黄连素通过调节miRNA-146a促进乳腺癌细胞凋亡  

Berberine promotes apoptosis of human breast cancer cells by regulating miRNA-146a

作　　者：孟昭杰 巴雅尔[4] 张明[3] 陈立[3] 

机构地区：[1]中山大学材料科学与工程学院,广东广州510275 [2]深圳市海普瑞药业股份有限公司,广东深圳518057 [3]吉林大学基础医学院药理系,吉林长春130021 [4]包头市肿瘤医院综合内科,内蒙古包头014030

出　　处：《中国病理生理杂志》2016年第11期1966-1971,共6页Chinese Journal of Pathophysiology

基　　金：吉林省科委基金(No.20140203011YY;No.20150311013YY)

摘　　要：目的:探讨黄连素对乳腺癌MCF-7细胞凋亡的影响及其作用机制。方法:乳腺癌细胞MCF-7采用含10%小牛血清的1640培养基培养,实验分为对照组、黄连素低剂量组、黄连素中剂量组和黄连素高剂量组。给药处理24 h后,采用MTT法检测各组中MCF-7细胞的存活率;Hoechst 33258染色及流式细胞技术观察细胞的凋亡情况;采用Western blot法检测各组MCF-7细胞中NF-κB P65磷酸化水平及促凋亡蛋白Bax和抑凋亡蛋白Bcl-2的表达水平;RT-q PCR法检测细胞中microRNA-146a(miRNA-146a)的水平。为进一步探讨黄连素影响乳腺癌MCF-7细胞凋亡的作用机制,本实验还检测了转染miRNA-146a siRNA后黄连素对促凋亡蛋白Bax和抑凋亡蛋白Bcl-2的mRNA水平的影响。结果:MTT实验结果显示,与对照组相比,黄连素中、高剂量给药组MCF-7细胞的存活率明显降低,且呈一定的剂量依赖性(P<0.01);Hoechst 33258染色观察到给药组细胞核呈致密浓染,或呈碎块状致密浓染;流式细胞技术实验结果亦显示,黄连素给药组MCF-7细胞凋亡率显著增加(P<0.05);Western blot实验结果显示,与对照组比较,黄连素给药组的p-P65和Bcl-2表达水平明显降低,Bax表达水平明显升高,且呈一定的剂量依赖性(P<0.05);RT-q PCR实验结果显示,与对照组比较,黄连素给药组的miRNA-146a表达水平明显升高,且呈一定的剂量依赖性(P<0.05)。黄连素给药联合转染miRNA-146a siRNA后,与黄连素单独给药组相比,MCF-7细胞中Bax mRNA水平显著下降(P<0.05),Bcl-2 mRNA水平显著上调(P<0.05)。结论:黄连素能够促进乳腺癌MCF-7细胞凋亡,其机制可能有部分是通过miRNA-146a抑制NF-κB P65磷酸化,最终影响凋亡相关蛋白Bax/Bcl-2的表达。AIM： To observe the effect of berberine on apoptosis of MCF-7 cells and its potential mechanism.METHODS： The MCF-7 cells were divided into control group and the groups with 3 different doses of berberine. The cell viability was detected by MTT assay,while the cell apoptosis was measured by Hoechst 33258 staining and flow cytometry assay. The protein levels of p-P65,Bax and Bcl-2 were Western blot. The levels of microRNA-146a（ miRNA-146a） in the MCF-7 cells were detected by RT-q PCR. The miRNA-146 a siRNA was transfected to the MCF-7 cells after an evaluation of transfection efficacy,which was co-incubated with berberine to observe its effects on the mRNA levels of Bax and Bcl-2.RESULTS： Compared with control group,the cell viabilities were decreased significantly in medium and high doses of berberine treatment groups with a dose-dependent manner（ P〈0. 01）. The cell apoptosis was increased significantly in medium and high doses of berberine treatment groups dose-dependently（ P〈0. 05）. The protein levels of Bax were up-regula-ted,while those of Bcl-2 and p-P65 were down-regulated significantly by the treatment of berberine（ P〈0. 05）. In addition,the miRNA-146 a levels were increased significantly in medium and high doses of berberine treatment groups（ P〈0. 05） and showed a dose-dependent manner. The mRNA levels of Bax were decreased,while the mRNA levels of Bcl-2were increased after transfection with miRNA-146 a siRNA and co-incubated with berberine. CONCLUSION： Berberine promotes apoptosis of MCF-7 cells. The mechanism may be related to inhibit the activity of NF-κB by incresing the levels of miRNA-146 a.

关 键 词：黄连素 MicroRNA-146a 乳腺癌 细胞凋亡 

分 类 号：R730.23[医药卫生—肿瘤]