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环状RNA同源结构域相互作用蛋白激酶3(circHIPK3)通过海绵化miR-124-3p促进胶质瘤细胞的生长和转移

[Circular RNA homeodomain-interacting protein kinase 3 (circHIPK3) promotes growth and metastasis of glioma cells by sponging miR-124-3p].

作者信息

Xia Lijun, Yi Fuxin, Zhai Xu, Zhang Mingyuan

机构信息

Department of Neurosurgery, First Affiliated Hospital of Jinzhou Medical University, Jinzhou 121000, China.

Department of Neurosurgery, First Affiliated Hospital of Jinzhou Medical University, Jinzhou 121000, China. *Corresponding author, E-mail:

出版信息

Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi. 2020 Jul;36(7):609-615.

Abstract

Objective To investigate the effect of circular RNA homeodomain-interacting protein kinase 3 (circHIPK3) on the proliferation and metastasis of glioma cells via sponging miR-124-3p. Methods T98G cells were transfected with circHIPK3 short hairpin RNA (sh-circHIPK3), pcDNA3.1-circHIPK3, miR-124-3p mimics or pcDNA3.1-WEE1 using Lipofectamine 3000 reagent following the manufacturer's instructions. Real-time quantitative PCR was performed to evaluate the expression of circHIPK3 and miR-124-3p in glioma tissues and cell lines. CCK-8 assay was employed to assess the proliferation of T98G cells. Transwell assay was applied to validate the invasion of T98G cells. The targeting relationship among miR-124-3p, circHIPK3 and serine/threonine kinase WEE1 were verified by dual-luciferase reporter gene assay. The expression of WEE1 and epithelial mesenchymal transition (EMT)-related factors (E-cadherin, N-cadherin and vimentin) were measured by Western blot analysis. In addition, after the competitive binding of circHIPK3 and WEE1 to miR-124-3p, the proliferation of T98G cells was detected by CCK-8 assay; the invasion of T98G cells was evaluated by Transwell assay. Results The circHIPK3 was upregulated in glioma tissues and cell lines. Knockdown of circHIPK3 repressed the proliferation, invasion and EMT of T98G cells. Dual-luciferase reporter gene assay confirmed that miR-124-3p was the target gene of circHIPK3, while WEE1 was the target gene of miR-124-3p. The miR-124-3p was over-expressed simultaneously with circHIPK3 or WEE1. Co-transfected sh-circHIPK3 and pcDNA3.1-WEE1 restored the inhibitory effect of miR-124-3p overexpression on the proliferation, invasion and EMT of T98G cells. Conclusion The circRNA-HIPK3 and WEE1 can promote the proliferation, invasion and EMT of glioma cells by sponging miR-124-3p.

摘要

目的 探讨环状RNA同源结构域相互作用蛋白激酶3(circHIPK3)通过吸附miR-124-3p对胶质瘤细胞增殖和转移的影响。方法 按照制造商说明,使用Lipofectamine 3000试剂将circHIPK3短发夹RNA(sh-circHIPK3)、pcDNA3.1-circHIPK3、miR-124-3p模拟物或pcDNA3.1-WEE1转染至T98G细胞。采用实时定量PCR检测胶质瘤组织和细胞系中circHIPK3和miR-124-3p的表达。使用CCK-8法评估T98G细胞的增殖。采用Transwell法验证T98G细胞的侵袭能力。通过双荧光素酶报告基因检测验证miR-124-3p、circHIPK3和丝氨酸/苏氨酸激酶WEE1之间的靶向关系。采用蛋白质免疫印迹分析检测WEE1和上皮间质转化(EMT)相关因子(E-钙黏蛋白、N-钙黏蛋白和波形蛋白)的表达。此外,在circHIPK3和WEE1与miR-124-3p竞争性结合后,通过CCK-8法检测T98G细胞的增殖;采用Transwell法评估T98G细胞的侵袭能力。结果 circHIPK3在胶质瘤组织和细胞系中表达上调。敲低circHIPK3可抑制T98G细胞的增殖、侵袭和EMT。双荧光素酶报告基因检测证实miR-124-3p是circHIPK3的靶基因,而WEE1是miR-124-3p的靶基因。miR-124-3p与circHIPK3或WEE1同时过表达。共转染sh-circHIPK3和pcDNA3.1-WEE1可恢复miR-124-3p过表达对T98G细胞增殖、侵袭和EMT的抑制作用。结论 circRNA-HIPK3和WEE1可通过吸附miR-124-3p促进胶质瘤细胞的增殖、侵袭和EMT。

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