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Title: [Effect of miR-513a-3p targeting MDM2 on proliferation, migration and invasion of gastric cancer cells].
Author: Ma YG, Zhang ZS, Yu Y, Xu XF, Yuan L.
Journal: Zhonghua Zhong Liu Za Zhi; 2020 Jan 23; 42(1):30-36. PubMed ID: 32023766.
Abstract:
Objective: To investigate the effects of miR-513a-3p on proliferation, migration and invasion of gastric cancer cells and its mechanism. Methods: 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. Results: 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 (P<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 (P<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 (P<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 (P<0.05). Conclusion: 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. 目的： 探讨miR－513a－3p靶向鼠双微体基因2(MDM2)对胃癌细胞的增殖、迁移和侵袭的影响及其作用机制。 方法： 采用脂质体法将miR－NC、miR－513a－3p、anti－miR－NC、anti－miR－513a－3p、si－NC、si－MDM2、miR－513a－3p＋pcDNA3.1和miR－513a－3p＋pcDNA3.1－MDM2转染至BGC－823细胞中，采用实时荧光定量聚合酶链反应(qRT－PCR)检测miR－513a－3p的表达水平，采用Western blot检测cyclin D1、MMP－2、p21、E－cadherin和MDM2蛋白的表达水平，四甲基偶氮唑蓝法检测各组胃癌细胞BGC－823的活性，Transwell法检测各组胃癌细胞BGC－823的迁移和侵袭能力，双荧光素酶报告基因检测实验检测miR－513a－3p与MDM2的靶向关系。 结果： 胃癌细胞BGC－823、MGC－803中miR－513a－3p的表达水平分别为0.21±0.02和0.34±0.03，与胃上皮细胞GES－1(0.76±0.08)比较，差异均有统计学意义(均P<0.05)。培养24、48和72 h后，miR－NC组细胞的吸光度(A)值分别为0.57±0.05、1.03±0.10和1.43±0.14，miR－513a－3p组细胞的A值分别为0.36±0.03、0.48±0.05和0.63±0.06，差异均有统计学意义(均P<0.05)；miR－NC组细胞的迁移和侵袭数分别为(130±11.80)个和(117±10.60)个，miR－513a－3p组细胞分别为(58±5.64)个和(50±5.13)个，差异均有统计学意义(均P<0.05)。培养24、48和72 h后，si－NC组细胞的A值分别为0.53±0.05、0.95±0.10和1.36±0.14，si－MDM2组细胞的A值分别为0.39±0.04、0.57±0.06和0.80±0.08；si－NC组细胞的迁移和侵袭数分别为(141±12.02)个和(109±10.60)个，si－MDM2组的迁移和侵袭数分别为(66±6.67)个和(61±6.18)个，差异均有统计学意义(均P<0.05)。培养24、48和72 h后，miR－513a－3p＋pcDNA3.1组细胞的A值分别为0.34±0.03、0.46±0.05和0.61±0.06，miR－513a－3p＋pcDNA3.1－MDM2组细胞的A值分别为0.48±0.05、0.82±0.08和1.17±0.12，差异均有统计学意义(均P<0.05)；miR－513a－3p＋pcDNA3.1组细胞的迁移和侵袭数分别为(56±5.71)个和(51±5.16)个，miR－513a－3p＋pcDNA3.1－MDM2组分别为(113±10.28)个和(104±10.02)个，差异均有统计学意义(均P<0.05)。 结论： miR－513a－3p可抑制胃癌细胞的增殖、迁移和侵袭能力，其机制可能与靶向调控MDM2的表达有关，可为胃癌的预防和治疗提供新靶点。.

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