微小RNA-3677靶向调控B细胞易位基因1表达及对肝细胞癌细胞侵袭迁移的影响  

Targeted regulation of microRNA-3677 on BTG1 expression and its effect on invasion and migration of hepatocellular carcinoma cells

作　　者：魏世平[1] 尹晓燕[1,2] 田新利[1,2] 李宏勇[1,2] 李娜[1,2] WEI Shiping;YIN Xiaoyan;TIAN Xinli;LI Hongyong;LI Na(Department of Pathology,the Second Affiliated Hospital of Xingtai Medical College,Xingtai 054000,China)

机构地区：[1]邢台医学高等专科学校附属二院病理科,河北邢台054000 [2]邢台医学高等专科学校基础医学部病理教研室,054000

出　　处：《临床肿瘤学杂志》2019年第8期690-694,共5页Chinese Clinical Oncology

摘　　要：目的探讨微小RNA-3677(miR-3677)对B细胞易位基因1(BTG1)的靶向调控作用及肝细胞癌(HCC)细胞HepG2侵袭和迁移的影响。方法采用脂质体Lipofectamine 2000向HepG2细胞转染miR-3677抑制剂(Inhibitor组)和阴性对照序列(NC组)并以未行转染的细胞为对照组,采用实时定量PCR(QPCR)检测miR-3677水平,划痕实验和Transwell实验分别检测划痕宽度和穿膜细胞数目,双荧光素酶报告基因实验验证miR-3677与BTG1的靶向关系,QPCR检测BTG1、基质金属蛋白酶-9(MMP-9)和信号转导和转录激活因子3(STAT3)的mRNA水平,Western blotting检测BTG1、MMP-9、STAT3和磷酸化信号转导和转录激活因子3(p-STAT3)的蛋白水平。结果QPCR结果显示Inhibitor组的miR-3677水平为0.241±0.059,低于对照组的1.012±0.123和NC组的1.147±0.168(P<0.05);Inhibitor组的穿膜细胞数和划痕相对宽度分别为(96.5±9.2)个和(62.15±3.29)%,均优于对照组的(174.5±12.9)个和(37.45±2.16)%及NC组的(181.6±16.4)个和(39.47±2.82)%,差异有统计学意义(P<0.05)。miR-3677模拟物抑制了BTG1野生型3′端非翻译区的荧光素酶活性,而对突变型的无影响。与其余两组相比,Inhibitor组的BTG1表达水平升高,而MMP9和p-STAT3的水平降低(P<0.05)。结论下调miR-3677水平可抑制HCC细胞的迁移侵袭,该miRNA发挥癌基因的作用,可能与通过靶向BTG1以激活JAK/STAT3信号传导有关,为HCC提供一个新的潜在治疗靶点。Objective To investigate the the targeted regulation of microRNA-3677(miR-3677)on B cell translocation gene 1(BTG1)and invasion and migration of hepatocellular carcinoma(HCC)HepG2 cells.Methods The HepG2 cells were transfected with miR-3677 inhibitor(Inhibitor group)and negative control(NC group)by the liposome Lipofectamine 2000,and untransfected cells were used as Control group.Real-time quantitative polymerase chain reaction(QPCR)was used to detect the level of miR-3677.The scratch width and number of transmembrane cells were detected by cell scratch test and Transwell test,respectively.Double luciferase reporter gene assay was used to verify the targeting relationship between miR-3677 and BTG1.The mRNA levels of BTG1,matrix metalloproteinase-9(MMP-9)and signal transducer and activator of transcription 3(STAT3)were detected by QPCR.The protein levels of BTG1,MMP-9,STAT3 and phospho-STAT3(p-STAT3)were detected by Western blotting.Results QPCR showed that the miR-3677 level in Inhibitor group was 0.241±0.059,lower than 1.012±0.123 in Control group and 1.147±0.168 in NC group(P<0.05).The number of transmembrane cells and relative scratch width in Inhibitor group were 96.5±9.2 and(62.15±3.29)%,superior to 174.5±12.9 and(37.45±2.16)%in Control group and 181.6±16.4 and(39.47±2.82)%in NC group(P<0.05).MiR-3677 mimics significantly inhibited the relative luciferase activity of wild-type 3′untranslated region of BTG1,but had no effect on mutant BTG1 luciferase activity.Compared with other two groups,the increased levels of BTG1 and decreased levels of MMP-9 and p-STAT3 were observed in Inhibitor group(P<0.05).Conclusion Downregulation of miR-3677 can inhibit the migration and invasion of HCC cells.This miRNA plays an oncogene role,which may be related to the activation of JAK/STAT3 signal transduction by targeting BTG1,thus providing a new potential therapeutic target for HCC.

关 键 词：肝细胞癌 微小RNA-3677 侵袭迁移 B细胞易位基因1 

分 类 号：R735.7[医药卫生—肿瘤]