微小RNA-423靶向调控BAP1表达及对肺鳞癌细胞增殖和凋亡的影响  

Targeted regulation of microRNA-423 on BAP1 expression and its effect on proliferation and apoptosis of lung squamous cell carcinoma cells

作　　者：周小敏 赵展庆 林奇栋 余秉昌 黄国定 ZHOU Xiaomin;ZHAO Zhanqing;LIN Qidong;YU Bingchang;HUANG Guoding(Department of Critical Care Medicine,Western Central Hospital of Hainan,Danzhou 571799,China)

机构地区：[1]海南西部中心医院重症医学科

出　　处：《临床肿瘤学杂志》2019年第8期678-683,共6页Chinese Clinical Oncology

摘　　要：目的探讨微小RNA-423(miR-423)对BRCA1相关蛋白1(BAP1)表达及肺鳞癌细胞NCI-H2170增殖和凋亡的影响。方法采用miR-Pathway和Kaplan-Meier Plotter在线分析肺腺癌和肺鳞癌中miR-423的表达、调控信号通路及与预后的关系。脂质体法向NCI-H2170细胞分别转染miR-423模拟物(过表达组)和抑制物(抑制组),以未转染的细胞为对照组;采用实时定量PCR(QPCR)检测NCI-H2170细胞的miR-423水平,活细胞计数CCK-8试剂盒检测增殖情况,AnnexinⅤ-FITC/PI双染流式细胞术检测凋亡情况,双荧光素酶报告基因实验验证miR-423与BAP1的靶向关系,Western blotting检测BAP1水平。结果在线分析结果提示miR-423在肺腺癌中调控8条信号通路,在鳞癌中调控35条信号通路;与正常组织相比,miR-423在肺腺癌组织表达降低而在肺鳞癌中表达升高;miR-423水平与腺癌的预后无关,而仅与鳞癌的预后有关(HR=0.63,95%CI:0.46~0.87,P=0.004)。QPCR结果显示与对照组(1.024±0.206)相比,过表达组的miR-423水平升高为3.162±0.856,而抑制组降低为0.215±0.043,差异有统计学意义(P<0.05);与对照组相比,过表达组48、72 h的细胞增殖水平升高,而抑制组降低,差异有统计学意义(P<0.05)。流式细胞术结果显示与对照组的(7.352±1.434)%相比,过表达组的凋亡率降低为(2.851±0.723)%,而抑制组的凋亡率升高为(16.028±4.206)%,差异有统计学意义(P<0.05)。miR-423模拟物可抑制BAP1野生型3′端非翻译区的相对荧光素酶活性,但对突变型BAP1的无影响;与对照组(0.611±0.016)相比,过表达组的BAP1水平降低为0.214±0.054,而抑制组升高为0.851±0.074(P<0.05)。结论miR-423在肺鳞癌的发生发展中发挥促癌基因的作用,可以通过靶向BAP1来调控癌细胞的增殖并抑制凋亡,有望成为肺鳞癌诊断和新型生物治疗的靶点。Objective To investigate the effect of microRNA-423(miR-423)on the expression of BRCA1-associated protein 1(BAP1)as well as the proliferation and apoptosis of lung squamous cell carcinoma NCI-H2170 cells.Methods The expression of miR-423 in lung adenocarcinoma and squamous cell carcinoma,its regulation signaling pathway and relationship with prognosis were analyzed by using miR-Pathway and Kaplan-Meier Plotter online.NCI-H2170 cells were transfected with miR-423 mimics(Overexpression group)and inhibitors(Inhibition group)by liposome method,and the untransfected cells were used as Control group.Real-time quantitative PCR(QPCR)was used to detect the miR-423 level in NCI-H2170 cells.CCK-8 kit was used to detect the proliferation of NCI-H2170 cells and AnnexinⅤ-FITC/PI double-staining flow cytometry was used to detect apoptosis.Double luciferase reporter gene experiment was used to verify the targeting relationship between miR-423 and BAP1,and Western blotting was used to detect the level of BAP1.Results Online analysis indicated that miR-423 regulated 8 signaling pathways in lung adenocarcinoma and 35 signaling pathways in squamous cell carcinoma.Compared with normal tissues,miR-423 decreased in lung adenocarcinoma tissues but increased in lung squamous cell carcinoma tissues;miR-423 was only related to the prognosis of squamous cell carcinoma(HR=0.63,95%CI:0.46-0.87,P=0.004).Compared with the Control group(1.024±0.206),the miR-423 level in the Overexpression group increased to 3.162±0.856,while that in the Inhibition group decreased to 0.215±0.043 with a significant difference(P<0.05).Compared with the Control group,the cell proliferation level increased in the Overexpression group at 48 and 72 h,while decreased in the Inhibition group(P<0.05).The results of flow cytometry showed that compared with the Control group(7.352±1.434)%,the apoptotic rate of the Overexpression group decreased to(2.851±0.723)%,while that of the Inhibition group increased to(16.028±4.206)%with a significant difference(P<0.05).MiR-42

关 键 词：肺鳞癌 微小RNA-423 增殖 凋亡 BRCA1相关蛋白1 

分 类 号：R734.2[医药卫生—肿瘤]