Hu Y, Liang D, Chen X, Chen L, Bai J, Li H, Yin C, Zhong W
Department of Pathology, Weifang Medical University, Weifang 261053, China.
First Department of Joint Surgery, Weifang Medical University, Weifang 261053, China.
Nan Fang Yi Ke Da Xue Xue Bao. 2021 Oct 20;41(10):1562-1568. doi: 10.12122/j.issn.1673-4254.2021.10.16.
To explore the role of miR-671-5p in regulating the migration and invasion of osteosarcoma and the underlying mechanisms.
The differentially expressed microRNAs (miRNAs) in osteosarcoma were screened in the NCBI online database, and the target proteins of these miRNAs were predicted and their functions were analyzed. Osteosarcoma cells were transfected with a plasmid overexpressing miR-671-5p, and the transfection efficiency was assessed using quantitative real-time PCR (qRT-PCR). The changes in the migration and invasion of the transfected cells were examined with Transwell assay, and the expressions of proteins related with epithelial-mesenchymal transition (EMT) were detected using Western blotting. Dual-luciferase reporter assay was performed to determine whether the 3'UTR of SMAD3 contained a targeted binding site of miR-671-5p.
MiR-671-5p was significantly down-regulated in both osteosarcoma tissues and osteosarcoma cells ( < 0.05). The osteosarcoma cells overexpressing miR-671-5p showed significantly reduced migration and invasion abilities ( < 0.05) with obviously lowered expressions of EMT-related proteins ( < 0.05). SMAD3 was highly expressed in osteosarcoma cells ( < 0.05), and dual-luciferase reporter assay confirmed the presence of a targeted binding site between miR-671-5p and the 3'UTR of SMAD3 ( < 0.05). In osteosarcoma cells transfected with a SMAD3-overexpressing plasmid ( < 0.05), the high expression of SMAD3 significantly inhibited by miR-671-5p overexpression ( < 0.05). Transwell assay demonstrated that SMAD3 overexpression significantly promoted the migration and invasion of osteosarcoma cells ( < 0.05), and while miR-671-5p overexpression obviously reversed this effect ( < 0.05).
MiR-671-5p can inhibit the invasion and migration of osteosarcoma cells by negatively regulating SMAD3.
探讨miR-671-5p在调控骨肉瘤迁移和侵袭中的作用及其潜在机制。
在NCBI在线数据库中筛选骨肉瘤中差异表达的微小RNA（miRNA），预测这些miRNA的靶蛋白并分析其功能。用过表达miR-671-5p的质粒转染骨肉瘤细胞，采用定量实时PCR（qRT-PCR）评估转染效率。用Transwell实验检测转染细胞迁移和侵袭能力的变化，用蛋白质印迹法检测上皮-间质转化（EMT）相关蛋白的表达。进行双荧光素酶报告基因实验以确定SMAD3的3'UTR是否包含miR-671-5p的靶向结合位点。
miR-671-5p在骨肉瘤组织和骨肉瘤细胞中均显著下调（<0.05）。过表达miR-671-5p的骨肉瘤细胞迁移和侵袭能力显著降低（<0.05），EMT相关蛋白表达明显降低（<0.05）。SMAD3在骨肉瘤细胞中高表达（<0.05），双荧光素酶报告基因实验证实miR-671-5p与SMAD3的3'UTR之间存在靶向结合位点（<0.05）。在转染了过表达SMAD3质粒的骨肉瘤细胞中（<0.05），miR-671-5p过表达显著抑制了SMAD3的高表达（<0.05）。Transwell实验表明，SMAD3过表达显著促进骨肉瘤细胞的迁移和侵袭（<0.05），而miR-671-5p过表达明显逆转了这种作用（<0.05）。
miR-671-
