摘要
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。
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.
出处
《中南大学学报（医学版）》
CAS
CSCD
北大核心
2016年第8期789-795,共7页
Journal of Central South University ：Medical Science
基金
湖南省自然科学基金(14JJ7019)~~
