阿帕替尼抑制鼻咽癌CNE-2Z细胞的生长及机制研究  

Study on Apatinib Inhibiting the Growth of Nasopharyngeal Carcinoma CNE-2Z Cells and Its Mechanism

作　　者：詹德超 陈梓宏 杨东红 杨柳 林晓花 ZHAN Dechao;CHEN Zihong;YANG Donghong;YANG Liu;LIN Xiaohua(Cancer Center,Affiliated Hospital of Guangdong Medical University,Zhanjiang 524000,China)

机构地区：[1]广东医科大学附属医院肿瘤中心,广东湛江524000

出　　处：《中国现代应用药学》2021年第24期3136-3142,共7页Chinese Journal of Modern Applied Pharmacy

基　　金：湛江市科技计划项目(190709174541704)。

摘　　要：目的探讨阿帕替尼对鼻咽癌CNE-2Z细胞生长抑制作用及其可能机制。方法将鼻咽癌CNE-2Z细胞与不同浓度阿帕替尼(0,0.5,1,2μmol·L^(-1))共培养48 h,并分别定义为0,0.5,1,2μmol·L^(-1)组。MTT法检测细胞生长抑制率,Annexin V-FITC/PI联合流式细胞仪检测细胞凋亡率,Western blotting检测Bcl-2、Bax、caspase-3、caspase-9、PI3K、p-PI3K、Akt、p-Akt蛋白表达,RT-PCR检测p-PI3K、p-Akt mRNA表达。按药物不同,将鼻咽癌CNE-2Z细胞分为4组:对照组、阿帕替尼组、740YP组、阿帕替尼+740YP组,48 h后Annexin V-FITC/PI联合流式细胞仪检测细胞凋亡率,Western blotting检测p-PI3K和p-Akt蛋白表达。结果MTT检测结果表明,阿帕替尼呈浓度依赖性抑制CNE-2Z细胞的增殖(P<0.05),阿帕替尼对鼻咽癌细胞的半数抑制浓度(IC50值)为1.518μmol·L^(-1)。Annexin-V/PI联合流式细胞仪检测结果表明,阿帕替尼呈浓度依赖性诱导CNE-2Z细胞凋亡(P<0.05)。Western blotting检测结果表明,阿帕替尼呈剂量依赖性地促进促Bax、caspase-3和caspase-9的表达(P<0.05),抑制Bcl-2的表达(P<0.05)。阿帕替尼能抑制p-PI3K、p-Akt的蛋白和mRNA表达(P<0.05),但对PI3K、Akt蛋白表达无影响。细胞培养48 h后,对照组细胞凋亡率为(10.5±0.96)%,阿帕替尼组为(25.3±1.67)%,740YP组为(6.30±0.52)%,阿帕替尼+740YP组为(11.9±0.61)%,与对照组相比,阿帕替尼组凋亡率上调(P<0.05),740YP组凋亡率下调(P<0.05),阿帕替尼和740YP联用时,凋亡率比740YP组上调(P<0.05)。与对照组相比,阿帕替尼组p-PI3K、p-Akt蛋白表达下调(P<0.05),740YP组p-PI3K、p-Akt蛋白表达上调(P<0.05),阿帕替尼和740YP联用时,p-PI3K、p-Akt蛋白表达比740YP组下调(P<0.05)。结论阿帕替尼可通过抑制PI3K/Akt信号转导通路而诱导鼻咽癌CNE-2Z细胞凋亡,从而达到抑制其增殖的目的。OBJECTIVE To explore the inhibitory effect of apatinib on nasopharyngeal carcinoma CNE-2Z cells and its possible mechanism.METHODS Nasopharyngeal carcinoma CNE-2Z cells combined with different concentrations of apatinib(0,0.5,1,2μmol·L^(-1))were co-cultured for 48 h,and defined as 0,0.5,1 and 2μmol·L^(-1) group,respectively.MTT method was used to detect cell growth inhibition rate.Annexin V-FITC/PI combined with flow cytometry were used to detect cell apoptosis rate.Western blotting was used to detect Bcl-2,Bax,caspase-3,caspase-9,PI3K,p-PI3K,Akt,p-Akt protein expression.RT-PCR was used to detect p-PI3K,p-Akt mRNA expression.According to different drugs,the nasopharyngeal carcinoma CNE-2Z cells were diveded into 4 groups:control group,apatinib group,740YP group,apatinib+740 YP group,48 h later,Annexin V-FITC/PI combined with flow cytometry were used to detect cells apoptosis rate,Western blotting was used to detect the expression of p-PI3K and p-Akt protein.RESULTS MTT test results showed that apatinib inhibited the proliferation of CNE-2Z cells in a concentration-dependent manner(P<0.05),and the half-inhibitory concentration(IC50 value)of apatinib on nasopharyngeal carcinoma cells was 1.518μmol·L^(-1).Annexin-V/PI combined with flow cytometry showed that apatinib induced apoptosis of CNE-2Z cells in a concentration-dependent manner(P<0.05).The result of Western blotting showed that apatinib promoted the expression of Bax,caspase-3 and caspase-9 in a dose-dependent manner(P<0.05),and inhibited the expression of Bcl-2(P<0.05).Apatinib could inhibit the expression of p-PI3K and p-Akt proteins and mRNA(P<0.05),but has no effect on the expression of PI3K and Akt proteins.After cell culture for 48 h,the apoptosis rate of the control group was(10.5±0.96)%,the apatinib group was(25.3±1.67)%,the 740YP group was(6.30±0.52)%,the apatinib+740YP group was(11.9±0.61)%.Compared with the control group,the apoptosis rate in the apatinib group was increased(P<0.05),while it was decreased in the 740YP group(P<0.05).When

关 键 词：阿帕替尼 鼻咽癌 CNE-2Z细胞 PI3K/AKT 凋亡 增殖 

分 类 号：R966[医药卫生—药理学]