MicroRNA-203抑制骨肉瘤细胞增殖和迁移的机制研究

发布时间：2018-07-14 09:57

【摘要】：目的探讨microR NA-203(miR-203)对骨肉瘤细胞增殖和迁移的影响及机制。方法通过逆转录聚合酶链反应(RT-PCR)测定miR-203在正常成骨细胞hF OB及骨肉瘤细胞系中的表达,将MG-63细胞系分成miR-203组、Scramble组、RAB22A组、NC组,Lipofectamine 2000分别转染miR-203 mimics、Scrambles、pcD NA3.1-RAB22A、空白对照质粒;通过噻唑蓝(MTT)实验测定增殖能力,细胞划痕实验测定迁移能力;通过Target Scan和双荧光素酶实验预测及验证miR-203与RAB22A基因的靶向调节关系;RT-PCR测定RAB22A mR NA表达水平;Western blot测定E-cadherin、N-cadherin、Vimentin蛋白表达水平。结果 miR-203在骨肉瘤细胞系SAOS-2、SOSP-9607、U2OS及MG-63中较hF OB细胞系表达降低。MTT显示,在培养48和72 h后,miR-203组相对细胞数少于Scramble组(P0.05);培养24 h后,Scramble组细胞相对划痕面积小于miR-203组(P0.05);双荧光素酶实验示,miR-203与RAB22A基因存在靶向调节关系。RAB22A组相对划痕面积小于NC组(P0.05);MTT显示,在培养24、48和72 h后,RAB22A组相对细胞数多于NC组(P0.05);RAB22A组与NC组比较,E-cadherin下调表达,N-cadherin上调表达,Vimentin上调表达。结论 miR-203通过下调RAB22A基因表达,抑制上皮间质转化,进而抑制骨肉瘤细胞增殖与迁移。
[Abstract]:Objective to investigate the effect and mechanism of microR NA-203 (miR-203) on proliferation and migration of osteosarcoma cells. Methods the expression of miR-203 in normal osteoblast hFOB and osteosarcoma cell line was determined by reverse transcription-polymerase chain reaction (RT-PCR). MG-63 cell line was divided into miR-203 group, scramble group, RAB22A group and NC group, respectively transfected with miR-203 mimicsScramblespcNA3.1-RAB22A, blank control plasmid. The proliferation ability was measured by MTT assay and the migration ability was measured by cell scratch assay. The target regulatory relationship between miR-203 and RAB22A gene was predicted and verified by Target scan and double luciferase assay. RT-PCR was used to detect the expression level of RAB22A mRNA. Western blot was used to detect the expression of E-cadherin N-cadherinnin Vimentin protein. Results the expression of miR-203 in osteosarcoma cell line SAOS-2OS-9607 U2OS and MG-63 was lower than that in HF-OB cell line. After 48 and 72 hours of culture, the number of relative cells in miR-203 group was less than that in scramble group (P0.05), the relative scratch area in scramble group was smaller than that in miR-203 group after 24 hours culture (P0.05), and the relative scratch area in RAB22A group was lower than that in NC group (P0.05). The number of relative cells in RAB22A group was higher than that in NC group (P0.05). The down-regulated expression of N-cadherin and Vimentin upregulation in RAB22A group were higher than those in NC group. Conclusion miR-203 inhibits the proliferation and migration of osteosarcoma cells by down-regulating RAB22A gene expression and inhibiting epithelial mesenchymal transformation.
【作者单位】：华中科技大学同济医学院附属武汉中心医院急诊创伤外科;
【分类号】：R738.1

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