Artemis影响人肝癌细胞BEL-7402/5-FU化疗药物敏感性的研究（英文）  

Effect of Artemis on chemotherapy sensitivity in hepatocellular carcinoma cell line BEL-7402/5-FU

作　　者：李宇婷[1] 张雪[1] 范芳[1] 李大玉[1] 余春波[1] 陈佳瑜[1] 刘云 李长福[1] Li Yuting 1,Zhang Xue 1,Fan Fang 1,Li Dayu 1,Yu Chunbo 1,Chen Jiayu 1,Liu Yun 2, Li Changfu 1(1.Department of Biochemistry,Zunyi Medical University,Zunyi Guizhou 563099,China; 2.Research Center for Medicine and Biology,Zunyi Medical University,Zunyi Guizhou 563099,China)

机构地区：[1]遵义医学院生物化学教研室,贵州遵义563099 [2]遵义医学院医学与生物学研究中心,贵州遵义563099

出　　处：《遵义医学院学报》2018年第3期264-271,共8页Journal of Zunyi Medical University

基　　金：贵州省社会发展攻关项目(NO:黔科合SY字[2013]3004);贵州省科技厅基金资助项目(NO:黔科合J字LKZ[2011]33);贵州省教育厅特色重点实验室建设项目(NO:[2014]212)

摘　　要：目的探讨Artemis对人肝癌耐药细胞株BEL-7402/5-FU化疗药物敏感性的影响及其可能机制。方法将p SGU6/GFP/Neo/sh Artemis干扰质粒转染人肝癌耐药细胞BEL-7402/5-FU(sh Artemis组),通过Real-time PCR法检测P-gp基因表达水平变化;MTT法检测sh Artemis组细胞对5-氟尿嘧啶、丝裂霉素及索拉菲尼的IC50值和RI值;利用Western-blot法检测p-Artemis和ATM蛋白水平表达变化;使用ATM抑制剂KU55933联合丝裂霉素作用细胞48 h,Westernblot法观察ATM、γ-H2AX和Artemis蛋白水平表达变化。结果与对照组比较,sh Artemis组的P-gp mRNA表达水平下降(P<0.05),且对5-氟尿嘧啶、丝裂霉素和索拉菲尼的敏感性增加,IC50值和RI值均降低(P<0.05)。sh Artemis组的pArtemis与ATM蛋白表达水平降低(P<0.05);KU55933联合丝裂霉素作用细胞后,有效减少ATM蛋白的表达(P<0.05),同时γ-H2AX蛋白表达量增加(P<0.05),但Artemis蛋白表达无明显变化(P>0.05)。结论靶向干扰Artemis的表达能够增强细胞BEL-7402/5-FU对化疗药物的敏感性。此效应可能与Artemis作为BEL-7402/5-FU DNA损伤修复的分子开关,进而通过影响ATM参与DNA损伤修复有关。Objective In this study,we seek to investigate whether Artemis is involved in multidrug resistance of the human hepatocellular carcinoma cell line,namely BEL-7402/5-FU cells,after knockdown of Artemis.Methods We transfected p SGU6/GFP/Neo/sh Artemis interference plasmids to BEL-7402/5-FU cells（ sh Artemis group） and examined Artemis and P-gp mRNA by Real-time PCR. We carried out MTT assay to calculate the IC50 values and RI of sh Artemis group induced by antitumor agents 5-fluorouracil,MMC and sorafenib for 48 h,respectively. We performed Western-blot to observe the expression levels of p-Artemis and ATM protein. We detected γ-H2 AX and Artemis proteins changes after treatment of inhibitors of ATM,KU55933,and MMC in BEL-7402/5-FU by Western-blot. Results We observed that sh Artemis group had markedly decreased expression levels of Artemis and P-gp mRNA（ P〈0. 05）,but also significantly reduced the IC50 values and RI of sh Artemis group following treatment with 5-fluorouracil,MMC and sorafenib（ P〈0. 05）. Subsequently,the results of the Western-blot indicated that p-Artemis and ATM proteins expressions were decreased in sh Artemis group（ P〈0. 05）. In this context,further treatment with KU55933 of BEL-7402/5-FU cells exposed to MMC resulted in a lower expression ATM（ P〈0. 05） and an increased expression γ-H2 AX（ P〈0. 05）,however,Artemis protein level was not astatistically significant one（ P〈0. 05）. Conclusion Our analysis suggested that the targeted inhibition of Artemis significantly enhanced chemo sensitivity of BEL-7402/5-FU cells,and this effect appears to be associated with the function of Artemis acting as a molecular switch involved in DNA damage repair by regulating ATM.

关 键 词：ARTEMIS 化疗敏感性 ATM BEL-7402/5-FU 肝癌 

分 类 号：R730.43[医药卫生—肿瘤]