| 韩林.微小 RNA-34a在紫草素抑制肺癌 H1299细胞增殖和侵袭中的作用及其机制[J].安徽医药,2023,27(1):83-87. |
| 微小 RNA-34a在紫草素抑制肺癌 H1299细胞增殖和侵袭中的作用及其机制 |
| Role and mechanism of miR-34a in shikonin inhibiting the proliferation and invasion of lung cancer H1299 cells |
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| DOI：10.3969/j.issn.1009-6469.2023.01.018 |
| 中文关键词: 肺肿瘤 紫草素 微小 RNA-34a 增殖 侵袭 YY1转录因子 |
| 英文关键词: Lung neoplasms Shikonin miR-34a Proliferation Invasion YY1 transcription factor |
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| 中文摘要: |
| 目的探讨微小 RNA-34a(microRNA-34a，miR-34a)在紫草素抑制肺癌 H1299细胞增殖和侵袭中的作用及其机制。方法 2019年 9月至 2020年 3月，采用 1、2.5和 5 mg/L紫草素处理体外培养的肺癌 H1299细胞后，采用噻唑蓝( MTT)法检测细胞的存活率；平板克隆实验和 Transwell实验分别检测细胞克隆形成能力和侵袭能力； RT-PCR检测细胞中 miR-34a的表达情况。采用生物信息软件预测、双萤光素酶报告基因实验检测 miR-34a与 Yin Yang-1(YY1)的靶向关系。采用脂质体法转染 miR-34a抑制剂和 pcDNA3.1-YY1-GFP过表达质粒后， RT-PCR检测细胞中 miR-34a的表达情况，蛋白质印迹法(western blotting)检测细胞 YY1蛋白的表达， MTT法、平板克隆实验和 Transwell小室实验观察下调 miR-34a和上调 YY1表达对紫草素处理的 H1299细胞增殖和侵袭的影响。结果与 0 mg/L紫草素相比， 1、2.5和 5 mg/L紫草素能够呈浓度依赖性抑制 H1299细胞存活率、克隆细胞数［( 127.63±7.38)、(108.85±6.34)、(91.72±6.15)比( 156.55±9.06)］和侵袭细胞数［( 93.12±5.56)、(75.25±4.68)、(59.85±3.20)比( 116.00±7.85)］并促进 miR-34a表达。生物信息软件预测到 miR-34a与 YY1存在互补的结合位点，双萤光素酶报告基因实验证实 YY1是 miR-3，4a的靶基因。下调 miR-34a可逆转紫草素对 H1299细胞增殖和侵袭的抑制作用，并促进 YY1蛋白的表达， |
| 英文摘要: |
| Objective To explore the role of miR-34a in the inhibition of shikonin on the proliferation and invasion of lung cancer H1299 cells and its mechanism.Methods Lung cancer H1299 cells in vitro were treated with 1, 2.5 and 5 mg/L shikonin from September 2019 to March 2020, and the cell viability was detected by methyl thiazolyl tetrazolium (MTT) method. Formation and invasionof cell cloning were detected by plate cloning assay and Transwell assay; RT-PCR was used to detect the expression of miR-34a. The targeting relationship between miR-34a and Yin Yang-1 (YY1) was detected by bioinformation software prediction and dual luciferase reporter gene assay. After transfection of miR-34a inhibitor and pcDNA3.1-YY1-GFP overexpression plasmid by liposome method, the expression of miR-34a was detected by RT-PCR, and the expression of YY1 protein was detected by Western blotting. MTT assay, platecloning experiments and Transwell chamber experiments were performed to observe the effects of down-regulated miR-34a and up-regulated YY1 expression on the proliferation and invasion of shikonin-treated H1299 cells.Results Compared with 0 mg/L shikonin, 1,2.5 and 5 mg/L shikonin could inhibit the survival rate of H1299 cells, the number of cloned cells [(127.63±7.38), (108.85±6.34),(91.72±6.15) vs. (156.55±9.06)] and the number of invasive cells [(93.12±5.56), (75.25±4.68), (59.85±3.20) vs. (116.00±7.85)] in a concentration-dependent manner, and promoted the expression of miR-34a. The bioinformation software predicted the presence of a complementary binding site for miR-34a and YY1, and the dual luciferase reporter gene assay confirmed that YY1 was the target gene of miR-34a. Down-regulation of miR-34a reversed the inhibitory effect of shikonin on proliferation and invasion of H1299 cells, and promoted the expression of YY1 protein. Up-regulation of YY1 expression reversed the inhibitory effect of shikonin on the proliferation and invasion of H1299 cells.Conclusions miR-34a plays an active role in the inhibition of proliferation and invasion of lung cancerH1299 cells by shikonin. The mechanism may be related to the targeted regulation of YY1 expression. |
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