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

CLC Number: 

• R782