CAJ | 학술논문

目的探索miR-200a对卵巢癌化疗敏感性的影响及其可能的作用机制.方法首先利用慢病毒表达载体,在卵巢癌细胞株SKOV-3和ES-2中,构建稳定上调miR-200a的细胞模型;接下来采用MTT实验检测miR-200a对紫杉醇及顺铂化疗敏感性的影响;进一步通过荧光定量PCR和免疫印迹实验,检测miR-200a对耐药相关三磷酸腺苷结合盒转运子(ABC)家族表达水平的影响;最后利用双荧光素酶实验验证miR-200a与ABCG2mRNA 3'末端非编码区(3'-UTR)的相互作用关系.结果 qRT-PCR验证携带miR-200a转基因的SKOV-3和ES-2卵巢癌细胞较对照细胞中miR-200a的表达水平分别升高了3.15和273.76倍;证实miR-200a过表达增加了SKOV-3和ES-2细胞株对紫杉醇的敏感性,但对顺铂的敏感性没有明显影响;miR-200a不同程度的下调了ABC家族(ABCB3,ABCC1,ABCC2,ABCC3,ABCG2)的表达水平,但与ABCG2的3'-UTR并不存在直接相互作用关系.结论 miR-200a可能通过调控耐药相关ABC家族基因的表达,增加了卵巢癌细胞对紫杉醇的敏感性,但其下调ABCG2表达的方式并非直接作用其mRNA的3'-UTR.
목적 탐색miR-200a대란소암화료민감성적영향급기가능적작용궤제.방법 수선이용만병독표체재체,재란소암세포주SKOV-3화ES-2중,구건은정상조miR-200a적세포모형;접하래채용MTT실험검측miR-200a대자삼순급순박화료민감성적영향;진일보통과형광정량PCR화면역인적실험,검측miR-200a대내약상관삼린산선감결합합전운자(ABC)가족표체수평적영향;최후이용쌍형광소매실험험증miR-200a여ABCG2mRNA 3'말단비편마구(3'-UTR)적상호작용관계.결과 qRT-PCR험증휴대miR-200a전기인적SKOV-3화ES-2란소암세포교대조세포중miR-200a적표체수평분별승고료3.15화273.76배;증실miR-200a과표체증가료SKOV-3화ES-2세포주대자삼순적민감성,단대순박적민감성몰유명현영향;miR-200a불동정도적하조료ABC가족(ABCB3,ABCC1,ABCC2,ABCC3,ABCG2)적표체수평,단여ABCG2적3'-UTR병불존재직접상호작용관계.결론 miR-200a가능통과조공내약상관ABC가족기인적표체,증가료란소암세포대자삼순적민감성,단기하조ABCG2표체적방식병비직접작용기mRNA적3'-UTR.
Objective To explore the role of miR-200a in chemosensitivity regulation of ovarian cancer.Methods Firstly miR-200a was up-regulated in ovarian cancer cell lines (SKOV-3 and ES-2) by lentiviral vector.Then the effects of miR-200a on cytotoxicity of paclitaxel and cisplatin were investigated by methyl thiazolyl tetrazolium (MTT).Furthermore miR-200a regulation of chemoresistance associated with ATP-binding cassette (ABC) family genes expression was detected by quantitative real-time polymerase chain reaction (PCR) and Western blot.Finally the interaction between miR-200a and ABCG2 mRNA 3' untranslated region (3'-UTR) was verified by dual-luciferase reporter assay.Results An over-expression of miR-200a were successfully achieved in SKOV-3 and ES-2 cells.MiR-200a enhanced the chemosensitivity of SKOV-3 and ES-2 to paclitaxel,but not to cisplatin.Chemoresistance associated ABC family (ABCB3,ABCC1,ABCC2,ABCC3,ABCG2) were down-regulated by miR-200a at several levels.However,the direct interaction between miR-200a and the 3'-UTR of ABCG2 mRNA was not found.Conclusion An over-expression of miR-200a may increase chemosensitivity to paclitaxel in ovarian cancer cells through negatively regulated chemoresistance associated ABC family.However,no direct action on 3'-UTR of ABCG2 was not found after its down-regulation by miR-200a.