miR-183对滑膜肉瘤细胞株SW982转移潜能及EGR1表达影响研究  

Effects of miR-183 on metastasis potential and EGR1 expression of Human Synovial Sarcoma cells line SW982

作　　者：董超[1] 周洋[1] 锡林宝勒日 鲁学良[1] 谢鹏鸣[1] 哈斯鲁[1] 

机构地区：[1]新疆医科大学附属肿瘤医院骨与软组织病区,新疆乌鲁木齐830011

出　　处：《中华肿瘤防治杂志》2015年第16期1270-1276,共7页Chinese Journal of Cancer Prevention and Treatment

摘　　要：目的探讨微小RNA-183(miRNA-183)对人滑膜肉瘤细胞株SW982转移潜能的影响,并分析miR-183对早期生长反应因子1(early growth response,EGR1)表达影响的研究。方法培养人滑膜肉瘤细胞株SW982细胞,采用脂质体转染方法转染miR-183mimics、inhibitor和negative control,并分为miR-183 mimics模拟物组(mimics组),miR-183inhibitor抑制剂组(inhibitor组),miR-183negative control阴性对照组(NC)以及正常对照组(normal)。流式细胞术检测各组细胞转染效率;MTT法检测各组细胞转染前后细胞增殖率的改变;Transwell实验检验各组转染前后SW982细胞迁移和侵袭能力的变化;蛋白质印迹法检测各组转染前后EGR1蛋白表达量的变化;双荧光素酶报告基因实验验证EGR1是否为miR-183靶基因。结果转染miR-183mimics、inhibitor和negative control后,SW982细胞中miR-183的相对含量明显改变;与正常对照组(1.16±0.52)相比,mimics组(75.30±9.90)miR-183含量明显增加(P<0.001),inhibitor组(0.22±0.09)miR-183含量明显降低(P<0.001),NC组(1.06±0.98)miR-183含量无明显变化,P=0.637。MTT法测得转染后24、48、72和96h,SW982细胞增殖率无明显变化。Transwell实验结果显示,在迁移实验中,mimics组和inhibitor组穿膜细胞数分别为(108.61±4.94)和(54.36±4.03)个,P<0.001。在侵袭实验中,mimics组和inhibitor组穿膜细胞数分别为(79.13±5.77)和(20.87±3.17)个,P<0.001。在EGR1表达分析中,转染48h后mimics组中EGR1的相对表达量(286.59±32.77)较正常对照组(373.00±38.40)明显降低,P<0.001;inhibitor组中EGR1的相对表达量(496.93±42.61)较正常对照组明显增高,P=0.012;双荧光素酶报告基因实验表明,miR-183特异性靶向EGR1基因。结论转染miR-183mimics后,miR-183表达增加,SW982细胞转移潜能增强,EGR1表达降低;转染miR-183inhibitor后,miR-183表达降低,SW982细胞转移潜能降低,EGR1表达增加。EGR1可作为miR-183的潜在靶点。OBJECTIVE To investigate the influence on metastasis potential of synovial sarcoma via regulating the expression of microRNA-183 in human synovial sarcoma cells line SW982 and analyze the effects of miR 183 on EGR1. METHODS We cultured the human synovial sarcoma cells line SW982. We changed the expression of miR 183 by trans- fecting the miR-183 mimics, inhibitor and NC to SW982 cells line, and divided the cells into four groups： miR-183 mimics group, miR-183 inhibitor group, miR 183 negative control group and normal group. Transfection efficiency was detected by RT-PCR and FACS. MTT was used to detect the cell proliferation after transfection. Transwell model was used to ex- am the change of migratory and invasive abilities after transfection. Western blot was used to analyze the expression of EGR1 after transfection. Dual-luciferase report assay was used to identify whether EGR1 was the direct target of miR 183. RESULTS Real time PCR analysis revealed that the expression of miR-183 was higher in mimics group （75.30±9.90） compared to the normal groups （1.16±0.52,P〈0. 001）. The miR-183 expression in inhibitor （0.22±0.09） was lower than that of normal groups （P〈0,001）. In normal and negative control （NC） groups（1.06±0.98）, the result was similar （P=0. 637）. Twenty-four hours, 48, 72 and 96 h after transfeetion, the proliferation of SW982 cells had no significant effect. Transwell analysis revealed that in migratory study the number of SW982 cells penetrated the membrane in mimics and inhibitors group were （108.61±4.94） and （54.36±4.03）, respectively（P〈0. 001）. In invasive study, the number of SW982 cells penetrated this membrane in mimics and inhibitors group were （79. 13±5.77） and （20.87±3. 17）, respectively （P〈0. 001）. The expression of EGR1 was detected by western blotting, the result showed that in mimics group （286.59±32.77） the relative expression of EGR1 was dramatically decreased compared with inhibi- tor group （373.00±38.40, P�

关 键 词：microRNA-183 EGR1 滑膜肉瘤 侵袭 迁移 

分 类 号：R738.5[医药卫生—肿瘤]