贵州医科大学学报

2018, v.43;No.219(12) 1412-1419

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miR-543对胶质瘤细胞SHG-44增殖和凋亡的影响
The Effect of miR-543 on Proliferation and Apoptosis in Glioma SHG-44 Cells

赵云东;宋玉琴;刘娜;孙冬弢;
ZHAO Yundong;SONG Yuqin;LIU Na;SUN Dongtao;The Second Affiliated Hospital of Shenyang Medical College;The Sixth People's Hospital of Shenyang;The Seventh People's Hospital of Shenyang;

摘要(Abstract):

目的:观察miR-543对人胶质瘤细胞SHG-44增殖和凋亡的作用。方法:设计并合成miR-543寡核苷酸模拟物(miR-543 mimic)及阴性对照mimic NC序列、寡核苷酸抑制物(miR-543 inhibitor)及阴性对照inhibitor NC序列,将人胶质瘤细胞SHG-44株分为Control(对照组)、mimic NC组、inhibitor NC组、miR-543 mimic组和miR-543 inhibitor组,各组细胞转染相应序列; Real-time PCR检测miR-543及其靶基因氨酰基-tRNA合成酶相互作用多功能蛋白3(AIMP3) mRNA的表达,应用噻唑蓝细胞增殖实验(MTT)评估SHG-44细胞的增殖,应用流式细胞术检测细胞周期和细胞凋亡,应用Western Blot检测细胞周期蛋白cyclin D1、cyclin E与细胞凋亡相关蛋白Bcl-2、bax的表达变化。结果:与对照组相比,转染miR-543 mimic后,SHG-44细胞中miR-543 RNA表达水平显著上调(P <0. 01),其靶基因AIMP3 mRNA表达水平显著下调(P <0. 01);转染miR-543 inhibitor后,细胞中miR-543 RNA表达水平显著下调(P <0. 01),AIMP3 mRNA表达水平显著上调(P <0. 01);转染miR-543 mimic后,细胞增殖水平显著降低(P <0. 01);转染miR-543 inhibitor后,细胞增殖水平显著升高(P <0. 01);转染miR-543 mimic后,细胞处于G1期的细胞数量明显增加(P <0. 01),同时细胞凋亡率也显著增加(P <0. 01),而转染miR-543 inhibitor后细胞凋亡率显著减少(P <0. 01); Westrn Blot结果显示,与对照组相比,转染miR-543 mimic后,细胞周期蛋白cyclin D1、cyclin E及抗凋亡相关基因Bcl-2蛋白表达显著下调(P <0. 01),促凋亡相关基因bax蛋白表达显著上调(P <0. 01);转染miR-543 inhibitor后,细胞中cyclin D1、cyclin E、Bcl-2蛋白表达显著上调(P<0. 01),bax蛋白表达显著下调(P <0. 01)。结论:miR-543可能通过下调细胞周期蛋白cyclin D1、cyclin E及抗凋亡相关基因Bcl-2的表达,上调促凋亡基因bax的表达进而抑制细胞增殖,促进细胞凋亡。
Objective: To observe the role of miR-543 on the proliferation and apoptosis of human glioma SHG-44 cells and to investigate its effect on the protein expression of cyclin D1,cyclin E,Bcl-2 and bax. Methods: miR-543 oligonucleotide mimics( miR-543 mimic) and oligonucleotide inhibitor( miR-543 inhibitor) were used to transfect SHG-44 cells. Real-time PCR was used to detect the expression of miR-543 and AIMP3 RNA; methylthiazol tetrazoliu( MTT) assay was used to evaluat cell proliferation; Flow cytometry was applied to detect cell cycles and apoptosis; The expression of cyclin D1,cyclin E,Bcl-2 and bax were detected by Western Blot. Results: Compared with the control group,the expression of miR-543 in SHG-44 cells was significantly up-regulated( P < 0. 01) and the expression of AIMP3 mRNA was significantly down-regulated( P < 0. 01) after transfection of miR-543mimic; the expression of miR-543 in SHG-44 cells was significantly down-regulated( P < 0. 01) and that of AIMP3 mRNA was significantly up-regulated( P < 0. 01) after transfection of miR-543 inhibitor. The cell proliferation level significantly decreased after transfection of miR-543 mimic( P <0. 01),but after transfection of miR-543 inhibitor,the cell proliferation level significantly increased;The number of cells in G1 phase and also the apoptosis rate significantly increased( P < 0. 01) after transfection of miR-543 mimic( P < 0. 01); but the apoptosis rate significantly decreased( P < 0. 01)after transfection of miR-543 inhibitor. Compared with the control group,Western Blot results showed that the expression of cyclin D1,cyclin E and Bcl-2 was significantly down-regulated( P < 0. 01),meanwhile the expression of bax protein was significantly up-regulated( P < 0. 01) after transfection of miR-543 mimic,while the expression of cyclin D1,cyclin E and Bcl-2 was significantly up-regulated( P < 0. 01) and the expression of bax protein was significantly down-regulated( P < 0. 01) after transfection of miR-543 inhibitor,. Conclusion: miR-543 can inhibit cell proliferation and promote apoptosis by lowering the cyclin D1,cyclin E and Bcl-2 expression level meanwhile elevating bax expression level.

关键词(KeyWords): miR-543;胶质瘤细胞;细胞周期蛋白类;Bcl-2;bax;细胞增殖;细胞凋亡
miR-543;glioma cells;cyclins;Bcl-2;bax;cell proliferation;apoptosis

Abstract:

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基金项目(Foundation): 辽宁省科学技术计划项目(20170540873)

作者(Author): 赵云东;宋玉琴;刘娜;孙冬弢;
ZHAO Yundong;SONG Yuqin;LIU Na;SUN Dongtao;The Second Affiliated Hospital of Shenyang Medical College;The Sixth People's Hospital of Shenyang;The Seventh People's Hospital of Shenyang;

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DOI: 10.19367/j.cnki.1000-2707.2018.12.010

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