| 范清丽,张敏,吴楠,等.纳布啡通过调控 miR-4301/BRD4抑制肝癌细胞增殖、迁移和侵袭的机制研究[J].安徽医药,2022,26(1):21-25. |
| 纳布啡通过调控 miR-4301/BRD4抑制肝癌细胞增殖、迁移和侵袭的机制研究 |
| Study on the mechanism of nalbuphine inhibiting the proliferation, migration and invasion of hepatoma cells by regulating miR-4301/BRD4 |
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| DOI：10.3969/j.issn.1009-6469.2022.01.005 |
| 中文关键词: 肝肿瘤 纳布啡 微小 RNA-4301 溴结构域蛋白 4 肿瘤侵润 增殖 |
| 英文关键词: Liverneoplasms Nalbuphine miRNA-4301 Bromodomain-containingprotein4 Neoplasminvasiveness Proliferation |
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| 中文摘要: |
| 目的探讨纳布啡对肝癌细胞增殖、迁移和侵袭的影响及作用机制。方法本研究时间为 2019年 1—7月。肝癌细胞株 Huh7购自美国 ATCC，将 Huh7细胞分为对照组、不同浓度纳布啡( 50 μmol、100 μmol、200 μmol)组、微小 RNA-4301(miR-4301)组、 miR-4301模拟物阴性对照( miR-NC)组、纳布啡 +miR-4301抑制表达载体( anti-miR-4301)组、纳布啡 +miR-4301抑制表达载体阴性对照( anti-miR-NC)组。四甲基偶氮唑盐比色法( MTT)检测 Huh7细胞的增殖抑制率； Transwell检测 Huh7细胞的迁移和侵袭数；蛋白质印迹检测细胞周期蛋白依赖性激酶抑制剂 1A(p21)、细胞周期蛋白 D1(Cyclin D1)、基质金属蛋白酶 2(MMP-2)、基质金属蛋白酶 9(MMP-9)和溴结构域蛋白 4(BRD4)蛋白表达水平；实时荧光定量聚合酶链式反应( RT-qPCR)检测 miR-4301和 BRD4 mRNA表达水平；荧光素酶报告实验检测 miR-4301和 BRD4的靶向关系。结果与对照组相比，不同浓度纳布啡组 Huh7细胞抑制率升高［(11.25±1.12)%、(22.63±2.25)%、(37.65±3.74)%比( 0.00±0.01)%］迁移数减少［(79.32±7.38)个、(61.36±5.48)个、(45.69±5.37)个比( 98.63±9.54)个］，侵袭数减少［( 67.53±6.65)个、(53.41±5.2，2)个、(37.64±3.87)个比(81.36±8.13)个］p21表达水平升高， Cyclin D1、MMP-2、MMP-9表达水平降低， miR-4301表达水平升高， BRD4mRNA和蛋白表达水平降低( P<0.0，5)，且呈浓度依赖性。与 miR-NC组相比， miR-4301组 Huh7细胞抑制率升高［( 31.25±3.15)%比( 6.32± |
| 英文摘要: |
| Objective To investigate the effect and mechanism of nalbuphine on the proliferation, migration and invasion of hepato-ma cells.Methods This experiment started in January and ended in July 2019. The liver cancer cell line Huh7 was purchased fromATCC in the United States, and Huh7 cells were assigned into control group, different concentrations of nalbuphine (50 μmol, 100 μmol,200 μmol) group, MicroRNA-4301 (miR-4301) group, miR-4301 mimic negative control (miR-NC) group, nalbuphine+miR-4301 inhibi-tory expression vector (anti-miR-4301) group, nalbuphine+miR-4301 suppression expression vector negative control (anti-miR-NC)group. Tetramethylazolium salt colorimetric method (MTT) was used to detect the proliferation inhibition rate of Huh7 cells; Transwellwas used to detect the number of migration and invasion of Huh7 cells; Western blot was used to detect cyclin-dependent kinase inhibitor 1A (p21), Cyclin D1, matrix metalloproteinase 2 (MMP-2), matrix metalloproteinase 9 (MMP-9) and bromodomain protein 4 (BRD4) pro-tein expression levels; the expression levels of miR-4301 and BRD4 mRNA were detected by real-time PCR (RT-qPCR); the luciferase re-porter assay was used to detect the targeting relationship of miR-4301 and BRD4.Results Compared with the control group, the inhibi-tion rate of Huh7 cells in the different concentrations of nalbuphine group was increased [(11.25±1.12)%, (22.63±2.25)%, (37.65±3.74)% vs. (0.00±0.01)%], migration number was reduced [(79.32±7.38), (61.36±5.48), (45.69±5.37) vs. (98.63±9.54)], and the number of inva-sions was reduced [(67.53±6.65), (53.41±5.22), ( 37.64±3.87) vs. (81.36±8.13)], p21 expression level was increased, Cyclin D1, MMP-2, MMP-9 expression levels were decreased, miR-4301 expression level was increased, BRD4 mRNA and protein expression level was de-creased(P<0.05), and it was concentration-dependent. Compared with the miR-NC group, the inhibition rate of Huh7 cells in the miR-4301 group was increased [(31.25±3.15)% vs. (6.32±0.63)%], and the number of migration was decreased [(52.33±5.26) vs. (96.32±9.54), the number of invasions was decreased [(41.69±4.32) vs. (83.15±8.33)], the expression level of p21 was increased, and the expression lev-els of Cyclin D1, MMP-2, MMP-9 were decreased(P<0.05). miR-4301 targeted and regulated the expression of BRD4, and inhibited the expression of miR-4301 to reverse the effect of nalbuphine on Huh7 cells.Conclusion Nalbuphine can inhibit the proliferation, migra-tion and invasion of liver cancer Huh7 cells, and its mechanism may be related to miR-4301 and BRD4. |
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