miR-101-3p靶向TGFβR1/Smad抑制视网膜母细胞瘤细胞侵袭和迁移  

miR-101-3p inhibits invasion and migration of retinoblastoma cells by targeting TGFβR1/Smad

作　　者：王爽[1] 段素芳[1] 牛秉轩 闫琼 李晓鹏[1] WANG Shuang;DUAN Su-Fang;NIU Bing-Xuan;YAN Qiong;LI Xiao-Peng

机构地区：[1]新乡医学院第三附属医院眼科

出　　处：《眼科新进展》2019年第10期924-928,共5页Recent Advances in Ophthalmology

摘　　要：目的探索miR-101-3p对视网膜母细胞瘤细胞侵袭和迁移的影响及作用机制。方法RT-PCR检测miR-101-3p及转化生长因子β受体1(transforming growth factor beta receptor 1,TGFβR1)表达水平;生物信息学预测miR-101-3p与TGFβR1的靶向关系并通过荧光素酶报告实验进行验证。将Y79细胞分为Control组、miR-101-3p mimic组(转染miR-101-3p mimic)、pcDNA-TGFβR1组(转染pc-TGFβR1)及miR-101-3p mimic+pc-TGFβR1组(共转染miR-101-3p mimic和pc-TGFβR1),MTT检测细胞增殖速率,流式细胞术检测细胞凋亡,Transwell实验检测细胞侵袭,划痕实验检测细胞迁移,Western blot检测TGFβR1、Ki67、Bcl-2、Bax、cleaved Caspase-9、基质金属蛋白酶(matrix metalloproteinase,MMP)-2、MMP-9、血管内皮生长因子、p-Smad2及p-Smad3的表达水平。结果在视网膜母细胞瘤组织中,miR-101-3p的表达为0.35±0.05,明显低于正常视网膜组织(P<0.01);视网膜母细胞瘤细胞株Y79中miR-101-3p的表达为0.24±0.04,明显低于视网膜上皮细胞株ARPE-19(P<0.01);与Control组相比,miR-101-3p mimic组miR-101-3p mRNA水平显著升高、TGFβR1 mRNA水平显著降低(均为P<0.01);miR-101-3p与TGFβR1之间存在连续结合片段。与Control组相比,miR-101-3p mimic组TGFβR1的表达水平显著降低,增殖速率及细胞中Ki67表达均显著降低,细胞凋亡率显著增加,Bax及cleaved Caspase-9表达显著升高、Bcl-2表达显著降低,侵袭细胞数目和划痕闭合率显著降低,MMP-2、MMP-9、血管内皮生长因子、TGFβR1、p-Smad2和p-Smad3的表达水平显著降低(均为P<0.01)。结论miR-101-3p通过靶向下调TGFβR1抑制视网膜母细胞瘤细胞的侵袭和迁移。Objective To explore the effect of miR-101-3p on invasion and migration of retinoblastoma cells and its mechanism.Methods The expression levels of miR-101-3p and transforming growth factor beta receptor 1(TGFβR1)were detected by RT-PCR.The targeting relationship between miR-101-3p and TGFβR1 was predicted by bioinformatics and verified by luciferase reporter assay.Y79 cells were divided into Control group,miR-101-3p mimic group(cells were transfected with miR-101-3p mimic),pcDNA-TGFβR1 group(cells were transfected with pc-TGFβR1)and miR-101-3p mimic+pc-TGFβR1 group(cells were co-transfected with miR-101-3p mimic and pc-TGFβR1).Cell proliferation was detected by MTT,apoptosis was detected by flow cytometry,cell invasion was detected by Transwell,cell migration was detected by scratch test,and the expression levels of TGFβR1,Ki67,Bcl-2,Bax,Caspase-9,matrix metalloproteinase 2(MMP-2),MMP-9,vascular endothelial growth factor(VEGF),p-Smad2 and p-Smad3 were measured by Western blot.Results The expression of miR-101-3p in retinoblastoma tissue was 0.35±0.05,which was significantly lower than that of normal retinal tissue(P<0.01).The expression of miR-101-3p in retinoblastoma cell line Y79 was 0.24±0.04,which was significantly lower than that of retinal epithelial cell line ARPE-19(P<0.01).Compared with the Control group,the miR-101-3p mRNA level of the miR-101-3p mimic group was significantly increased,and the level of TGFβR1 mRNA was significantly decreased(both P<0.01).There was a continuous binding fragment between miR-101-3p and TGFβR1.Compared with the Control group,the expression level of TGFβR1 in the miR-101-3p mimic group was significantly decreased,the proliferation rate and the expression of Ki67 were significantly decreased,the apoptosis rate was significantly increased,the expression of Bax and cleaved Caspase-9 was significantly increased,the expression of Bcl-2 was significantly decreased,the number of invasive cells and the rate of scratch closure were significantly decreased,and the expressi

关 键 词：miR-101-3p TGFβR1 视网膜母细胞瘤 细胞侵袭 细胞迁移 

分 类 号：R774[医药卫生—眼科]