胡宁,吴磊,徐建华,郑森,黄亚辉,孟勇
HU Ning,WU Lei,XU Jianhua,ZHENG Sen,HUANG Yahui,MENG Yong

• 1:咸宁市中心医院胃肠外科

摘要(Abstract)：

目的 探讨微小RNA-543(miR-543)通过RNA结合蛋白(quaking, QKI)对结直肠癌HCT116/FU细胞5-氟尿嘧啶(5-FU)敏感性的调控作用。方法 不同浓度5-FU处理HCT116和HCT116/FU后，根据细胞增殖活力计算IC_(50)。将HCT116/FU细胞分为阴性对照组(NC组)、miR-543抑制物组(miR-543 inhibitor组)、过表达QKI组(QKI组)及过表达QKI+miR-543模拟物组(QKI+miR-543组)。分别检测细胞增殖活力、细胞凋亡及凋亡相关蛋白、QKI和Cyclin D1表达、细胞迁移和细胞周期。结果 miR-543在HCT116/FU细胞中高表达，并且HCT116/FU对5-FU的IC_(50)高于HCT116。敲降miR-543减少HCT116/FU细胞增殖活力、迁移数和细胞周期；增加细胞凋亡和cleaved caspase-3、Bax蛋白表达，并且减少Bcl-2和Cyclin D1表达。生物信息学和双荧光素酶报告系统表明QKI是miR-543的下游靶基因。过表达QKI后减少HCT116/FU细胞增殖活力、迁移数和细胞周期，增加细胞凋亡和cleaved caspase-3、Bax蛋白表达，并且减少Bcl-2和Cyclin D1表达；过表达miR-543后缓解过表达QKI对HCT116/FU细胞的不利影响。结论 miR-543靶向下调QKI降低结直肠癌HCT116/FU细胞对5-FU敏感性。
Objective To investigate the regulatory effect of microRNA-543(miR-543)on the 5-fluorouracil(5-FU)sensitivity of colorectal cancer HCT116/FU cells through Quaking(QKI).Methods HCT116 and HCT116/FU cellswere treated with different concentrations of 5-FU and IC_(50) were calculated.HCT116/FU cells were divided into NC group, miR-543 inhibitor group, QKI group and QKI+ miR-543 group.The cell viability, the expression of apoptosis-related proteins, cell migration, apoptosis and cell cycle were determined.Results miR-543 highly expressed in HCT116/FU cells and the IC_(50) were higher in HCT116/FU compared with HCT116 by 5-FU treatment.Knockdown of miR-543 decreased the viability, migration and cycle of HCT116/FU cells, increased cell apoptosis and apoptosis-related proteins of cleaved caspase-3 and Bax expressions, but reduced Bcl-2 and Cyclin D1 proteins expression.Dual-luciferase reporter gene indicated that QKI is target gene of miR-543.Overexpression of QKI significantly reduced the viability, migration and cycle of HCT116/FU cells, which were reversed with overexpression of miR-543.Overexpression of QKI significantly increased cell apoptosis and apoptosis-related proteins of cleaved caspase-3 and Bax expressions but was reversed with overexpression of miR-543.On the contrary, the expression of Bcl-2 and Cyclin D1 proteins were decreased after overexpression of QKI but were reversed with overexpression of miR-543.Conclusion miR-543 reduced the 5-FU sensitivity of colorectal cancer HCT116/FU cells by targeting downregulation QKI.

关键词(KeyWords)： 结直肠癌;5-氟尿嘧啶;敏感性;微小RNA-543;RNA结合蛋白QKI
colorectal cancer;5-fluorouracil;sensitivity;microRNA-543;QKI

Abstract:

Keywords:

基金项目(Foundation): 湖北省卫生和计划生育委员会科研资助项目(WJ2017F113)

作者(Author): 胡宁,吴磊,徐建华,郑森,黄亚辉,孟勇
HU Ning,WU Lei,XU Jianhua,ZHENG Sen,HUANG Yahui,MENG Yong

DOI: 10.14066/j.cnki.cn21-1349/r.2020.1241

参考文献(References)：

• [1] FANG L,JIANG Y,YANG Y,et al.Determining the optimal 5-FU therapeutic dosage in the treatment of colorectal cancer patients[J].Oncotarget,2016,7(49):81880-81887.
• [2] LIU G,ZHOU J,DONG M.Down-regulation of miR-543 expression increases the sensitivity of colorectal cancer cells to 5-Fluorouracil through the PTEN/PI3K/AKT pathway[J].Bio Rep,2019,39(3):BSR20190249.
• [3] HE B,GAO S Q,HUANG L D,et al.MicroRNA-155 promotes the proliferation and invasion abilities of colon cancer cells by targeting quaking[J].Mol Med Rep,2015,11(3):2355-2359.
• [4] LIU Y,ZHANG Y,WU H,et al.miR-10a suppresses colorectal cancer metastasis by modulating the epithelial-to-mesenchymal transition and anoikis[J].Cell Death and Dis,2017,8(4):e2739.
• [5] LV L,LI Q,CHEN S,et al.miR-133b suppresses colorectal cancer cell stemness and chemoresistance by targeting methyltransferase DOT1L[J].Exper Cell Res,2019,385(1):111597.
• [6] CHEN J C,HSIEH Y Y,LO H L,et al.In vitro and in silico mechanistic insights into miR-21-5p-mediated topoisomerase drug resistance in human colorectal cancer cells[J].Biomolecules,2019,9(9):467.
• [7] LI R R,LI C L,QIN F,et al.Effects of germacrone on lipopolysaccharide-induced oxidative stress injury and apoptosis of cardiac myocytes by regulating miR-9a-5p/PTEN[J].The Chinese Journal of Clinical Pharmacology(中国临床药理学杂志),2020,36(12):1643-1647.
• [8] ZHANG S,XIA W J,DONG G C,et al.Cyclic RNA molecule circ_0007766 promotes the proliferation of lung adenocarcinoma cells by up-regulating the expression of cyclin D1/cyclinE1/CDK4[J].Chinese Journal of Lung Cancer(中国肺癌杂志),2019,22(5):271-279.
• [9] XIAO J,LV D,ZHOU J,et al.Therapeutic inhibition of miR-4260 suppresses colorectal cancer via targeting MCC and SMAD4[J].Theranostics,2017,7(7):1901-1913.
• [10] DU Y,LIU X H,ZHU H C,et al.MiR-543 promotes proliferation and epithelial-mesenchymal transition in prostate cancer via targeting RKIP[J].Cell Phy and Bio,2017,41(3):1135-1146.
• [11] SUN J,ZHOU J,DONG M,et al.Dysregulation of MicroRNA-543 expression in colorectal cancer promotes tumor migration and invasion[J].Mol Carcino,2017,56(1):250-257.
• [12] MUKOHYAMA J,ISOBE T,HU Q,et al.miR-221 targets QKI to enhance the tumorigenic capacity of human colorectal cancer stem cells[J].Cancer Res,2019,79(20):5151-5158
• [13] WU J C,JIANG H M,YANG X H,et al.ING5-mediated antineuroblastoma effects of suberoylanilide hydroxamic acid[J].Cancer Med,2018,7(9):872-936.
• [14] WANG Y C,HE W Y,DONG C H,et al.lncRNA HCG11 regulates cell progression by targeting miR-543 and regulating AKT/mTOR pathway in prostate cancer[J].Cell Biol Int,2019,43(12):1453-1462.
• [15] ZHAO H,DIAO C,WANG X,et al.MiR-543 promotes migration,invasion and epithelial-mesenchymal transition of esophageal cancer cells by targeting phospholipase A2 group IVA[J].Cell Phy and Bio,2018,48(4):1595-1604.