MicroRNA-483-3p对人神经胶质瘤细胞的作用

发布时间：2018-03-20 05:04

本文选题：MicroRNA--p　切入点：神经胶质瘤　出处：《中国病理生理杂志》2017年07期 　论文类型：期刊论文

【摘要】：目的:探讨microRNA(miRNA)-483-3p对人神经胶质瘤细胞A172生长和迁移能力的影响及潜在作用机制。方法:实时荧光定量聚合酶链式反应(RT-q PCR)检测人肾胚细胞系HEK-293和不同神经胶质瘤细胞株(A172、U251和SHG44)中miRNA-483-3p的表达水平。转染miRNA-483-3p抑制序列(miRNA-483-3p inhibitor)下调A172细胞中miRNA-483-3p的表达,采用CCK-8法和流式细胞术检测细胞活力和周期分布;Transwell实验检测细胞的迁移;Western blot检测周期相关调控因子及上皮-间充质转化相关蛋白的水平。双萤光素酶报告基因分析法预测及验证其可能的靶基因。结果:miRNA-483-3p在各型神经胶质瘤细胞中高表达。沉默miRNA-483-3p后,A172细胞的活力下降并呈现出明显的周期阻滞,且细胞迁移率也显著降低。同时细胞中cyclin D1、周期蛋白依赖性激酶4、磷酸化视网膜母细胞瘤蛋白、N-cadherin及vimentin的蛋白表达水平均显著降低,E-cadherin和β-catenin的蛋白表达水平显著升高。双萤光素酶报告基因分析显示Smad4是miRNA-483-3p的可能作用靶点,A172细胞共转染miRNA-483-3p inhibitor和Smad4 siRNA可部分逆转miRNA-483-3p介导的细胞增殖及迁移抑制。结论:沉默miRNA-483-3p可通过靶向Smad4抑制神经胶质瘤细胞株A172的生长及迁移。
[Abstract]:Objective: to investigate the effect of microRNA(miRNA)-483-3p on the growth and migration of human glioma cell line A172 and its potential mechanism. Methods: Real-time fluorescent quantitative polymerase chain reaction (RT-q PCR) was used to detect human renal embryo cell line HEK-293 and glioma fine cells. The expression level of miRNA-483-3p in A172 U251 and SHG44) was down-regulated by transfection of miRNA-483-3p inhibitory sequence (miRNA-483-3p inhibitor), and the expression of miRNA-483-3p in A172 cells was down-regulated by transfection of miRNA-483-3p inhibitor. CCK-8 assay and flow cytometry were used to detect cell viability and cell cycle distribution. Western blot was used to detect the levels of cycle related regulatory factors and epithelial-mesenchymal transformation related proteins. Results: miRNA-483-3p was highly expressed in glioma cells. After silencing miRNA-483-3p, the activity of A172 cells decreased and the cell cycle arrest was observed. At the same time, the expression of cyclin D1, cyclin dependent kinase 4, phosphorylated retinoblastoma protein N-cadherin and vimentin decreased significantly, and the expression of E-cadherin and 尾 -catenin increased significantly. Double luciferase reporter gene analysis showed that Smad4 is a possible target of miRNA-483-3p in miRNA-483-3p inhibitor and Smad4 siRNA cotransfection with miRNA-483-3p inhibitor and Smad4 siRNA can partially reverse miRNA-483-3p mediated cell proliferation and migration inhibition. Conclusion: silencing miRNA-483-3p can inhibit nerve by targeting Smad4. Growth and migration of glioma cell line A172.
【作者单位】：郑州大学附属肿瘤医院神经外科;
【基金】：国家自然科学基金资助项目(No.81372269)
【分类号】：R739.41

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