the cell activities were determined for 7 d. The activities (A_{450 mm}) in miRNA-221 and miRNA-222 inhibition groups were down-regulated about 1.5-2.0 times compared with that in pEX-5 group, and there were significant differences between the 2 groups from the 3rd d(t=6.7, P<0.01; t=5.3, P<0.01). The wound healing test showed that the migration abilities of miRNA-221 inhibition group and miRNA-222 inhibition group decreased than that of pEX-5 group. Moreover, the expression levels of SIRT1 protein in miRNA-221 inhibition group(0.26±0.021) and miRNA-222 inhibition group(0.21±0.005 8) were up-regulated compared with that in pEX-5 group(0.14±0.017), and there were statistical significances (t=6.3 and 6.4, P<0.05 ). The SIRT1 mRNA expression levels had no statistical significance among the 3 groups(P>0.05). Conclusions The miRNA-221 and miRNA-222 promote the proliferation and migration of prostate cancer cell PC-3 and have the influence on SIRT1 expression in protein level.

Key words: Micro RNA-221, Micro RNA-222, Silent information regulator 1, Prostate cancer

中图分类号:

杨晓, 甘蓉, 杨英梅, 赵苓旭, 薄金双, 官雁鸣, 孟庆贺, 吕建新. miRNA-221和miRNA-222对前列腺癌细胞PC-3的增殖、迁移及SIRT1表达的调控[J]. 检验医学, 2014, 29(5): 446-451.

YANG Xiao, GAN Rong, YANG Yingmei, ZHAO Lingxu, BO Jinshuang, GUAN Yanming, MENG Qinghe, L Jianxin.. Modulation of the proliferation, migration and SIRT1 expression of prostate cancer cell PC-3 by miRNA-221 and miRNA-222[J]. , 2014, 29(5): 446-451.

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