miR-590靶向SIP1抑制胆管癌细胞上皮间充质转化的实验研究  

miR-590 targeting SIP1 inhibits epithelial-mesenchymal transition in cholangiocarcinoma cells

作　　者：张梅 崔新厂 刘烨 ZHANG Mei;CUI Xinchang;LIU Ye(Quality Control Department,Dongying Second People s Hospital,Dongying 257335,China)

机构地区：[1]东营市第二人民医院质控科,山东东营257335 [2]东营市第二人民医院肿瘤科,257335 [3]清华大学药学院,100084

出　　处：《临床肿瘤学杂志》2018年第9期785-789,共5页Chinese Clinical Oncology

摘　　要：目的探讨mi R-590在胆管癌细胞上皮间充质转化(EMT)中的作用及可能机制。方法采用荧光定量PCR(QPCR)检测人正常肝内胆管上皮细胞系HIBEC以及人胆管癌细胞系HUCCT1、HCCC-9810、RBE中mi R-590的表达情况。双荧光素酶报告试验评价mi R-590对SIP1的靶向调控作用。向HUCCT1细胞转染mi R-590 mimics(转染组)和无义核苷酸序列(NC组),Western blotting检测两组EMT相关蛋白的表达。向HUCCT1细胞转染SIP1 si RNA,Western blotting检测干扰SIP1后EMT相关蛋白的表达。结果 HUCCT1、HCCC-9810、RBE细胞系中mi R-590水平分别为0. 37±0. 084、0. 31±0. 071和0. 53±0. 089,显著低于人正常肝内胆管上皮细胞系HIBEC的1. 12±0. 201,差异具有统计学意义(P<0. 05)。mi R-590可抑制野生型SIP1 3’UTR报告基因载体的荧光素酶活性,而对突变型SIP1 3’UTR-MUT的荧光素酶活性无影响。转染mi R-590mimics可以下调HUCCT1细胞中Vimentin、N-cadherin、ZEB1、ETS1、SNAIL1及TWIST1蛋白表达,上调E-cadherin蛋白表达。SIP1沉默能上调上皮标志物E-cadherin蛋白表达,并下调间充质蛋白Vimentin和N-cadherin蛋白表达。结论 mi R-590通过靶向SIP13’-UTR阻断其翻译,最终抑制胆管癌细胞上皮间充质转化。Objective To investigate the role and possible mechanism of miR-590 in epithelial-mesenchymal transition （EMT） of cholangiocarcinoma cells.Methods Fluorescence quantitative PCR （QPCR） was used to detect the expression of miR-590 in human normal intrahepatic bile duct epithelial cell line HIBEC, human cholangiocarcinoma cell line HUCCT1, HCCC-9810 and RBE. Dual luciferase reporter assay was used to evaluate the regulatory effect of miR-590 on SIP1. MiR-590 mimics （transfected group） and nonsense nucleic aicd sequence （NC group） were transfected into HUCCT1 cells, and Western blotting was used to detect the expression of EMT related protein in two groups. SIP1 siRNA was transfected into HUCCT1 cells and Western blotting was used to detect EMT related protein expression after interfering SIP1.Results The expression levels of miR-590 in HUCCT1, HCCC-9810 and RBE cell lines were 0.37±0.084,0.31±0.071 and 0.53±0.089, which was significantly lower than that of normal intrahepatic bile duct epithelial cell line HIBEC （1.12±0.201）, and the difference was statistically significant （ P 〈0.05）. MiR-590 mimics significantly reduced luciferase expression in the psiCHECK2-SIP1-3’UTR group （0.37±0.041, P 〈0.001）,while not affecting the expression of the psiCHECK2-SIP1-3’UTR-MUT group （1.21±0.211, P 〉0.05）. Overexpressed of miR-590 could down-regulated the mesenchymal proteins Vimentin, N-cadherin, ZEB1, ETS1, SNAIL1, TWIST1 and up-regulated the epithelial marker E-cadherin. SIP1 silencing could up-regulate the expression of E-cadherin protein and down-regulate the expression of Vimentin and N-cadherin protein.Conclusion MiR-590 blocked its translation by targeting SIP1-3’UTR and ultimately inhibited epithelial mesenchymal transition of cholangiocarcinoma cells.

关 键 词：胆管癌 miR-590 SIP1 上皮间充质转化 

分 类 号：R739.41[医药卫生—肿瘤]