摘要
目的探究miR-153-3p与转化生长因子β2(transforming growth factor-beta 2,TGFβ2)的靶向关系以及对体外培养胶质瘤细胞侵袭、迁移和上皮间质转化的影响。方法实时荧光定量PCR和Western blot检测miR-153-3p对SHG-44细胞TGFβ2mRNA和蛋白表达的影响;生物信息学软件预测miR-153-3p与TGFβ2的结合位点;荧光素酶报告实验分析miR-153-3p与TGFβ2的靶向关系;体外侵袭实验和划痕实验检测miR-153-3p对SHG-44细胞侵袭和迁移能力的影响;Western blot检测侵袭、迁移和上皮间质转化相关蛋白的表达。结果miR-153-3p mimic抑制SHG-44细胞中TGFβ2mRNA和蛋白质的表达;软件预测显示miR-153-3p与TGFβ2存在连续的结合区域;miR-153-3p mimic与TGFβ2野生型报告载体共转后,细胞中荧光素酶的活性降低(P <0.01);miR-153-3p mimic与TGFβ2突变型报告载体共转后,细胞中荧光素酶的活性没有明显变化。miR-153-3p mimic转染SHG-44细胞后,侵袭细胞数目减少,划痕闭合率降低,基质金属蛋白酶2(matrix metalloproteinase,MMP-2)、MMP-9、血管内皮生长因子(vascular endothelial growth factor,VEGF)的表达降低,神经钙黏蛋白(N-cadherin)、Snail-2、波形蛋白(Vimentin)和Twist的表达降低,上皮钙黏蛋白(E-cadherin)的表达升高(P<0.01);TGFβ2高表达缓解miR-153-3p mimic对SHG-44细胞侵袭、上皮间质转化相关蛋白的表达。结论 miR-153-3p靶向TGFβ2抑制体外培养胶质瘤细胞侵袭、迁移和上皮间质转化。
Objective To investigate the targeting relationship between microRNA-153-3p and transforming growth factor-beta 2 (TGFβ2) and its effects on invasion, migration and epithelial-mesenchymal transformation of cultured glioma cells.Methods Real-time fluorescence quantitative PCR and Western blotting were used to detect the effects of miR-153-3p on the expressions of TGFβ2 mRNA and protein.Bioinformatics software was used to predict the binding sites of miR-153-3p to TGFβ2.Luciferase reporter assay was used to analyze the targeting relationship between miR-153-3p and TGFβ2.Transwell and wound healing assays were used to detect the effects of miR-153-3p on SHG-44 cells invasion and migration.Western blotting was used to detect the expressions of proteins related to invasion, migration and epithelial-mesenchymal transition.Results miR-153-3p mimic inhibited the expressions of TGFβ2 mRNA and protein in SHG-44 cells.Software predicted results showed a continuous binding region between miR-153-3p and TGFβ2.miR-153-3p mimic co-transfected with TGFβ2 wild-type reporter vector significantly decreased the activity of luciferase in the cells ( P <0.01).miR-153-3p mimic co-transfected with TGFβ2 mutant reporter showed no significant change in luciferase activity.The number of invasive cells and scratch closure rate of SHG-44 cells transfected with miR-153-3p mimic were significantly decreased.The expression levels of MMP-2, MMP-9 and VEGF were significantly decreased.The expression levels of N-cadherin, Snail-2, Vimentin and Twist were significantly decreased, and the expression level of E-cadherin was significantly increased ( P < 0.01 ).The high expression of TGFβ2 alleviated the effect of miR-153-3p on the cell invasion and epithelial mesenchymal transition related proteins in SHG-44.Conclusion miR-153-3p inhibits the invasion and migration of cultured glioma cells and epithelial mesenchymal transition by targeting TGFβ2.
作者
石洪仁
陈礼刚
SHI Hong-ren;CHEN Li-gang(Department of Neurosurgery, The Affiliated Hospital ofSouthwest Medical University, Luzhou 646000, China)
出处
《西安交通大学学报（医学版）》
CAS
CSCD
北大核心
2019年第5期722-727,共6页
Journal of Xi’an Jiaotong University（Medical Sciences）
基金
四川省医学会科研课题(No.S16083)~~
关键词
胶质瘤
转化生长因子Β2
上皮间质转化
上皮钙黏蛋白
glioma
transforming growth factor beta 2
epithelial mesenchymal transition
epithelial cadherin
