miR-18a-3p在肝癌中的表达及其调控ADCY1表达对肝癌细胞侵袭及增殖的影响  

Expression of miR-18a-3p in liver cancer and the influence of its regulating ADCY1 expression on invasion and proliferation of liver cancer cells

作　　者：彭婀敏 欧阳锡武 王文龙[3] 郑璐 姚磊 白宁 PENG Emin;OUYANG Xiwu;WANG Wenlong;ZHENG Lu;YAO Lei;BAI Ning(Xiangya School of Public Health,Central South University,Changsha 410078,China;International Medical Department,Xiangya Hospital,Central South University,Changsha 410008,China;Department of General Surgery,Xiangya Hospital,Central South University,Changsha 410008,China)

机构地区：[1]中南大学湘雅公共卫生学院,湖南长沙410078 [2]中南大学湘雅医院国际医疗部,湖南长沙410008 [3]中南大学湘雅医院普通外科,湖南长沙410008

出　　处：《中国普通外科杂志》2022年第5期648-657,共10页China Journal of General Surgery

基　　金：湖南省财政厅科教专项基金资助项目[湘发改委(2019)412号]。

摘　　要：背景与目的:研究表明多种micro RNA(miRNA)可能在肝癌的发生发展中发挥重要作用,其作用机制仍值得进一步研究和探讨。因此,本研究从已报道的肝癌差异表达miRNA中进一步筛选关键miRNA,并验证和探讨其作用机制。方法:从已发表的研究中筛选出肝癌组织及肝癌患者血清/血浆中与正常肝组织及正常血清/血浆中共同的差异表达miRNA;用q RT-PCR在正常肝细胞与肝癌细胞中对筛选出的目标miRNA表达情况进行验证;用过表达和抑制的方法观察目标miRNA对肝癌细胞侵袭能力(Transwell实验)与增殖能力(MTT实验)的影响,以及在30例临床标本中检测目标miRNA的表达并通过KM plotter网站分析其对肝癌患者生存的影响;通过miRDB和GEPIA数据库预测和分析目标miRNA的靶基因,并用逆转实验和双荧光素酶报告实验进一步验证。结果:在肝癌组织(vs.正常肝组织)及肝癌患者血清/血浆(vs.正常人血清/血浆)中共同高表达的miRNA有4个(miR-18a-3p、miR-221-3p、miR-222-3p、miR-224-3p),共同低表达的miRNA有2个(miR-26a-3p、miR-125b-3p)。q RT-PCR实验证实,与正常肝细胞比较,miR-18a在肝癌细胞中高表达,miR-26a在肝癌细胞中低表达(均P<0.05)。过表达/抑制miR-18a-3p表达能促进/降低肝癌细胞的侵袭及生长能力(均P<0.05),而过表达/抑制miR-26a-3p对肝癌细胞的侵袭及生长能力影响无不法确定。分析结果显示,ADCY1是miR-18a-3p的靶基因,过表达ADCY1能部分逆转miR-18a-3p对肝癌细胞的上述作用,同时,表达上调的miR-18a-3p能通过结合到ADCY1 m RNA 3’UTR抑制ADCY1的表达。结论:miR-18a-3p可能在肝癌的发生发展中起了关键作用,其在肝癌中表达上调,并能通过抑制下游靶基因ADCY1的表达增强进肝癌细胞的侵袭和增殖能力。Background and Aims:Studies have shown that a variety of micro RNAs(miRNAs)may play crucial roles in the occurrence and development of liver cancer,and their action mechanisms still need to be studied and determined.Therefore,this study was conducted to further screen and validate the key miRNAs from the differentially-expressed miRNAs that have been reported in liver cancer,and then investigate their action mechanisms.Methods:The common differentially-expressed miRNAs in liver cancer tissue and serum/plasm of liver cancer patients versus normal liver tissue and serum/plasm of healthy subjects were screened from the published studies.The expressions of goal miRNAs in normal liver cells and liver cancer cells were verified by q RT-PCR method.Using overexpression and inhibition strategies,the influences of goal miRNAs on invasion ability(Transwell invasion assay)and proliferation capacity(MTT assay)of liver cancer cells were observed,and also the expressions of goal miRNAs in 30 samples of clinical specimens were detected and their impacts on survival of liver cancer patients were analyzed through KM plotter website.The target genes of goal miRNAs were predicted and analyzed using miRDB and GEPIA databases and were further validated by reverse experiment and dual-luciferase reporter assay.Results:There were 4 common up-regulated miRNAs(miR-18 a-3 p,miR-221-3 p,miR-222-3 p,miR-224-3 p)and 2 down-regulated miRNAs(miR-26 a-3 p,miR-125 b-3 p)in liver cancer tissue(vs.normal liver tissue)and serum/plasma of liver cancer patients(vs.serum/plasma of healthy individuals).The q RT-PCR experiment confirmed that miR-18 a was highly expressed and miR-26 a was weakly expressed in liver cancer cells compared with normal liver cells(both P<0.05).Overexpression/inhibition of miR-18 a expression promoted/reduced the invasion and proliferation capacities of liver cancer cells(all P<0.05),while overexpression/inhibition of miR-26 a exerted confused influences on the invasion abilities and proliferation capacities of liver cancer cell

关 键 词：癌 肝细胞 微RNAS 肿瘤侵润 细胞增殖 

分 类 号：R735.7[医药卫生—肿瘤]