MiR-515-5p通过调控硫酸软骨素蛋白聚糖4表达对卵巢癌A2780细胞增殖和转移的影响及其机制

Effects of miR-515-5p on the proliferation and metastasis of ovarian cancer cell A2780 by regulating the expression of chondroitin sulfate proteoglycan 4 and its mechanism

目的探讨miR-515-5p对硫酸软骨素蛋白聚糖4(CSPG4)的靶向调控作用,以及对卵巢癌细胞系A2780细胞增殖和转移的影响。方法通过生物信息学工具预测miR-515-5p的靶基因。Real-time PCR和Western blotting法检测65例卵巢癌组织和与其对应的癌旁组织中miR-515-5p和CSPG4的表达。将A2780细胞分为miR-NC组、miR-515-5p组、si-NC组、si-CSPG4组、miR-515-5p+pcDNA组和miR-515-5p+pcDNA-CSPG4组。MTT法检测细胞增殖活力,Transwell检测细胞迁移和侵袭数,Western blotting检测细胞周期蛋白D1(cyclinD1)、P21、基质金属蛋白酶(MMP)-2和MMP-9蛋白的表达。双荧光素酶报告基因实验和Western blotting验证miR-515-5p对CSPG4基因的调控作用。结果生物信息学分析显示,CSPG4是miR-515-5p的潜在靶基因之一。与癌旁组织相比,卵巢癌组织miR-515-5p的表达较低,CSPG4的表达较高(P<0.05)。与miR-NC组、miR-515-5p组相比,A2780细胞cyclinD1、MMP-2和MMP-9蛋白的表达较低,P21蛋白的表达较高,细胞活力较低,细胞迁移和侵袭数较少(P<0.05)。与si-NC组相比,si-CSPG4组A2780细胞cyclinD1、MMP-2和MMP-9蛋白的表达较低,P21蛋白的表达较高,细胞活力较低,细胞迁移和侵袭数较少(P<0.05)。与miR-515-5p+pcDNA组相比,miR-515-5p+pcDNA-CSPG4组A2780细胞cyclinD1、MMP-2和MMP-9蛋白的表达较高,P21蛋白的表达较低,细胞活力较高,细胞迁移和侵袭数较多(P<0.05)。MiR-515-5p靶向并负性调控CSPG4表达。结论MiR-515-5p通过靶向CSPG4可抑制卵巢癌细胞A2780增殖和转移。

Objective To investigate the targeted regulation of miR-515-5 p on chondroitin sulfate proteoglycan 4(CSPG4)and its effect on the proliferation and metastasis of ovarian cancer cell A2780.Methods The target genes of miR-515-5 p were predicted by bioinformatics tools.Real-time PCR and Western blotting were used to detect the expression of miR-515-5 p and CSPG4 in 65 cases ovarian cancer tissues and adjacent tissues.Ovarian cancer cells A2780 were divided into miR-NC group,miR-515-5 p group,si-NC group,si-CSPG4 group,miR-515-5 p+pcDNA group,miR-515-5 p+pcDNA-CSPG4 group.MTT assay was used to detect cell proliferation.Transell was applied to detect cell migration and invasion,and Western blotting was selected to detect cyclinD1,P21,matrix metalloproteinase(MMP)-2 and MMP-9 protein expression.The double luciferase reporter gene experiment and Western blotting confirmed the regulation effect of miR-515-5 p on CSPG4 gene.Results Bioinformatics analysis showed that CSPG4 was one of the potential target genes of miR-515-5 p.Compared with the adjacent tissues,the expression of miR-515-5 p was lower in ovarian cancer tissues,and the expression of CSPG4 was higher(P<0.05).Compared with the miR-NC group,the expression of cyclinD1,MMP-2 and MMP-9 protein was lower in A2780 cells of miR-515-5 p group,the expression of P21 protein was higher,the cell viability was lower,and the number of cell migration and invasion was lower(P<0.05).Compared with the si-NC group,the expression of cyclinD1,MMP-2 and MMP-9 proteins were lower in A2780 cells in si-CSPG4 group,P21 protein was higher,cell viability was lower,and the number of cell migration and invasion was lower(P<0.05).Compared with the miR-515-5 p+pcDNA group,the expression of cyclinD1,MMP-2 and MMP-9 proteins was higher in A2780 cells of miR-515-5 p+pcDNA-CSPG4 group,the expression of P21 protein was lower,and the cell viability was higher,the number of cell migration and invasion was higher(P<0.05).MiR-515-5 p targeted and negatively regulated CSPG4 expression.Conclusion MiR-515-5 p could inhibit the proliferation and metastasis of ovarian cancer cell A2780 by targeting CSPG4.