miR-432对骨肉瘤细胞增殖、迁移、侵袭的影响及机制

发布时间：2018-04-18 02:26

本文选题：骨肉瘤 + 微小RNA-　；参考：《山东医药》2017年48期

【摘要】：目的探讨微小RNA-432(miR-432)对骨肉瘤细胞增殖、迁移和侵袭的影响及其作用机制。方法采用qRT-PCR技术检测人骨肉瘤细胞MG-63、人成骨细胞h FOB1.19中miR-432表达。将对数生长期MG-63细胞随机分为三组,分别转染miRNA对照质粒(miR-NC组)、miR-432抑制质粒(miR-432 inhibitor组)及miR-432过表达质粒(miR-432组),转染48 h时进行接种培养,分别检测细胞增殖(吸光度值)、迁移和侵袭情况。利用miRWalk和miRanda生物信息学软件预测miR-432靶基因可能为致癌基因CX3CL1,分别构建含有miR-432结合位点的CX3CL1基因3'-UTR区野生型(CX3CL1-wt)和突变型(CX3CL1-mut)荧光素酶报告载体。将MG-63细胞分别共转染CX3CL1-wt或CX3CL1-mut报告载体和miR-432载体(观察组),并设置共转染miR-NC载体作为对照组。转染48 h时,采用双荧光素酶试剂盒检测相对荧光素酶活性。结果 MG-63细胞中miR-432相对表达量低于h FOB1.19细胞(P0.01)。miR-NC组、miR-432 inhibitor组、miR-432组细胞中miR-432相对表达量分别为1.00±0.01、0.47±0.03、3.62±0.03,组间比较P均0.01。miR-432组培养24、36、48 h的细胞吸光度值均明显低于miR-NC组、miR-432 inhibitor组,miR-432 inhibitor组均明显高于miR-NC组。miR-432 inhibitor组及miR-432组细胞相对迁移率分别为(122±7)%、(65±4)%,相对侵袭率分别为(189±25)%、(49±11)%;上述指标组间比较P均0.05。CX3CL1-wt和miR-432共转染MG-63细胞后,观察组相对荧光素酶活性低于对照组(P0.05);CX3CL1-mut和miR-432共转染细胞后,对照组和观察组相对荧光素酶活性分别为1.00±0.11、0.93±0.13,两组比较P0.05。结论 miR-432过表达可抑制骨肉瘤细胞增殖、迁移和侵袭,靶向调控致癌基因CX3CL1可能是其作用机制。
[Abstract]:Objective to investigate the effects of microRNA-432miR-432) on proliferation, migration and invasion of osteosarcoma cells and its mechanism.Methods qRT-PCR technique was used to detect the expression of miR-432 in human osteosarcoma cell line MG-63 and human osteoblast h FOB1.19.The MG-63 cells in logarithmic growth phase were randomly divided into three groups. They were transfected with miRNA control plasmid / miR-NC group and miR-432 overexpression plasmid / mmiR-432 inhibitor group respectively. The cells were inoculated and cultured at 48 h after transfection.Cell proliferation (absorbance value), migration and invasion were measured.MiRWalk and miRanda bioinformatics software were used to predict that the miR-432 target gene might be the oncogene CX3CL1. The luciferase report vectors of CX3CL1 gene 3'-UTR region CX3CL1-wtand mutant CX3CL1-mutase containing miR-432 binding sites were constructed, respectively.MG-63 cells were cotransfected into CX3CL1-wt or CX3CL1-mut report vector and miR-432 vector respectively (observation group), and cotransfected miR-NC vector was set as control group.The relative luciferase activity was detected by double luciferase kit at 48 h after transfection.Results the relative expression of miR-432 in MG-63 cells was lower than that in h FOB1.19 cells (P 0.01). The relative expression of miR-432 in miR-432 inhibitor group was 1. 00 卤0. 01 卤0. 01 卤0. 03 卤3. 62 卤0. 03 respectively. The absorbance of miR-432 in P 0.01.miR-432 group was significantly lower than that in miR-NC group after 24 ~ 36 48 h culture.The relative mobility of the cells in the miR-NC group was significantly higher than that in the miR-NC group. The relative mobility of the cells in the inhibitor group and the miR-432 group were 122 卤7 and 65 卤4, respectively, and the relative invasion rates were 189 卤25 and 49 卤11. The P 0.05.CX3CL1-wt and miR-432 cotransfected MG-63 cells were compared between the two groups.The activity of relative luciferase in the observation group was lower than that in the control group (P 0.05), CX3CL1-mut and miR-432 cotransfected cells. The relative luciferase activity in the control group and the observation group was 1.00 卤0.110.93 卤0.13, respectively, compared with that in the two groups (P 0.05).Conclusion overexpression of miR-432 can inhibit proliferation, migration and invasion of osteosarcoma cells, and target regulation of oncogene CX3CL1 may be its mechanism.
【作者单位】：济南军区总医院;
【分类号】：R738.1

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