内皮素受体B基因过表达对乳腺癌ZR-75-1细胞活力和周期的影响  

Effect of endothelin receptor B gene overexpression on the viability and cycle of breast cancer ZR-75-1 cells

作　　者：李建超 岳丽玲 高秀丽 朱文斌 高宇 郑立红[3] 刘立琨 LI Jian-chao;YUE Li-ling;GAO Xiu-li;ZHU Wen-bin;GAO Yu;ZHENG Li-hong;LIU Li-kun(School of Medical Science and Technology,Qiqihar Medical University,Qiqihar 161006,Heilongjiang Province,China;Research Institute of Medicine and Pharmacy,Qiqihar Medical University,Qiqihar 161006,Heilongjiang Province,China;Basic Medical Science College,Qiqihar Medical University,Qiqihar 161006,Heilongjiang Province,China)

机构地区：[1]齐齐哈尔医学院医学技术学院,黑龙江齐齐哈尔161006 [2]齐齐哈尔医学院医药科学研究院,黑龙江齐齐哈尔161006 [3]齐齐哈尔医学院基础医学院,黑龙江齐齐哈尔161006

出　　处：《中国临床药理学杂志》2023年第5期664-668,共5页The Chinese Journal of Clinical Pharmacology

基　　金：齐齐哈尔市科技计划联合引导基金资助项目(LHYD-2021006);齐齐哈尔医学科学院基金资助项目(2021-ZDPY-010)。

摘　　要：目的探讨过表达内皮素受体B(EDNRB)基因对人乳腺癌ZR-75-1细胞活力和细胞周期的影响。方法设计EDNRB基因特异性引物,与GV492载体连接构建含EDNRB基因全长序列的过表达载体,慢病毒包装后转染ZR-75-1细胞,构建稳定过表达EDNRB的乳腺癌细胞系。用半定量逆转录聚合酶链反应和蛋白质印迹法检测EDNRB mRNA和蛋白的表达水平,用噻唑蓝法检测EDNRB过表达对细胞生长的影响,用流式细胞术检测EDNRB过表达对细胞周期的影响,用蛋白质印迹法检测EDNRB过表达对细胞周期相关蛋白表达的影响。结果未处理组、GV492转染组和GV492-EDNRB转染组的EDNRB mRNA表达水平分别为0.22±0.13、0.13±0.10和1.79±0.12,EDNRB蛋白相对表达水平分别为0.75±0.04、0.80±0.21和1.43±0.10,GV492-EDNRB转染组的上述指标与未处理组和GV492转染组比较,差异均有统计学意义(均P<0.01)。未处理组和GV492-EDNRB转染组的细胞存活率分别为0.98±0.58和0.62±0.15,G1期细胞百分比分别为(52.10±1.25)%和(63.35±1.06)%,抑癌蛋白53(p53)蛋白相对表达水平分为0.19±0.03和0.36±0.05,p21蛋白相对表达水平分为0.42±0.04和0.78±0.17,细胞周期蛋白1(CCND1)蛋白相对表达水平分为0.34±0.11和0.79±0.27,周期蛋白依赖性激酶4(CDK4)蛋白相对表达水平分为0.83±0.14和0.36±0.01,差异均有统计学意义(均P<0.05)。结论本实验成功构建了EDNRB基因过表达的乳腺癌ZR-75-1细胞系;EDNRB过表达抑制了ZR-75-1细胞生长,并将细胞周期阻滞在G1期,其机制可能与p53-p21-CCND1/CDK4通路有关。Objective To investigate the effect of endothelin receptor B(EDNRB)gene overexpression on the viability and cell cycle in breast cancer ZR-75-1 cells.Methods The specific primers of EDNRB gene were designed,the full-length sequence of EDNRB gene was amplified,and the overexpression vector containing the full-length sequence of EDNRB gene was constructed by lentivirus lection and transfected into ZR-75-1 cells to construct the breast cancer cell line stably overexpressing EDNRB.The expression of EDNRB on mRNA and protein levels were determined by semi-quantitative reverse transcription-polymerase chain reaction and western blot(gray value).The effect on the cell viability was measured by methyl thiazolyl tetrazolium method.The cell cycle status was measured by flow cytometry;the expression level of cycle-related proteins in both groups was detected by western blot(gray value).Results The mRNA expression levels of EDNRB in untreated group,GV492transfected group and GV492-EDNRB transfected group were 0.22±0.13,0.13±0.10 and 1.79±0.12;the protein expression levels of EDNRB in these groups were 0.75±0.04,0.80±0.21 and 1.43±0.10.The differences of above indicators were statistically significant between the GV492-EDNRB transfected group and the untreated and GV492transfected groups(all P<0.05).The survival rates of untreated and GV492-EDNRB transfected groups were0.98±0.58 and 0.62±0.15;the percentage of cells in each phase of the cell cycle were(52.10±1.25)%and(63.35±1.06)%in G1 phase;the expression levels of tumor suppressor protein 53(p53)were 0.19±0.03 and0.36±0.05;the expression levels of p21 were 0.42±0.04 and 0.78±0.17;the expression levels of Cyclin D1(CCND1)were 0.34±0.11 and 0.79±0.27;the expression levels of cyclin dependent kinase 4(CDK4)were0.83±0.14 and 0.36±0.01.The differences of the above indicators were statistically significant(all P<0.05).Conclusion We successfully constructed the breast cancer ZR-75-1 cell line with EDNRB gene overexpression.Overexpression of EDNRB inhibited the gr

关 键 词：内皮素受体B ZR-75-1细胞 细胞生长 G1期阻滞 抑癌蛋白53-抑癌蛋白21-细胞周期蛋白1/周期蛋白依赖性激酶4信号通路 

分 类 号：R97[医药卫生—药品]