微小RNA-150在胆囊癌上皮-间充质转化中的作用及其机制

The relationship between microRNA-150 expression and epithelial-mesenchymal transition in gallbladder cancer and its mechanism

目的:探讨微小RNA(miRNA,miR)-150与胆囊癌肿瘤上皮-间充质转化(EMT)的关系及其作用机制。方法:实时定量聚合酶链反应(qPCR)法检测河北医科大学第二医院2018年1月至2019年5月手术切除的30例胆囊癌肿瘤组织中miR-150的表达,以30例慢性胆囊炎组织为对照;qPCR法同时检测组织中增殖细胞核抗原(PCNA)、E-钙黏蛋白(E-cadherin)、N-钙黏蛋白(N-cadherin)、基质金属蛋白酶(MMP)-9的mRNA表达,分析miR-150与这些基因的关系。应用miR-150 mimics转染人胆囊癌细胞株GBC-SD,划痕实验及Transwell小室实验检测细胞迁移侵袭的改变;qPCR及蛋白质印迹法(Western blot)技术检测各基因mRNA和蛋白的变化。两样本均数比较采用t检验。多样本均数比较时采用方差分析,方差分析后续的两两比较采用最小显著差异法(LSD)。两样本之间相关性比较采用线性相关分析。结果:胆囊癌组织中miR-150和E-cadherin的mRNA表达均低于慢性胆囊炎组织(t=-7.035、-14.146,P<0.01);PCNA、N-cadherin、MMP-9的mRNA与慢性胆囊炎组织比较均明显增强(t=3.813、9.339、5.616,P<0.01)。miR-150表达与患者的肿瘤分化程度、临床分期、淋巴结转移有关(t=3.228、2.396、-2.604,P<0.05)。Pearson相关分析显示,miR-150与E-cadherin呈正相关(r=0.630,P<0.05),与PCNA、N-cadherin、MMP-9表达呈负相关(r=-0.769、-0.628、-0.616,P<0.05)。转染miR-150 mimics后GBC-SD细胞的迁移侵袭能力减弱(F=1965.860、114.940,P<0.01);转染后PCNA、N-cadherin、MMP-9表达降低,而E-cadherin表达增高(P<0.05)。结论:胆囊癌组织中miR-150表达降低是肿瘤进展的因素,其机制可能在于miR-150通过调节一些基因表达参与肿瘤EMT过程。

Objective To analyze the relationship between the expression of microRNA(miRNA,miR)-150 and epithelial-mesenchymal transition(EMT)in gallbladder cancer and its mechanism.Methods The expression of miR-150 in 30 cases of gallbladder cancer was detected by real-time quantitative polymerase chain reaction(qPCR),and 30 cases of chronic cholecystitis tissues was used as control.Simultaneously,the expression of proliferating cell nuclear antigen(PCNA),E-cadherin,N-cadherin and matrix metalloproteinase(MMP)-9 was detected,and the relationship of miR-150 and these genes was analyzed.Human gallbladder cancer cell line GBC-SD was transfected with miR-150 mimics.Then scratch experiments and Transwell chamber experiments were used to detect changes in cell migration and invasion;qPCR and Western blotting were used to detect changes in mRNA and protein of each gene.Results The expressions of miR-150 and E-cadherin in gallbladder cancer tissues were lower than those in chronic cholecystitis tissues(t=-7.035,-14.146,P<0.01).The expressions of PCNA,N-cadherin and MMP-9 were significantly higher compared with chronic cholecystitis tissues(t=3.813,9.339,5.616,P<0.01).The expression of miR-150 was related to the degree of tumor differentiation,clinical stage,and lymph node metastasis(t=3.228,2.396,-2.604,P<0.05).Pearson correlation analysis showed that miR-150 was positively correlated with E-cadherin(r=0.630,P<0.05),and negatively correlated with PCNA,N-cadherin,and MMP-9(r=-0.769,-0.628,-0.616,P<0.05).After transfection,the migration and invasion ability of GBC-SD cells decreased(F=1965.860,114.940,P<0.01).The expressions of PCNA,N-cadherin,and MMP-9 were decreased,while the expression of E-cadherin was increased(P<0.05).Conclusion Decreased miR-150 expression in gallbladder cancer plays a key role in tumor progression,and the mechanism may be that miR-150 participates in tumor EMT process by regulating some genes.