miR-17-5p在大鼠垂体泌乳素腺瘤MMQ细胞耐药中的作用  

Roles of microRNA-17-5p in the resistance of MMQ cells to Cabergoline

作　　者：管佳清 许家栋[1] 王春勇 苏志鹏[1] 蔡霖[1] 陈贤斌[1] 郑伟明[1] GUAN Jiaqing;XU Jiadong;WANG Chunyong;SU Zhipeng;CAI Lin;CHEN Xianbin;ZHENG Weiming(Department of Neurosurgery,the First Affiliated Hospital of Wenzhou Medical University,Wenzhou 325015,China)

机构地区：[1]温州医科大学附属第一医院神经外科,浙江温州325015

出　　处：《温州医科大学学报》2019年第4期258-262,280,共6页Journal of Wenzhou Medical University

基　　金：浙江省自然科学基金资助项目(LY17H160052;LY19C070002);温州市公益性科技计划项目(Y20170088)

摘　　要：目的:探讨microRNA-17-5p(mi R-17-5p)在大鼠垂体泌乳素腺瘤MMQ细胞耐药中的作用及相关机制。方法:利用miR-17-5p类似物及拮抗物分别过表达和抑制大鼠垂体泌乳素腺瘤MMQ细胞中miR-17-5p的表达,并采用实时荧光定量PCR(RT-q PCR)法进行检测。利用CCK-8法检测miR-17-5p表达调控前后细胞增殖以及对卡麦角林(CAB)治疗反应的变化。通过生物信息学方法预测miR-17-5p的靶点基因,然后采用荧光素酶报告检测法进行验证,继而用qRT-PCR及Western blot法检测miR-17-5p对靶基因表达的影响。慢病毒细胞转染过表达MMQ细胞中的PTEN基因,用Western blot检测PTEN的表达水平,并采用CCK-8法检测PTEN过表达对MMQ细胞耐药性的影响。结果:mi R-17-5p过表达可促进MMQ细胞的增殖,而miR-17-5p敲减则能抑制MMQ细胞的增殖。mi R-17-5p敲减能上调MMQ细胞对CAB治疗的敏感性,相反,miR-17-5p过表达则显著降低MMQ细胞对CAB治疗的敏感性。生物信息学方法预测PTEN为miR-17-5p潜在作用靶点,荧光素酶报告检测法进一步验证了两者的相互作用关系,同时miR-17-5p过表达的MMQ细胞中PTEN mRNA及蛋白表达量显著下降,而miR-17-5p敲减的MMQ细胞中PTEN mRNA及蛋白表达量显著上升。PTEN表达上调能部分逆转miR-17-5p过表达引起的MMQ细胞对CAB治疗敏感性降低。结论:miR-17-5p可通过下调PTEN的表达促进MMQ细胞增殖并降低其对CAB治疗的反应。Objective:To study the roles of microRNA-17-5p may play in the resistance of MMQ cells to dopamine agonists.Methods:MMQ cells were either treated with miR-17-5p mimics or miR-17-5p inhibitor to regulate the expression of miR-17-5p in vitro qPCR was used to verify the transfection efficiency.After treatment,cell viability and its response to CAB was determined with CCK-8 assays.The target gene of miR-17-5p was predicted and confirmed via bioinformatics analysis and luciferase reporter assays.The expression of PTEN was analyzed by qPCR and Western Blotting.In addition we deployed CCK-8 assay to evaluate the response to CAB of MMQ cells which were initially treated with lentiviral vector containing PTEN.Results:Overexpression of miR-17-5p could promote the proliferation of MMQ cells and suppress CAB cytotoxicity.On the contrary,down-regulated of the expression of miR-17-5p could effectively inhibit proliferation and boost susceptibility to CAB in MMQ cells.The bioinformatics database identified the potential target of miR-17-5p and the luciferase reporter assay confirmed that PTEN was the direct target of miR-17-5p.While the expression of PTEN was down-regulated in miR-17-5p over-expressed MMQ cells,the expression of PTEN was up-regulated in miR-17-5p knockdown MMQ cells.Furthermore,PTEN over-expression was able to reverse the drug resistance induced by miR-17-5p on CAB treatment.Conclusion:miR-17-5p promotes MMQ proliferation and mediates CAB resistance by targeting PTEN.

关 键 词：microRNA-17-5p 泌乳素瘤 卡麦角林 PTEN 大鼠 

分 类 号：R736.4[医药卫生—肿瘤]