微RNA-449b-5p通过调控NOTCH1基因的表达抑制甲状腺乳头状癌细胞增殖、迁移和侵袭  

MiR-449b-5p inhibits proliferation,migration and invasion of thyroid papillary carcinoma cells by regulating NOTCH1 expression

作　　者：王丹[1] 秦珍珠[1] 梁运霞[1] 潘翔珍[1] 张玉文[1] Wang Dan;Qin Zhenzhu;Liang Yunxia;Pan Xiangzhen;Zhang Yuwen(Department of Pathology,Shangqiu First People’s Hospital,Shangqiu,Henan 476000,China)

机构地区：[1]河南省商丘市第一人民医院病理科,476000

出　　处：《中华生物医学工程杂志》2019年第6期698-704,共7页Chinese Journal of Biomedical Engineering

基　　金：河南省科技攻关计划项目(2018020949)。

摘　　要：目的探究miR-449b-5p通过调控NOTCH1基因的表达抑制甲状腺乳头状癌细胞增殖、迁移和侵袭的机制。方法实时荧光定量PCR、免疫印迹检测甲状腺乳头状癌细胞中miR-449b-5p和NOTCH1的表达,过表达miR-449b-5p,沉默或者过表达NOTCH1基因,检测细胞增殖、迁移和侵袭情况;双荧光素酶实验检测miR-449b-5p和NOTCH1的联系。结果与Nthy-ori3-1细胞比较,甲状腺乳头状癌细胞BCPAP、TPC-1、K1中miR-449b-5p表达水平显著降低,NOTCH1mRNA及蛋白水平均显著升高(P<0.05);过表达miR-449b-5p后,与Control组、miR-NC组比较,miR-449b-5p组miR-449b-5p水平显著升高,NOTCH1水平显著降低,OD值、迁移、侵袭细胞数显著降低,CyclinD1、MMP-2蛋白水平显著降低,p21、E-cadherin蛋白水平显著升高(P<0.05);双荧光素酶报告实验显示,免疫印迹证实miR-449b-5p对NOTCH1的影响,miR-449b-5p可与NOTCH1结合负向调控NOTCH1水平;沉默NOTCH1后,与Control组、si-NC组比较,si-NOTCH1组NOTCH1蛋白水平显著降低,OD值、迁移和侵袭细胞数显著减少,CyclinD1、MMP-2蛋白水平显著降低,p21、E-cadherin蛋白水平显著升高(P<0.05);与miR-449b-5p+pcDNA3.1组比较,miR-449b-5p+pcDNA3.1-NOTCH1组NOTCH1水平显著升高,OD值、迁移和侵袭细胞数目显著升高,CyclinD1、MMP-2蛋白水平显著升高,p21、E-cadherin蛋白水平显著降低(P<0.05)。结论过表达miR-449b-5p抑制甲状腺乳头状癌细胞的增殖、迁移和侵袭,过表达NOTCH1基因能逆转过表达miR-449b-5p对甲状腺乳头状癌细胞增殖、迁移和侵袭的抑制作用。Objective To explore the mechanism underlying the inhibitory effect of miR-449b-5p on proliferation,migration and invasion of thyroid papillary carcinoma cells by regulating the expression of NOTCH1 gene.Methods Real-time fluorescence quantitative PCR and Western blotting were used to detect the expression of miR-449b-5p and NOTCH1,cell proliferation,migration and invasion after over-expression of miR-449b-5p and silencing or over-expression of NOTCH1 gene in thyroid papillary carcinoma cells.Dual luciferase reporter assay was used to detect the relationship between miR-449b-5p and NOTCH1.Results Compared with Nthy-ori3-1 cells,the miR-449b-5p expression levels were significantly decreased,and the gene and protein expression levels of NOTCH1 were significantly increased,in thyroid papillary carcinoma BCPAP,TPC-1 and K1 cells(P<0.05).After overexpression of miR-449b-5p,compared with the control group and miR-NC group,the miR-449b-5p group showed significantly higher expression level of miR-449-5p,decreased expression level of NOTCH1,lowered OD value,fewer migrating and invasive cells,down-regulated protein expression of CyclinD1 and MMP-2 and up-regulated protein expression of p21 and E-cadherin(P<0.05).Dual luciferase reporter assay showed that miR-449b-5p may bind to NOTCH1 and thereby negatively regulate its expression.After NOTCH1-silencing,compared with the control group and the si-NC group,the si-NOTCH1 group showed significantly decreased expression levels of NOTCH1 protein,lowered OD value,fewer migrating and invasive cells,down-regulated expression of CyclinD1 and MMP-2 proteins and up-regulated expression of p21 and E-cadherin proteins(P<0.05).Compared with the miR-449b-5p+pcDNA3.1 group,the miR-449b-5p+pcDNA3.1-NOTCH1 group showed significantly expression levels of NOTCH1,higher OD value,more migrating and invasive cells,up-regulated expression of CyclinD1 and MMP-2 proteins,and down-regulated expression of p21 and E-cadherin proteins(P<0.05).Conclusion Overexpression of miR-449b-5p may inhibit prolifer

关 键 词：miR-449b-5p NOTCH1 甲状腺乳头状癌 增殖 迁移 侵袭 

分 类 号：R73[医药卫生—肿瘤]