| miR-23a调控ESRP1干预直肠癌细胞增殖和凋亡 |
| 投稿时间：2016-11-14 修订日期：2016-11-29 点此下载全文 |
| 引用本文：劳玲娟,宋新江,徐佳.miR-23a调控ESRP1干预直肠癌细胞增殖和凋亡[J].医学研究杂志,2017,46(6):121-125 |
| DOI：
10.11969/j.issn.1673-548X.2017.06.031 |
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| 基金项目:湖南省岳阳市2014年第三批科技基金资助项目 |
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| 中文摘要:目的 分析微小RNA-23a （microRNAs-23a，miR-23a）在直肠癌组织及细胞系中的相对表达水平，以及miR-23a对直肠癌细胞增殖和凋亡的作用，并分析其可能作用机制。方法 采用qRT-PCR检测miR-23a在36例直肠癌组织和癌旁正常组织中的表达水平；利用qRT-PCR检测miR-23a在直肠癌SW480细胞及人正常结肠上皮细胞株NCM460中的表达；合成miR-23a抑制剂（inhibitor） RNA片段和抑制剂阴性对照RNA片段（inhibitor negative control，inhibitor NC），并将其分别转染至SW480细胞后，通过CCK-8法检测miR-23a inhibitor转染SW480细胞后对细胞增殖的影响；流式细胞术检测转染后细胞凋亡率；Western blot法检测上皮剪接调节蛋白1（epithelial splicing regulatory protein 1，ESRPl）蛋白在SW480细胞中的表达水平；构建野生型pGL3-ESRP1-3'UTR （wt-pGL3-ESRP1-3'UTR）或突变型pGL3-ESRP1-3'UTR （mut-pGL3-ESRP1-3'UTR）质粒，HEK293和SW480细胞经上述质粒分别与miR-23a inhibitor或inhibitor NC共转染后测定双荧光素酶活性。结果 与癌旁正常组织相比较，miR-23a在直肠癌组织中的相对表达水平明显上调，差异有统计学意义（P=0.000）；与NCM460细胞相比较，miR-23a在SW480细胞中的表达量显著上调，差异有统计学意义（P=0.000）；SW480细胞转染miR-23a inhibitor后，细胞增殖较inhibitor NC组下降35.54%±5.27%，两组结果差异有统计学意义（P=0.000）；转染miR-23a inhibitor后SW480细胞早期凋亡率明显高于inhibitor NC组（P=0.000）；荧光素酶报告基因结果表明ESRP1是miR-23a的直接靶基因；SW480细胞转染inhibitor NC后对ESRP1蛋白表达无明显影响，而转染miR-23a inhibitor至SW480细胞后ESRP1蛋白表达水平明显升高。结论 miR-23a在直肠癌组织和细胞系中的表达均显著升高，miR-23a可通过下游靶基因ESRP1从而调控直肠癌细胞增殖和凋亡。 |
| 中文关键词:直肠癌 miR-23a ESRP1 凋亡 |
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| miR-23a Regulates Proliferation and Apoptosis of Rectal Cancer via Targeting Gene ESRP1 |
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| Abstract:Objective To elucidate the relative level of miR-23a RNA in rectal cancer tissues and cell line as well as the effects of miR-23a on the cell proliferation and apoptosis of rectal cancer cells in vitro. Methods Quantitative reverse-transcription polymerase chain reaction (qRT-PCR) was applied in assessment of the transcription of miR-23a in rectal cancer tissues and in vitro cells. The RNA fragment of miR-23a inhibitor and inhibitor NC were synthesized and transfected into SW480 cells. Cell proliferation was evaluated with Cell Counting Kit-8 (CCK-8) assay. The apoptotic rate was analyzed by flow cytometry. The expression of ESRP1 was detected by western blot. Wild-type pGL3-ESRP1-3'UTR(wt-pGL3-ESRP1-3'UTR) or mutant pGL3-ESRP1-3'UTR(mut-pGL3-ESRP1-3'UTR) plasmids and miR-23a inhibitor RNA fragments or inhibitor NC RNA fragments were co-transfected into HEK293 and SW480 cells, then the Promega dual luciferase reporter gene assay kit was used to examine the dual luciferase activity in SW480 cells. Results The relative RNA level of miR-23a was significantly promoted in both rectal cancer tissue samples and SW480 cells. After SW480 cells were transfected with miR-23a inhibitor, human rectal cancer cell line SW480 with down-regulation of miR-23a showed significant inhibition of cell proliferation compared with negative control (P=0.000). Furthermore, our data demonstrated clearly that the inhibition of miR-23a promoted apoptosis in SW480 cells (P=0.000). Luciferase assay showed that ESRP1 was a direct target gene of miR-23a. Conclusion The expression of miR-23a is clearly associated with the growth and apoptosis of human rectal cells by targeting ESRP1, whilst miR-23a may be used as a potential therapeutic target for the treatment of rectal cancer in the future. |
| keywords:Rectal cancer miR-23a ESRP1 Apoptosis |
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