《Journal of Oral and Maxillofacial Surgery》 ›› 2020, Vol. 30 ›› Issue (3): 156-162. doi: 10.3969/j.issn.1005-4979.2020.03.006

• Basic Scientific Study • Previous Articles     Next Articles

Mechanism of MiR-579-3p Regulating Proliferation and Apoptosis of Oral Squamous Cell Carcinoma Cells by Targeting ACTR3B

GAO Gongjie1(), WANG Haiye2, LI Xiaoyan3   

  1. 1 Zhide Stomatology Hospital of Zhengzhou, Zhengzhou 450052
    2 Department of Stomatology, the First Hospital, Zhengzhou University, Zhengzhou 450033
    3 Department of General Surgery, Zhengzhou People's Hospital East District Branch, Zhengzhou 450003, Henan Province, China
  • Received:2019-11-25 Revised:2020-03-17 Online:2020-06-28 Published:2020-06-26

MiR-579-3p靶向肌动蛋白相关蛋白3B调控口腔鳞状细胞癌细胞增殖和凋亡的机制研究

高功杰1(), 王海业2, 李晓彦3   

  1. 1 郑州植得口腔医院,河南 郑州 450052
    2 郑州大学第一附属医院口腔科,河南 郑州 450033
    3 郑州人民医院郑东院区普外科,河南 郑州 450003
  • 通讯作者: 高功杰,主治医师. E-mail: glq198505@163.com
  • 作者简介:

    高功杰(1979—),男,河南南阳人,主治医师,学士

Abstract:

Objective: To investigate the effect and mechanism of miR-579-3p on proliferation and apoptosis of oral squamous cell carcinoma (OSCC) cells. Methods: The normal human oral keratinocytes (NHOK) and oral squamous carcinoma cell lines CAL27, CAL33 and SCC15 were cultured. The expression levels of miR-579-3p and actin-related protein 3B (ACTR3B) mRNA were detected by quantitative real-time PCR(qRT-PCR). The expression level of ACTR3B protein was detected by western blot. Taking CAL33 cells as the research object, CAL33 cells with miR-579-3p overexpressed or ACTR3B expression silenced were constructed. Then cell proliferation was detected by methylthiazoletrazolium(MTT) assay, apoptosis was detected by flow cytometry, and the expression levels of CyclinD1 and C-caspase-3 protein were detected by western blot. Starbase Bioinformatics software predicts that ACTR3B may be the target gene of miR-579-3p, and the dual luciferase reporter gene assay validates the regulatory relationship between miR-579-3p and ACTR3B. Results: Compared with NHOK cells, the expression levels of miR-579-3p in oral squamous cell carcinoma cell lines CAL27, CAL33 and SCC15 were decreased (P<0.05), and the expression levels of ACTR3B mRNA and protein were increased (P<0.05). Overexpression of miR-579-3p or silencing ACTR3B could decrease the survival rate of CAL33 cells and the expression level of CyclinD1 protein (P<0.05), and increase the apoptosis rate of CAL33 cells and the expression level of C-caspase-3 protein (P<0.05). MiR-579-3p targets negative regulation of ACTR3B expression. Overexpression of ACTR3B reduced the effect of overexpression of miR-579-3p on CAL33 cell viability, apoptotic rate, and the expression level of CyclinD1 and C-caspase-3 protein. Conclusion: Overexpression of miR-579-3p may inhibit the proliferation of oral squamous carcinoma cells and induce apoptosis by down-regulating the expression of ACTR3B.

Key words: miR-579-3p, actin-related protein 3B (ACTR3B), oral squamous cell carcinoma, cell proliferation, apoptosis

摘要:

目的: 探讨miR-579-3p对口腔鳞状细胞癌(oral squamous cell carcinoma, OSCC)细胞增殖和凋亡的影响及作用机制。方法: 培养正常人口腔角质形成细胞(normal human oral keratinocyte,NHOK)和OSCC细胞系CAL27、CAL33、SCC15,实时荧光定量PCR(quantitative real-time PCR,qRT-PCR)检测细胞中miR-579-3p和肌动蛋白相关蛋白3B(actin-related protein 3B,ACTR3B)信使RNA(mRNA)的表达水平,蛋白质印迹法(western blot)检测ACTR3B蛋白表达水平。以CAL33细胞为研究对象,构建过表达miR-579-3p或沉默ACTR3B表达的CAL33细胞,四甲基噻唑蓝染色法(methylthiazoletrazolium,MTT)检测细胞增殖,流式细胞仪检测细胞凋亡,western blot检测细胞周期蛋白D1(CyclinD1)和活化的半胱天冬酶-3(C-caspase-3)蛋白水平。Starbase生物信息学软件预测ACTR3B可能是miR-579-3p的靶基因,双荧光素酶报告基因实验验证miR-579-3p和ACTR3B调控关系。结果: 与NHOK细胞比较,OSCC细胞系CAL27、CAL33和SCC15中miR-579-3p表达水平降低(P<0.05),ACTR3B mRNA和蛋白表达水平升高(P<0.05)。过表达miR-579-3p或沉默ACTR3B表达均可降低CAL33细胞存活率和CyclinD1蛋白表达水平(P<0.05),提高CAL33细胞凋亡率和C-caspase-3蛋白表达水平(P<0.05)。miR-579-3p靶向负调控ACTR3B表达。过表达ACTR3B降低了过表达miR-579-3p对CAL33细胞存活率、凋亡率及CyclinD1和C-caspase-3蛋白表达的影响。结论: 过表达miR-579-3p可能通过下调ACTR3B表达抑制OSCC细胞增殖,并诱导细胞凋亡。

关键词: miR-579-3p, 肌动蛋白相关蛋白3B, 口腔鳞状细胞癌, 细胞增殖, 凋亡

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