摘要
目的研究微小RNA-134（miRNA-134）对人脑胶质瘤细胞系U87在侵袭、迁移方面的作用并探讨可能的作用机制。方法通过人工合成miR-134 mimic瞬时转染脑胶质瘤细胞系U87,实时定量荧光聚合酶链式反应（PCR）检测细胞miR-134的表达水平;并通过四唑盐比色法（MTT）检测U87细胞增殖情况;Transwell小室法检测各组U87细胞株迁移和侵袭能力;实时定量荧光PCR及蛋白质印迹法（Western blot）检测各组基质金属蛋白酶3（MMP-3）的表达水平。统计学数据处理釆用SPSS 19.0软件处理,组间比较采用单因素方差分析,P〈0.05有统计学意思。结果荧光显微镜下观察转染效率90%以上,实时定量荧光PCR结果显示与未处理的空白对照组（control）及空白转染组（Mock）比较,miR-134转染组的miR-134表达水平明显上调,可达到对照组的5-6倍（P〈0.05）;MTT实验结果表明miR-134转染组细胞增殖能力明显下降（P〈0.05）,同时Transwell实验也显示miR-134转染组细胞的迁移能力明显降低（P〈0.05）;实时定量荧光PCR及Western blot分析MMP-3表达情况发现,过表达miR-134可以明显减少MMP-3基因及蛋白的表达水平（P〈0.05）。结论本研究表明miR-134能够抑制胶质瘤U87细胞的增殖及侵袭迁移能力,其机制可能与抑制MMP-3的表达有关,提示miR-134可能成为胶质瘤治疗的新靶点。
Objective The effect of micmRNA-D4 on biological behaviors of human glioma cell line U87 by observing the changes of cell proliferation and the invasion is discussed and the possible mechanism is explored, Methods Synthetic miR-134 mimic was transiently transfected into glioma cell lines U87, and then the expression levels of miR- 134 were tested by real-time polymerase chain reaction （PCR） ; and U87 cell proliferation was detected by Mosmann tetrazolium test （MTT） assay; Transwell chamber assay was used to detect lines migration and invasion in U87 cells of each group; real-time PCR and Western blot were used to detect the expression of matrix metalloproteinase 3 （MMP-3） expression. Statistical data processing precluded by SPSS 19. 0 software. The groups were compared using one-way ANOVA, with P 〈0. 05 as statistical significance. Results Transfection efficiency observed under a fluorescence microscope was more than 90%. Real-time PCR results showed that miR-134 expression in transfected group was significantly up-regulated and could reach 5 to 6 times compared with the untreated control group （control） and blank transfection group （Mock） （P 〈 0. 05） ; MTr results showed that cell proliferation was significantly decreased in miR-134 transfected group （P 〈0. 05）, while Transwell experiments also showed that the migration of cells was significantly lower in miR-134 transfected group （P 〈 0. 05） ; real-time PCR and Western blot analysis found that over-expression of miR-134 could significantly reduce the levels of MMP-3 gene and protein （ P 〈0. 05）. Conclusion This study shows that miR-134 can inhibit the proliferation and invasion of U87 glioma cell migration, which may be related to the inhibition of MMP-3 expression, suggesting that miR-134 may become a new target for the treatment of gliomas.
出处
《中华神经外科疾病研究杂志》
CAS
2015年第2期105-108,共4页
Chinese Journal of Neurosurgical Disease Research
