[1]王爽,段素芳,牛秉轩,等.miR-101-3p靶向TGFβR1/Smad抑制视网膜母细胞瘤细胞侵袭和迁移[J].眼科新进展,2019,39(10):924-928.[doi:10.13389/j.cnki.rao.2019.0210]
　WANG Shuang,DUAN Su-Fang,NIU Bing-Xuan,et al.miR-101-3p inhibits invasion and migration of retinoblastoma cells by targeting TGFβR1/Smad[J].Recent Advances in Ophthalmology,2019,39(10):924-928.[doi:10.13389/j.cnki.rao.2019.0210]
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miR-101-3p靶向TGFβR1/Smad抑制视网膜母细胞瘤细胞侵袭和迁移/HTML
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《眼科新进展》[ISSN:1003-5141/CN:41-1105/R]

卷:
39卷
期数:
2019年10期
页码:
924-928
栏目:
实验研究
出版日期:
2019-10-05

文章信息/Info

Title:
miR-101-3p inhibits invasion and migration of retinoblastoma cells by targeting TGFβR1/Smad
作者:
王爽; 段素芳; 牛秉轩; 闫琼; 李晓鹏
453000　河南省新乡市，新乡医学院第三附属医院眼科
Author(s):
WANG Shuang; DUAN Su-Fang; NIU Bing-Xuan; YAN Qiong; LI Xiao-Peng
Department of Ophthalmology，the Third Affiliated Hospital of Xinxiang Medical University,Xinxiang 453000,Henan Province,China
关键词:
miR-101-3p; TGFβR1; 视网膜母细胞瘤; 细胞侵袭; 细胞迁移
Keywords:
miR-101-3p; transforming growth factor beta receptor 1; retinoblastoma; cell invasion; cell migration
分类号:
R774
DOI:
10.13389/j.cnki.rao.2019.0210
文献标志码:
A
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。
Abstract:
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 expression levels of MMP-2,MMP-9,VEGF,TGFβR1,p-Smad2 and p-Smad3 were significantly decreased (all P<0.01).Conclusion　miR-101-3p inhibits the invasion and migration of retinoblastoma cells by targeting down-regulation of TGFβR1.

参考文献/References:

［1］　GENG W,QIN F,REN J X,XU X H,WANG A Y.Effects of ribosomal protein L41 on proliferation and apoptosis of human retinoblastoma Y79 cells and its mechanism［J］.Rec Adv Ophthalmol,2018,38(3):214-217.
耿雯,秦丰,任佳绪,徐晓鸿，王爱媛.核糖体蛋白L41对人视网膜母细胞瘤Y79细胞增殖和凋亡的影响及其机制［J］.眼科新进展,2018,38(3):214-217.
［2］　MANJANDAVIDA F P,HONAVAR S G,REDDY V A,KHANNA R.Management and outcome of retinoblastoma with vitreous seeds［J］.Ophthalmology,2014,121(12):517-524.
［3］　SHIELDS C L,FULCO E M,ARIAS J D,ALARCON C,PELLEGRINI M,RISHI P,et al.Retinoblastoma frontiers with intravenous,intra-arterial,periocular,and intravitreal chemotherapy［J］.Eye (Lond),2013,27(2):253-264.
［4］　ZHANG X,HE X,LIU Y,ZHANG H Q,CHEN H,GUO S X,et al.MiR-101-3p inhibits the growth and metastasis of non-small cell lung cancer through blocking PI3K/AKT signal pathway by targeting MALAT-1［J］.Biomed Pharmacother,2017,93(12):1065-1073.
［5］　LEI Q,SHEN F,WU J,ZHANG W,WANG J,ZHANG L.MiR-101,downregulated in retinoblastoma,functions as a tumor suppressor in human retinoblastoma cells by targeting EZH2［J］.Oncol Rep,2014,32(1):261-269.
［6］　SHIELDS C L,ALSET A E,SAY E A,CAYWOOD E,JABBOUR P,SHIELDS J A.Retinoblastoma control with primary intra-arterial chemotherapy:Outcomes before and during the intravitreal chemotherapy era［J］.J Pediatr Ophthalmol Strabismus,2016,53(5):275-284.
［7］　GORE A J,DEITZ S L,PALAM L R,CRAVEN K E,KORC M.Pancreatic cancer-associated retinoblastoma 1 dysfunction enables TGF-beta to promote proliferation［J］.J Clin Invest,2014,124(1):338-352.
［8］　ZHANG L,CHENG X,GAO Y,ZHANG C,BAO J,GUAN H.Curcumin inhibits metastasis in human papillary thyroid carcinoma BCPAP cells via down-regulation of the TGF-beta/Smad2/3 signaling pathway［J］.Exp Cell Res,2016,341(2):157-165.
［9］　YANG J,LU Y,LIN Y Y,ZHENG Z Y,FANG J H,HE S,et al.Vascular mimicry formation is promoted by paracrine TGF-beta and SDF1 of cancer-associated fibroblasts and inhibited by miR-101 in hepatocellular carcinoma［J］.Cancer Lett,2016,383(1):18-27.
［10］　HOU Y,LI L,JU Y,LU Y,CHANG L,XIANG X.miR-101-3p regulates the viability of lung squamous carcinoma cells via targeting EZH2［J］.J Cell Biochem,2017,118(10):3142-3149.
［11］　MA Y,LUO W,BUNCH B L,PRATT R N,TRUMP D L,JOHNSON C S.1,25D3 differentially suppresses bladder cancer cell migration and invasion through the induction of miR-101-3p［J］.Oncotarget,2017,8(36):60080-60093.
［12］　BAUVOIS B.New facets of matrix metalloproteinases MMP-2 and MMP-9 as cell surface transducers:outside-in signaling and relationship to tumor progression［J］.Biochim Biophys Acta,2012,18(25):29-36.
［13］　BHATTACHARYA R,FAN F,WANG R,YE X,XIA L,BOULBES D,et al.Intracrine VEGF signalling mediates colorectal cancer cell migration and invasion［J］.Br J Cancer,2017,117(6):848-855.

更新日期/Last Update: 2019-10-12