CAJ | 학술논문

目的分析微小RNA(miR)-373在肝门部胆管癌的表达与临床病理因子之间的关系,探讨miR-373靶向甲基CpG结合蛋白2(MBD2)-3’端非翻译区(3' UTR)的机制.方法 TaqMan miRNA Assay检测miR-373的表达,采用QuantiTect_Primer Assays和Western blot法分析甲基CpG结合蛋白(MBPs) mRNA和蛋白水平；双荧光素酶报告基因检测miR-373对MBD2-3' UTR的靶向作用.结果 miR-373在肝门部胆管癌中卜调2.94倍(P＜0.01).与高表达组比较,miR-373低表达与肿瘤细胞低分化(P＜0.05)、高分期(P＜0.05)、短总生存和无病生存时间(P＜0.05)相关.miR-373低表达肿瘤组织中,MBD2的mRNA和蛋白表达分别上调3.75倍(P＜0.01)、2.34倍(P＜0.05)；miR-373前体抑制MBD2-3' UTR荧光素酶活性1.79倍；外源性miR-373和anti-miR-373分别下调QBC939细胞、上调人肝内胆管上皮细胞(HIBEpic)中MBD2表达2.56倍(P＜0.01)和2.13倍(P＜0.01).结论 miR-373通过结合在MBD2-3' UTR,抑制MBD2表达.肝门部胆管癌中,miR-373下调导致MBD2表达增高,与肿瘤进展和预后密切相关.
목적 분석미소RNA(miR)-373재간문부담관암적표체여림상병리인자지간적관계,탐토miR-373파향갑기CpG결합단백2(MBD2)-3’단비번역구(3' UTR)적궤제.방법 TaqMan miRNA Assay검측miR-373적표체,채용QuantiTect_Primer Assays화Western blot법분석갑기CpG결합단백(MBPs) mRNA화단백수평；쌍형광소매보고기인검측miR-373대MBD2-3' UTR적파향작용.결과 miR-373재간문부담관암중복조2.94배(P＜0.01).여고표체조비교,miR-373저표체여종류세포저분화(P＜0.05)、고분기(P＜0.05)、단총생존화무병생존시간(P＜0.05)상관.miR-373저표체종류조직중,MBD2적mRNA화단백표체분별상조3.75배(P＜0.01)、2.34배(P＜0.05)；miR-373전체억제MBD2-3' UTR형광소매활성1.79배；외원성miR-373화anti-miR-373분별하조QBC939세포、상조인간내담관상피세포(HIBEpic)중MBD2표체2.56배(P＜0.01)화2.13배(P＜0.01).결론 miR-373통과결합재MBD2-3' UTR,억제MBD2표체.간문부담관암중,miR-373하조도치MBD2표체증고,여종류진전화예후밀절상관.
Objective To study the expression and targeting to methyl-CpG-binding domain 2 (MBD2) of microRNA (miR)-373 in hilar cholangiocarcinoma.Methods The miR-373 expression was detected by using TaqMan miRNA assay.The mRNA and protein expression of methyl-CpG binding domain proteins (MBPs) was examined by using QuantiTect_Primer assays and Western blotting,respectively.The targeting at MBD2-3' UTR by miR-373 was evaluated by using dual-luciferase reporter gene assay.Results The miR-373 expression was decreased 2.94 times and was closely associated with poor cell differentiation (P ＜ 0.05),advanced clinical stage (P ＜ 0.05),and shorter survival (P ＜ 0.05) in hilar cholangiocarcinoma.In high miR-373 group,the MBD2 mRNA and protein expression was increased 3.75 times (P ＜0.01) and 2.34 times (P ＜ 0.05),respectively.Precursor miR-373 inhibited the luciferase activity of MBD2-3' UTR by 1.79 times.Exogenous miR-373 and anti-miR-373 suppressed MBD2 in QBC939 cells by 2.56 times (P ＜ 0.01),and enhanced MBD2 in human intrahepatic biliary epithelial cells(HIBEpic) by 2.13 times (P ＜ 0.01).Conclusion miR-373 is one negative regulator of MBD2.The increase of MBD2 induced by down-expression of miR-373 plays an important role in development and prognosis of hilar cholangiocarcinoma.