miR-181b通过靶向调控N-myc下游调节基因2影响骨肉瘤细胞的迁移和侵袭

目的探讨miR-181b对骨肉瘤细胞迁移和侵袭的影响。方法培养骨肉瘤细胞，实时荧光定量PCR检测miR-181b在骨肉瘤细胞中的表达。抑制miR-181b的表达，Transwell检测对骨肉瘤迁移和侵袭的影响；生物信息学分析miR-181b靶基因并采用荧光素酶报告基因分析miR-181b和靶基因在骨肉瘤中是否直接作用，同时采用Western Blotting检测靶基因在骨肉瘤细胞中的表达以及Transwell检测靶基因对骨肉瘤迁移和侵袭的影响。结果实时荧光定量PCR结果表明miR-181b在骨肉瘤细胞中高表达。抑制miR-181b 的表达能够抑制骨肉瘤细胞的迁移和侵袭；生物信息学分析表明N-myc 下游调节基因2（NDRG2）是miR-181b直接作用的靶基因，荧光素酶报告基因分析验证了该结果；Western blotting检测表明NDRG2在骨肉瘤中低表达，而抑制miR-181b 能够显著提高NDRG2 的表达。抑制miR-181b 的表达的同时抑制靶基因NDRG2 的表达，能够逆转抑制miR-181b的表达对骨肉瘤细胞的迁移和侵袭的影响，从而显著提高骨肉瘤细胞的迁移和侵袭。结论miR-181b在骨肉瘤中过表达，NDRG2为miR-181b直接调控靶基因，抑制miR-181b能够提高NDRG2的表达从而抑制骨肉瘤细胞的迁移侵袭。

microRNA-181b promotes migration and invasion of osteosarcoma cells by targeting N-myc downstream regulated gene 2

Objective To investigate the effects of miR-181b on the migration and invasion of osteosarcoma cells. Methods Threecultured osteosarcoma cell lines and MG-63 cells transfected with miR-181b inhibitor were examined for miR-181b expressionusing qRT-PCR analysis. The cell migration and invasion of the transfected cells were assessed with Transwell assay. Thetargets of miR-181b were predicted using a miRNA target prediction software and the results were verified with luciferasereporter assay. The target protein expression in osteosarcoma cells lines was determined by Western blotting, and the cellmigration and invasion changes following inhibition of miR-181b or its target protein were assessed using Transwell assay.Results All the 3 osteosarcoma cells lines showed significantly up-regulated miR-181b expression. Inhibition of miR-181bexpression obviously suppressed the migration and invasion of MG-63 cells. Based on luciferase reporter assay, N-mycdownstream regulated gene 2 (NDRG2) was identified as the direct target gene of miR-181b, and inhibition of NDRG2expression significantly reversed the effect of miR-181b on cell migration and invasion in MG-63 cells. Conclusion miR-181b isover-expressed in osteosarcoma cells, and inhibition of miR-181b, which directly targets NDRG2, can suppress the migrationand invasion of osteosarcoma cells.