沉默环状RNA circ_0052012对骨肉瘤细胞增殖和侵袭的影响  

Knockdown of circ_0052012 attenuated proliferation and invasion of osteosarcoma cells

作　　者：吴晗[1] 朱旭[1] 吴学建[1] Wu Han;Zhu Xu;Wu Xuejian

机构地区：[1]郑州大学第一附属医院骨科,450052

出　　处：《中华实验外科杂志》2020年第12期2274-2277,共4页Chinese Journal of Experimental Surgery

摘　　要：目的观察骨肉瘤中环状RNA circ_0052012表达和沉默环状RNA circ_0052012表达对骨肉瘤细胞增殖和侵袭的影响。方法实时定量反转录聚合酶链反应(RT-qPCR)方法检测2017年10月至2018年12月郑州大学第一附属医院确诊的31对骨肉瘤标本癌组织和癌旁正常组织circ_0052012表达水平。使用针对circ_0052012的小干扰RNA(siRNA)转染骨肉瘤细胞系MG63,将体外培养的MG63细胞分为空白组(未转染)、阴性对照组(转染阴性对照si-NC)和沉默组(转染si-circ)。采用细胞增殖实验和Transwell实验,检测沉默RNA circ_0052012表达对MG63细胞增殖和侵袭的影响。采用生物信息学方法预测circ_0052012与微小RNA(miRNA,miR)-7的结合位点,利用双荧光素酶报告实验、RT-qPCR技术和Pearson相关分析,验证circ_0052012对miR-7的靶向调控作用及其在骨肉瘤中表达水平的相关性。两组间数据比较采用t检验,多组间数据比较采用单因素方差分析(ANOVA)。结果骨肉瘤组织中circ_0052012的表达水平显著高于对应癌旁正常组织,差异有统计学意义(2.452±1.061比1.007±0.644,t=10.180,P<0.01),且circ_0052012表达水平与骨肉瘤TNM分期、远处转移呈明显正相关(P<0.05)。在骨肉瘤细胞系MG63中,培养24、48、72 h后,circ_0052012沉默组细胞的增殖(0.297±0.012、0.607±0.074、0.797±0.067)均明显低于空白对照组(0.407±0.035、0.820±0.070、1.287±0.050)和阴性对照组(0.383±0.012、0.853±0.064、1.240±0.092),差异有统计学意义(24 h,t=-5.154、-9.192,P<0.01;48 h,t=-3.635、-4.391,P<0.05;72 h,t=-10.170、-6.778,P<0.01)。沉默组细胞的侵袭能力(39.400±6.066)也明显低于空白对照组(132.600±8.620)和阴性对照组(126.000±10.950),差异有统计学意义(t=-19.770、-15.460,P<0.01)。双荧光素酶报告实验证实circ_0052012与miR-7靶向结合,且在骨肉瘤组织中的表达呈明显负相关(r=-0.661,P<0.05)。通过抑制miR-7,可部分逆转沉默circ_0052012对MG63细胞增殖和侵袭的Objective To investigate circ_0052012 expression in osteosarcoma tissues and the effects of silencing circ_0052012 on proliferation and invasion of osteosarcoma cells.Methods The expression levels of circ_0052012 were detected by real-time quantitative reverse transcriptase-polymerase chain reaction(RT-qPCR)in 31 osteosarcoma specimens and adjacent normal tissues.By performing the transfection of small interfering RNA(siRNA)targeting circ_0052012,MG63 cells were therefore divided into blank group(not transfected),negative control group(transfected with negative control si-NC)and knockdown group(transfected with si-circ).Cell counting kit-8(CCK-8)and Transwell assays were used to detect the proliferation and invasion of MG63 osteosarcoma cells.Dual-luciferase reporting assay,RT-qPCR and correlation analysis were used to determine the targeted regulatory effects of circ_0052012 on microRNA(miRNA,miR)-7.SPSS 21.0 software was used to analyze the data.Results The expression level of circ_0052012 in osteosarcoma tissues was significantly higher than that in adjacent normal tissues(2.452±1.061 vs.1.007±0.644,t=10.180,P<0.01).Silenced circ_0052012 significantly inhibited the proliferation and invasion of MG63 cells,compared to blank group and si-NC group.Moreover,circ_0052012 could function as a molecular sponge targeting miR-7 with a negative correlation in osteosarcoma specimens(r=-0.661,P<0.05).By inhibiting miR-7,the suppress effect of circ_0052012 knockdown on proliferation and invasion of MG63 cells can be partially reversed.Conclusion Knockdown of circ_0052012 could suppress the proliferation and invasion of osteosarcoma cells by up-regulating the expression of miR-7.

关 键 词：骨肉瘤 circ_0052012 微小RNA-7 增殖 侵袭 

分 类 号：R738.1[医药卫生—肿瘤]