MiR-181a靶向RASSF1A促进骨肉瘤细胞的生长和转移

目的：探讨miR-181a是否靶向调控抑癌基因RAS相关区域家族1A(Ras-association domain family 1，isoform A，RASSF1A)，从而促进骨肉瘤细胞的生长和转移。方法：采用real-time PCR检测30例人骨肉瘤组织及其癌旁正常骨组织中miR-181a的表达，并分析miR-181a表达水平与骨肉瘤患者临床病理特征的相关性。在骨肉瘤细胞MG-63中瞬时转染miR-181a模拟物和miR-181a抑制剂，分别采用MTT和transwell检测miR-181a对骨肉瘤细胞生长和转移的作用。随后，预测并通过双荧光素酶报告基因验证miR-181a与RASSF1A基因3'-非编码区(3'-untranslated region，3'-UTR)的特异性结合作用。结果：与癌旁正常骨组织比较，miR-181a在骨肉瘤组织中高表达(P<0.001)，但其表达水平与骨肉瘤患者的性别、年龄、肿瘤大小及肿瘤病理分期等无明显相关性(P>0.05)。MTT及transwell结果显示过表达miR-181a促进了MG-63细胞生长、迁移和侵袭，显著高于阴性对照组(P<0.05)；而抑制miR-181a的表达降低了MG-63细胞生长、迁移和侵袭，显著低于阴性对照组(P<0.05)。双荧光素酶报告基因检测结果显示miR-181a可靶向结合RASSF1A基因3'-UTR，从而调控RASSF1A的表达。结论：miR-181a可特异性结合RASSF1A基因3'-UTR，下调RASSF1A基因的表达，从而促进人骨肉瘤细胞MG-63的生长和转移。

miR-181a promotes the proliferation and metastasis of osteosarcoma cells by targeting RASSF1A

Objective: To investigate the role of miR-181a in promoting the proliferation and metastasis of osteosarcoma cells by targeting RASSF1A. Methods: The level of miR-181a in 30 human osteosarcoma tissues and corresponding bone tissues was detected by real-time PCR, and the correlation between the level of miR-181a and clinicopathological characteristics of osteosarcoma was analyzed. Osteosarcoma cells MG-63 were transfected with chemically-synthesized miR-181a mimics and inhibitors, and the proliferation, migration and invasion of MG-63 cells were detected by MTT and Transwell assay. The specific binding ability of miR-181a to RASSF1A 3'-UTR was theoretically predicted and detected by the dual luciferase reporter gene assay. Results: The level of miR-181a in osteosarcoma tissues was statistically higher than that in the corresponding bone tissues (P<0.001). However, the level of miR-181a was not correlated with gender, age, tumor size and stage (P>0.05). MTT and Transwell assays showed that the growth rate, migration and invasion ability of MG-63 cells with up-regulation of miR-181a was significantly increased compared with negative control (P<0.05), while the growth rate, migration and invasion ability of MG-63 with down-regulated miR-181a was significantly decreased compared with negative control (P<0.05). Luciferase reporter gene assay showed that miR-181a targeted the 3'-UTR of RASSF1A and regulated the expression of RASSF1A. Conclusion: MiR-181a promotes the proliferation and metastasis of osteosarcoma cells through specifically binding to RASSF1A 3'-UTR and subsequent down-regulation of RASSF1A