miR-136对胶质瘤U251细胞生物学功能的影响

Effect of miR-136 on the biological functions of glioma cell line U251

目的:初步探讨miR-136对胶质瘤U251细胞增殖、侵袭和迁移的影响。方法:选取胶质瘤U251细胞系,分别转染miR-136 mimic(模拟物组)和miR-136模拟物阴性对照(对照组),采用qRT-PCR法检测转染效率,观察2组细胞miR-136表达差异;显微镜下观察转染24 h后细胞密度,并拍照记录;CCK-8法检测转染后细胞增殖活性;Transwell和划痕实验分别检测转染后细胞侵袭及迁移能力;检索miRnada生物信息学数据库,预测并分析miR-136下游靶基因;qRT-PCR和蛋白质印迹法分别检测细胞中FZD4 mRNA和FZD4蛋白的表达。结果:与对照组相比,模拟物组细胞miR-136表达水平明显升高(P<0.05);转染24 h之后,模拟物组细胞密度低于对照组;CCK-8检测显示模拟物组细胞增殖活性较对照组明显降低(P<0.05);Transwell和划痕实验显示模拟物组细胞侵袭和迁移能力较对照组降低;miRnada数据库分析显示FZD4是miR-136的下游靶基因;与对照组相比,模拟物组FZD4 mRNA和FZD4蛋白表达水平均明显降低(P均<0.05)。结论:miR-136可能通过靶向调控FZD4基因的表达抑制胶质瘤U251细胞的增殖、侵袭和转移。

To investigate the effect of miR-136on the biological functions of glioma U251cells.Methods:Glioma cell line U251selected in the study were divided into two groups,miR-136mimic(mimic group)and miR-136mimic negative control(control group)was transfected respectively.qRT-PCR was used to detect the expression of miR-136in the two groups cells;24h after transfection,the cell density of the two groups was observed under microscope;CCK-8assay was applied to evaluate the proliferative ability;the invasion and migration of glioma U251cells was evaluated by Transwell assay and scratch wound assay;the binding sites of miR-136were predicted by miRanda;the expression of FZD4mRNA and FZD4protein was detected by qRT-PCR and Western blotting,respectively.Results:The expression of miR-136in mimic group was significantly higher than control group(P<0.05);24h after transfection,the cells of mimic group were less than the control group;CCK-8showed that the proliferative activity of U251cells in mimic group was significantly lower than that of the control group(P<0.05);Transwell assays and scratch wound assays revealed that the invasion and migration of U251cells in mimic group were lower than the control group;according to the miRanda,we found that FZD4was a directly target gene of miR-136;the expression of FZD4mRNA and the expression of FZD4protein in mimic group were significantly lower than the control group(both P<0.05).Conclusion:miR-136might inhibit the proliferation,invasion and migration of U251cells by regulating FZD4.