miR-369靶向调节SOX4表达对骨肉瘤细胞增殖和凋亡的调控作用

目的:观察微小RNA（miRNA,miR）-369通过调节SOX4对骨肉瘤MG-63细胞增殖、凋亡的影响。方法:miR-369体外模拟物（mimics）转染，构建miR-369上调表达模型，以转染阴性对照链为对照组，CCK-8法和流式细胞技术分别检测细胞增殖率和凋亡率变化；Real-time PCR法检测SOX4 mRNA表达水平；双荧光素酶活检检测证实miR-369与SOX4相互作用关系；Western-blot法检测不同组间SOX4及Bcl-2家族相关蛋白表达水平变化。结果:miR-369 mimics转染后，MG-63细胞中miR-369表达水平较对照组明显提高(P=0.009)；miR-369 mimic转染后24 h及48 h后细胞相对存活率均明显低于对照组；流式细胞仪结果显示mimic组细胞凋亡率较对照组明显提高（P=0.031）；Real-time PCR和Western blot实验分别证实miR-369 mimic转染后，SOX4在mRNA和蛋白水平表达均明显抑制；而Bcl-2家族相关蛋白表达水平发生明显变化。结论:miR-369表达上调可以抑制骨肉瘤细胞增殖并诱导凋亡，这些作用与靶向下调SOX4表达有关。

miRNA-369 inhibited osteosarcoma proliferation and induced apoptosis by targeting SOX4

Objective:To investigate the effects of microRNA-369 on the proliferation and apoptosis against osteosarcoma MG-63 cell through SOX4.Methods:The expression of miR-369 in osteosarcoma MG-63 cell was detected by Real-time PCR after miR-369mimics transfection.After the validation of miR-369 over-expression,CCK-8 and flow cytometry assay were used to detect the proliferation and apoptosis rate.Then the expression levels of SOX4 mRNA were detected by Real-time PCR.The relationship between the miR-369 and SOX4 was investigated by dual luciferase assay.The expression of SOX4 and Bcl-2 related proteins were detected by Western-blot.Results:After miR-369 mimics transfection in MG-63 cell,relative expression of miR-369 in mimics group was significantly increased compared with the negative-control group(P=0.009).The proliferation at 24 h and 48 h after miR-369 mimics transfection were significantly suppressed compared with the NC group.The flow cytometry assay suggested the apoptosis cell was increased in mimic group compared with the NC group (P=0.031).The expressions of SOX4 in the mRNA and protein levels were reduced significantly in mimics group while the expression of Bcl-2 related proteins were regulated significantly after transfection.Conclusion:miR-369 inhibited osteosarcoma MG-63 cell proliferation and induced apoptosis by targeting SOX4.