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[微小RNA-513a-3p靶向鼠双微体2对胃癌细胞增殖、迁移和侵袭的影响]

[Effect of miR-513a-3p targeting MDM2 on proliferation, migration and invasion of gastric cancer cells].

作者信息

Ma Y G, Zhang Z S, Yu Y, Xu X F, Yuan L

机构信息

Department of Gastroenterology, the Fifth Affiliated Hospital of Zhengzhou University, Zhengzhou 450052, China.

出版信息

Zhonghua Zhong Liu Za Zhi. 2020 Jan 23;42(1):30-36. doi: 10.3760/cma.j.issn.0253-3766.2020.01.004.

Abstract

To investigate the effects of miR-513a-3p on proliferation, migration and invasion of gastric cancer cells and its mechanism. The miR-NC (miR-negative control mimics), miR-513a-3p (miR-513a-3p mimics), anti-miR-NC, anti-miR-513a-3p, si-NC, si-MDM2 (murine double minute 2), miR-513a-3p+ pcDNA3.1 (co-transfected with miR-513a-3p and pcDNA3.1), miR-513a-3p+ pcDNA3.1-MDM2 (co-transfected with miR-513a-3p and pcDNA3.1-MDM2) were transfected into BGC-823 cells, respectively. The expression of miR-513a-3p was detected by real-time quantitative reverse transcription polymerase chain reaction (qRT-PCR), and the protein expressions of cyclin D1, MMP-2, p21, E-cadherin, MDM2 were detected by western blot. The viability of BGC-823 cells of each group was detected by 3-(4, 5-dimethyl-2-thiazolyl)-2, 5-diphenyl-2-H-tetrazolium bromide (MTT) assay. The migration and invasion of each group were detected by Transwell, the targeting relationship between miR-513a-3p and MDM2 was detected by double luciferase reporter gene assay. The expression of miR-513a-3p in gastric epithelial cells GES-1 was 0.76±0.08, significantly higher than 0.21±0.02 in gastric cancer cells BGC-823 and 0.34±0.03 in MGC-803, respectively (<0.05). The cell viabilities of the miR-NC group at 24 h, 48 h and 72 h were 0.57±0.05, 1.03±0.10, 1.43±0.14, respectively, while those of the miR-513a-3p group were 0.36±0.03, 0.48±0.05, and 0.63±0.06, respectively. The migration and invasion numbers of miR-NC group were 130±11.80 and 117±10.60, respectively, those of miR-513a-3p group were 58±5.64 and 50±5.13, respectively, and the differences were statistically significant (<0.05). The cell viabilities of the si-NC group at 24 h, 48 h and 72 h were 0.53±0.05, 0.95±0.10, 1.36±0.14, respectively. Those of the si-MDM2 group were 0.39±0.04, 0.57±0.06, and 0.80±0.08, respectively. The cell migration and invasion of the si-NC group were 141±12.02 and 109±10.60, respectively, while those of the MDM2 group were 66±6.67 and 61±6.18, respectively, and the differences were statistically significant (<0.05). The cell viabilities of the miR-513a-3p+ pcDNA3.1 group at 24 h, 48 h and 72 h were 0.34±0.03, 0.46±0.05, and 0.61±0.06, respectively. Those of miR-513a-3p+ pcDNA3.1-MDM2 group were 0.48±0.05, 0.82±0.08, 1.17±0.12, respectively. The migration and invasion of miR-513a-3p+ pcDNA3.1 group were 56±5.71 and 51±5.16, respectively, while those of miR-513a-3p+ pcDNA3.1-MDM2 group were 113±10.28 and 104±10.02, respectively, and the differences were statistically significant (<0.05). miR-513a-3p may inhibit the proliferation, migration and invasion of gastric cancer cells through targeting regulation of MDM2, which will provide new targets for the prevention and treatment of gastric cancer.

摘要

探讨miR-513a-3p对胃癌细胞增殖、迁移和侵袭的影响及其机制。分别将miR-NC(miR阴性对照模拟物)、miR-513a-3p(miR-513a-3p模拟物)、抗miR-NC、抗miR-513a-3p、si-NC、si-MDM2(鼠双微体2)、miR-513a-3p+pcDNA3.1(与miR-513a-3p和pcDNA3.1共转染)、miR-513a-3p+pcDNA3.1-MDM2(与miR-513a-3p和pcDNA3.1-MDM2共转染)转染至BGC-823细胞。采用实时定量逆转录聚合酶链反应(qRT-PCR)检测miR-513a-3p的表达,采用蛋白质免疫印迹法检测细胞周期蛋白D1、基质金属蛋白酶-2、p21、E-钙黏蛋白、MDM2的蛋白表达。采用3-(4,5-二甲基-2-噻唑基)-2,5-二苯基-2-H-溴化四氮唑(MTT)法检测各组BGC-823细胞的活力。采用Transwell检测各组细胞的迁移和侵袭能力,采用双荧光素酶报告基因检测法检测miR-513a-3p与MDM2的靶向关系。胃上皮细胞GES-1中miR-513a-3p的表达为0.76±0.08,显著高于胃癌细胞BGC-823中的0.21±0.02和MGC-803中的0.34±0.03(<0.05)。miR-NC组在24 h、48 h和72 h的细胞活力分别为0.57±0.05、1.03±0.10、1.43±0.14,而miR-513a-3p组分别为0.36±0.03、0.48±0.05、0.63±0.06。miR-NC组的迁移和侵袭细胞数分别为130±11.80和117±10.60,miR-513a-3p组分别为58±5.64和50±5.13,差异具有统计学意义(<0.05)。si-NC组在24 h、48 h和72 h的细胞活力分别为0.53±0.05、0.95±0.10、1.36±0.14。si-MDM2组分别为0.39±0.04、0.57±0.06、0.80±0.08。si-NC组的细胞迁移和侵袭能力分别为141±12.02和109±10.60,而MDM2组分别为66±6.67和61±6.18,差异具有统计学意义(<0.05)。miR-513a-3p+pcDNA3.1组在24 h、48 h和72 h的细胞活力分别为0.34±0.03、0.46±0.05、0.61±0.06。miR-513a-3p+pcDNA3.1-MDM2组分别为0.48±0.05、0.82±0.08、1.17±0.12。miR-513a-3p+pcDNA3.1组的迁移和侵袭细胞数分别为56±5.71和51±5.16,而miR-513a-3p+pcDNA3.1-MDM2组分别为113±10.28和104±10.02,差异具有统计学意义(<0.05)。miR-513a-3p可能通过靶向调控MDM2抑制胃癌细胞的增殖、迁移和侵袭,为胃癌的防治提供新靶点。

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