环状RNA_0001495/微小RNA-527/Robo1轴对膀胱癌细胞增殖和转移的影响  

Effect of circular RNA_0001495/microRNA-527/Robo1 axis on proliferation and metastasis of bladder cancer cells

作　　者：程鹤鹏 冯留伟 高磊 侯淑超 李扬 Cheng Hepeng;Feng Liuwei;Gao Lei;Hou Shuchao;Li Yang(Department of Urology,Huaihe Hospital,Henan University,Kaifeng 475000,China)

机构地区：[1]河南大学淮河医院泌尿外科,开封475000

出　　处：《中华实验外科杂志》2023年第7期1355-1358,共4页Chinese Journal of Experimental Surgery

基　　金：河南省科技攻关项目(222102310717)。

摘　　要：目的探讨环状RNA(circRNA)_0001495/微小RNA(miR)-527/Robo1轴对膀胱癌细胞增殖和转移的影响。方法人膀胱癌细胞系T24培养至对数生长期,分为circRNA对照组和circRNA_0001495KD组,采用对照circRNA短发卡RNA(shRNA)和circRNA_0001495shRNA慢病毒感染T24细胞,建立稳转细胞系。分别采用细胞计数试剂盒(CCK-8)和5-乙炔基-2脱氧尿嘧啶核苷(EdU)染色分析两组细胞增殖能力;采用划痕实验和Transwell实验分析两组细胞迁移和侵袭能力。生物信息学和双荧光素酶报告基因分析circRNA_0001495的靶基因;采用荧光定量聚合酶链反应(PCR)分析circRNA_0001495靶基因miR-527表达水平;采用蛋白质印迹法(Westernblot)分析miR-527的靶蛋白。组间计量数据比较采用t检验。结果对照组细胞circRNA_0001495表达水平(1.02±0.06)明显高于circRNA_0001495KD组细胞(0.67±0.05),差异有统计学意义(t=10.750,P<0.05)。对照组细胞吸光度(A)值水平(1.97±0.06)明显高于circRNA_0001495KD组细胞(1.67±0.09),差异有统计学意义(t=6.262,P<0.05)。对照组细胞EdU阳性率水平(1.97±0.06)明显高于circRNA_0001495KD组细胞(1.67±0.09),差异有统计学意义(t=6.262,P<0.05)。对照组细胞划痕愈合率[(79.65±4.05)%]明显高于circRNA_0001495KD组细胞[(55.13±5.41)%],差异有统计学意义(t=8.880,P<0.05)。对照组细胞迁移数量[(114.50±12.37)个]明显高于circRNA_0001495KD组细胞[(84.50±12.90)个],差异有统计学意义(t=4.112,P<0.05)。miR-527是circRNA_0001495的靶基因。对照组细胞miR-527表达水平(0.88±0.05)明显低于circRNA_0001495KD组细胞(1.38±0.10),差异有统计学意义(t=10.690,P<0.05)。Westernblot结果显示,对照组细胞Robo1蛋白表达水平(1.25±0.14)明显高于circRNA_0001495KD组细胞(0.79±0.08),差异有统计学意义(t=7.196,P<0.05)。结论circRNA_0001495敲降可显著抑制膀胱癌细胞的增殖和转移,可能通过靶向调节miR-527/Robo1轴实现的。Objective To explore the effects of circular RNA(circRNA)_0001495/microRNA(miR)-527/Robol axis on the proliferation and metastasis of bladder cancer cells.Methods Human bladder cancer cell line T24 was cultured to logarithmic growth stage and divided into circRNA control group and circRNA_0001495 KD group.T24 cells were infected with control circRNA short hairpin RNA(shRNA)and circRNA_0001495 shRNA Lentivirus and stable cell lines were established.The prolifera-tion ability of the two groups was analyzed by cell counting kit-8(CCK-8)assay and 5-Ethynyl-2'-deoxyuri-dine(EdU)staining.The migration and invasion ability of the two groups was analyzed by wounding heal-ing and Transwell test.The target gene of circRNA_0001495 was analyzed by bioinformatics and Dual Lu-ciferase Reporter Gene.The miR-527 expression level was detected by fluorescence quantitative polymerase chain reaction(PCR).The target protein of miR-527 was detected by Western blotting.The comparison of measurement data between groups was conducted using t-test.Results The expression level of circRNA_0001495 in control group(1.02±0.06)was significantly higher than that in circRNA_0001495 KD group(0.67±0.05,t=10.750,P<0.05).The absorbance(A)value in the control group(1.97±0.06)was significantly higher than that of circRNA_0001495 KD group(1.67±0.09,t=6.262,P<0.05).The positive rate of EdU in the control group(78.47±6.11)was significantly higher than that in circRNA_0001495 KD group(51.27±3.62,t=6.262,P<0.05).The scratch healing rate in the control group[(79.65±4.05)%]was significantly higher than that in the circRNA_0001495 KD group[(55.13±5.41)%,t=8.880,P<0.05].The number of migrating cells[(114.50±12.37)cells]in the control group was significantly greater than that in the circRNA_0001495 KD group[(84.50±12.90)cells,t=4.112,P<0.05].The expression level of miR-527 in the control group(0.88±0.05)was significantly lower than that in the circRNA_0001495 KD group(1.38±0.10,t=10.690,P<0.05).Western blotting results showed that the expression leve

关 键 词：环状RNA 微小RNA ROBO1 膀胱癌 增殖 转移 

分 类 号：R737.14[医药卫生—肿瘤]