miRNA-320a通过靶向水通道蛋白4抑制神经胶质瘤细胞U251迁移侵袭的分析

Study of mi RNA-320a Inhibiting the Migration and Invasion of Glioma Cell Line U251 by Target AQP4

本研究初步探讨过表达miRNA-320a对抑制神经胶质瘤U251细胞迁移、侵袭的可能的机制。实验开始前我们利用生物信息学软件进行分析对比miRNA-320a与水通道蛋白4(AQP4)之间的靶点结合关系,然后我们用miRNA-320a mimic及Ncontrol对U251细胞株进行转染,48 h后进行下一步实验。首先用q-PCR验证过表达转染情况,以及水通道蛋白4(AQP4)m RNA的表达水平,其次用划痕和Transwell检测转染后细胞株的迁移侵袭能力,最后用Western blotting测定AQP4的表达水平。生物信息分析可得miRNA-320a在AQP4 m RNA 3'UTR区域能稳定结合,实验结果显示转染mimic后,过表达组明显升高,且过表达组AQP4 m RNA的表达明显被抑制,划痕和Transwell实验提示了过表达miRNA-320a后能抑制U251细胞株的迁移侵袭能力(p<0.01)。Western blotting结果显示,过表达miRNA-320a与对照组相比能明显抑制AQP4蛋白的表达。所有研究结果提示miRNA-320a能靶向作用AQP4 m RNA 3'UTR区域,并抑制其蛋白表达,从而抑制了肿瘤细胞U251的迁移侵袭能力,为临床治疗恶性胶质瘤提供新的参考。

This study preliminarily disscussed the possible mechanism of overexpression of miRNA-320a in inhibiting the migration and invasion of U251 gliomacell line. Before the experiment, we analyzed the target binding relationship between the miRNA-320a and AQP4 mRNA 3＇ UTR through bioinformatics software, and then we transfected U251 cell line by miRNA-320a mimic and Ncontrol for 48 h for the next step of the experiment. First, the overexpression transfection and the expression of mRNA of AQP4 were detected by q-PCR, and then migration and invasion ability of cell line were identified by scratch test and Transwell, and the expression level of AQP4 was determined by Western blotting at last. Bioinformatics analysis suggested that miRNA-320a could stablely combine in AQP4 mRNA 3＇ UTR region, and the q-PCR results showed that the expression level of overexpression group incerased significantly after transfected by mimic and the expression of AQP4 mRNA was obviously inhibited in overexpression group. The results of scratch test and Transwell indicated overexpression of miRNA-320a could inhibit the migration and invasion ability of U251 cell line. The result of Western blotting illustrated that overexpression ofmiRNA-320a could significantly inhibit the expression of AQP4 protein compared with the control group. In conclusion, miRNA-320a could target AQP4 mRNA 3＇ UTR region and inhibit its expression of protein, which inhibited the migration and invasion ability of glioma cell U251, and it might provide a new reference for clinical treatment of malignant glioma.