lncRNA LINC00173通过调节miR-130a-5p抑制骨肉瘤发生发展的机制研究

Mechanism of lncRNA LINC00173 inhibiting the development of osteosarcoma by regulating miR-130a-5p

目的:探讨长链非编码RNA(lncRNA)LINC00173通过调控miR-130a-5p抑制骨肉瘤发生发展的分子机制。方法:体外培养人骨肉瘤细胞系MG-63,分别将LINC00173过表达载体质粒或空载质粒转染至MG-63细胞,实时荧光定量PCR(qPCR)检测转染效率,采用细胞计数法(CCK-8)和Transwell实验分析MG-63细胞增殖、侵袭和迁移能力,双荧光素酶报告基因实验和qPCR检测LINC00173和miR-130a-5p的靶向调控关系。同时过表达LINC00173和miR-130a-5p,并检测MG-63细胞增殖、侵袭和迁移能力。结果:qPCR检测结果显示,与Mock组比较,pc-LINC00173组LINC00173在MG-63细胞中的表达量明显升高(P<0.01)。转染48 h和72 h后,pc-LINC00173组细胞增殖活力均低于Mock组和pcDNA组(P<0.01)。Transwell检测结果显示,pc-LINC00173组与Mock组比较侵袭和迁移细胞数明显减少(P<0.05)。LINC00173与miR-130a-5p之间存在特异性结合位点,与miR-NC组比较,转染miR-130a-5p mimics能够抑制LINC00173-Wt细胞的相对荧光素酶活性(P<0.01),与Mock组和pcDNA组比较,pc-LINC00173组MG-63细胞中miR-130a-5p的表达量均降低(P<0.01)。与pc-LINC00173组和pc-LINC00173+miR-NC组比较,pc-LINC00173+miR-130a-5p组MG-63细胞中miR-130a-5p的表达明显升高,吸光度值明显升高,侵袭和迁移细胞数明显增多(P<0.01)。结论:LINC00173能够抑制骨肉瘤MG-63细胞增殖、侵袭和迁移,其作用机制可能与靶向抑制miR-130a-5p的表达有关。

Objective:To investigate the molecular mechanism of long-chain non-coding RNA(lncRNA)LINC00173 through regulating miR-130a-5p to mediate the development of osteosarcoma.Methods:Human osteosarcoma cell line MG-63 was cultured in vitro,and LINC00173 overexpression vector plasmid or empty plasmid was transfected into MG-63 cells,real-time fluorescence quantitative PCR(qPCR)was used to detect the transfection efficiency.CCK-8 and Transwell assay was used to analyze the proliferation,invasion and migration ability of MG-63 cells,dual luciferase reporter gene experiment and qPCR were used to detect the targeted regulation relationship between LINC00173 and miR-130a-5p.At the same time,LINC00173 and miR-130a-5p were overexpressed,and the proliferation,invasion and migration ability of MG-63 cells were detect.Results:qPCR results showed that the expression of LINC00173 in MG-63 cells in pc-LINC00173 group was significantly higher than that in Mock group(P<0.01).After 48 and 72 hours of transfection,the cell proliferation activity of pc-LINC00173 group was lower than that of Mock group and pcDNA group(P<0.01).Transwell results showed that the number of invading and migrating cells in pc-LINC00173 group was significantly reduced compared with Mock group(P<0.05).There was a specific binding site between LINC00173 and miR-130a-5p.Compared with the miR-NC group,transfection of miR-130a-5p mimics could inhibit the relative luciferase activity of LINC00173-Wt cells(P<0.01).Compared with the Mock group and the pcDNA group,the expression of miR-130a-5p in MG-63 cells in pc-LINC0173 group decreased(P<0.01).Compared with pc-LINC00173 group and pc-LINC00173+miR-NC group,the expression of miR-130a-5p in MG-63 cells in pc-LINC00173+miR-130a-5p group was significantly increased,the absorbance value was significantly increased,and the number of invading and migrating cells was significantly increased(P<0.01).Conclusions:LINC00173 can inhibit the proliferation,invasion and migration of osteosarcoma MG-63 cells,and its mechanism may be related to the targeted inhibition of miR-130a-5p expression.