circ_0072088靶向miR-578促进类风湿关节炎滑膜成纤维细胞增殖和转移的机制研究

Mechanism of circ_0072088 promoting the proliferation and metastasis of rheumatoid arthritis synovial fibroblasts by targeting miR-578

目的:探讨circ_0072088对类风湿关节炎滑膜成纤维细胞增殖和转移的影响及其对miR-578的调控作用。方法:采用实时荧光定量聚合酶链反应(qRT-PCR)法检测正常滑膜组织、类风湿关节炎患者滑膜组织中circ_0072088和miR-578的表达量;原代分离培养类风湿关节炎滑膜成纤维细胞,将细胞分为si-NC组、si-circ_0072088组、miR-NC组、miR-578组、si-circ_0072088+anti-miR-NC组、si-circ_0072088+anti-miR-578组;采用细胞计数试剂盒(CCK-8)实验、平板克隆形成实验、划痕实验、Transwell小室实验分别检测细胞增殖、克隆形成、迁移及侵袭能力;双荧光素酶报告基因实验检测circ_0072088与miR-578的相互作用。结果:与正常滑膜组织比较,类风湿关节炎患者滑膜组织中circ_0072088的表达水平升高(P<0.05),miR-578的表达水平降低(P<0.05);与si-NC组比较,si-circ_0072088组细胞活力降低(P<0.05),克隆形成数及侵袭细胞数减少(P<0.05),划痕愈合率降低(P<0.05);双荧光素酶报告基因实验结果显示circ_0072088与miR-578特异性结合,并可调控miR-578的表达活性;与miR-NC组比较,miR-578组细胞活力降低(P<0.05),克隆形成数及侵袭细胞数均明显减少(P<0.05),划痕愈合率降低(P<0.05);与si-circ_0072088+anti-miR-NC组比较,si-circ_0072088+anti-miR-578组细胞活力升高(P<0.05),克隆形成数及侵袭细胞数均明显增多(P<0.05),划痕愈合率升高(P<0.05)。结论:抑制circ_0072088表达可通过靶向miR-578而降低类风湿关节炎滑膜成纤维细胞增殖、克隆形成、迁移及侵袭能力。

Objective:To explore the effect of circ_0072088 on the proliferation and metastasis of rheumatoid arthritis synovial fibroblasts and its regulation of miR-578.Methods:The quantitative real-time PCR(qRT-PCR)method was used to detect the expression of circ_0072088 and miR-578 in normal synovial tissue and synovial tissue of patients with rheumatoid arthritis.Rheumatoid arthritis synovial fibroblasts were primary isolated and cultured,rheumatoid arthritis synovial fibroblasts were divided into si-NC group,si-circ_0072088 group,miR-NC group,miR-578 group,si-circ_0072088+anti-miR-NC group,si-circ_0072088+anti-miR-578 group.The CCK-8 experiment,plate clone formation experiment,scratch experiment,and Transwell chamber experiment were used to detect cell proliferation,clone formation,migration and invasion capabilities,respectively.The dual luciferase reporter gene experiment was used to detect the interaction between circ_0072088 and miR-578.Results:Compared with normal synovial tissue,the expression level of circ_0072088 in the synovial tissue of patients with rheumatoid arthritis was increased(P<0.05),while the expression level of miR-578 was decreased(P<0.05).Compared with the si-NC group,the cell viability of the si-circ_0072088 group was decreased(P<0.05),the number of colonies and the number of invasive cells were decreased(P<0.05),the scratch healing rate was decreased(P<0.05).The results of the dual-luciferase reporter gene assay showed that circ_0072088 specifically binds to miR-578 and could regulate the expression activity of miR-578.Compared with the miR-NC group,the cell viability of the miR-578 group was reduced(P<0.05),the number of colonies and the number of invaded cells were significantly reduced(P<0.05),the scratch healing rate was reduced(P<0.05).Compared with the si-circ_0072088+anti-miR-NC group,the cell viability of the si-circ_0072088+anti-miR-578 group was increased(P<0.05),and the number of clone formation and the number of invasive cells were increased significantly(P<0.05),and the rate of scar healing was increased(P<0.05).Conclusion:Inhibition of the expression of circ_0072088 can reduce the proliferation,clone formation,migration and invasion of rheumatoid arthritis synovial fibroblasts by targeting miR-578.