顺铂耐药乳腺癌细胞株的建立及FANCF基因在耐药中的作用  

Establishment of cisplatin-resistant breast cancer cell line and role of FANCF gene in cisplatin resistance

作　　者：马芸[1,2] 李姝墨 蒋爱梅[2] 董坚[1] MA Yun;LI Shumo;JIANG Aimei;DONG Jian(The Third Affiliated Hospital of Kunming Medical University, Kunming 650106, Yunnan, China;Department of Breast Surgery, The First Affiliated Hospital of Kunming Medical University, Kunming 650032, Yunnan, China)

机构地区：[1]昆明医科大学第三附属医院,云南昆明650106 [2]昆明医科大学第一附属医院乳腺外科,云南昆明650032

出　　处：《中国肿瘤生物治疗杂志》2018年第6期607-612,共6页Chinese Journal of Cancer Biotherapy

基　　金：国家自然科学基金资助项目(No.81660472);云南省卫生系统领军人才培养计划资助项目(No.L-201212);云南省细胞治疗技术转化医学重点实验室资助项目(No.2015DG034)~~

摘　　要：目的:探讨三阴性乳腺癌顺铂(cisplatin,DDP)耐药细胞和敏感细胞中FA/BRCA通路关键基因FANCF的表达和功能,以及与DDP耐药的相关性。方法:DDP浓度递增法诱导建立乳腺癌细胞MDA-MB-231的DDP耐药细胞株MDA-MB-231/DDP;通过RNAi技术敲减MDA-MB-231敏感细胞和DDP耐药细胞中FANCF,并在m RNA和蛋白水平进行敲减效果验证。CCK-8法检测DDP耐药细胞株增殖活性,Western blotting法检测该细胞中FANCF蛋白表达,流式细胞仪检测MDA-MB-231细胞周期和凋亡情况,实时定量PCR(RT-q PCR)法检测FANCF m RNA的表达。结果:MDA-MB-231细胞DDP诱导3个月建立的MDA-MB-231/DDP细胞株耐药指数为13.5,其G0/G1期细胞增多、S期和G2/M期细胞减少。MDA-MB-231/DDP细胞中FANCF m RNA和蛋白表达水平显著升高(均P<0.01)。FANCF敲低后MDA-MB-231/DDP细胞凋亡增加,细胞对DDP的药物敏感性显著升高(均P<0.01)。结论:FANCF基因通过抗凋亡作用导致MDA-MB-231细胞对DDP的耐药性,FANCF是乳腺癌治疗的一个潜在靶点。Objective： To investigate the expression profile and function of FANCF gene（a key gene in FA/BRCA pathway） in both cisplatin（DDP）-resistant and DDP-sensitive human triple-negative breast cancer cell lines and to analyze its correlation with DDP-resistance in breast cancer. Methods： The DDP-resistant breast cancer MDA-MB-231 cell line（MDA-MB-231/DDP） was established by induction of gradient DDP. The expression of FANCF gene in both sensitive and resistant cell lines was knocked-down by RNAi interference technology and the knockdown efficiency was validated at both RNA and protein level. The cell viability of MDA-MB-231 cells and MDA-MB-231/DDP cells was determined by the CCK8 assay; Flow cytometry was used to examine the cell cycle distribution and apoptosis; the m RNA and protein expressions of FANCF gene were examined by using q RT-PCR and western blotting, respectively. Results： The resistance index of MDA-MB-231/DDP cells was 13.5 after 3-month induction. The m RNA and protein expressions of FANCF were significantly increased in MDA-MB-231/DDP cells（all P〈0.01）. Cell cycle analysis indicated that the DDP treatment significantly induced G0/G1 arrest and decreased the cell proportion in phase S and G2/M. si RNA-mediated knockdown of FANCF could not only be able to increase sensitivity of MDA-MB-231 to DDP but also promote the cell apoptosis（all P〈0.01）. Conclusion： FANCF attributes to the occurrence of DDP-resistance through anti-apoptosis effect, which might be served as a potential treatment target for drug-resistant human breast cancer.

关 键 词：乳腺癌 顺铂 MDA-MB-231/DDP细胞 耐药 FANCF基因 

分 类 号：R730.5[医药卫生—肿瘤] R737.9[医药卫生—临床医学]