miRNA-6516-5p通过靶向ODC1调控肾癌细胞的增殖和迁移  

miRNA-6516-5p regulates the proliferation and migration of renal cancer cells by targeting ODC1

作　　者：黄耿[1] 桂定文[1] 徐祖伟 付金伦 罗帅 赵云飞 万京桦 Huang Geng;Gui Dingwen;Xu Zuwei;Fu Jinlun;Luo Shuai;Zhao Yunfei;Wan Jinghua(Department of Urology,Huangshi Central Hospital,Affiliated Hospital of Hubei Institute of Technology,Edong Healthcare Group,Huangshi 435000,China;Hubei Provincial Key Laboratory of Renal Disease Occurrence and Intervention,Huangshi 435000,China;Hubei Provincial Key Laboratory of Occupational Hazard Identification and Control,Wuhan University of Science and Technology,Wuhan 430065,China)

机构地区：[1]鄂东医疗集团黄石市中心医院(湖北理工学院附属医院)泌尿外科,黄石435000 [2]肾脏疾病发生与干预湖北省重点实验室,黄石435000 [3]武汉科技大学职业危害识别与控制湖北省重点实验室,武汉430065

出　　处：《国际外科学杂志》2022年第3期194-198,I0006,共6页International Journal of Surgery

基　　金：湖北省卫生健康科研基金(WJ2019H158)。

摘　　要：目的探讨微小RNA(miRNA)-6516-5p在肾癌细胞系中的表达及调控肾癌细胞增殖和迁移的分子机制。方法用实时荧光定量聚合酶链反应(qRT-PCR)检测肾癌细胞系和正常近端肾小管上皮细胞系中miR-6516-5p的表达。用脂质体法分别将miR-6516-5p模拟物和无意义序列(NC)瞬时转染至miR-6516-5p表达最低的肾癌细胞,即miR-6516-5p组和NC组。qRT-PCR检测转染细胞miR-6516-5p的表达。CCK-8法和Transwell迁移实验检测转染细胞的增殖和迁移。生物信息学软件和双荧光素酶基因报告实验分别预测和验证miR-6516-5p对靶基因的调控。qRT-PCR和Western blotting法分别检测转染细胞中靶基因的表达。计量资料以均数±标准差(±s)表示,两组间比较采用t检验,多组间比较采用单因素方差分析。结果miR-6516-5p在肾癌细胞系中的表达均明显低于正常近端肾小管上皮细胞(P<0.01),786-O细胞中miR-6516-5p的表达最低(F=27.69,P<0.01)。NC组和miR-6516-5p组786-O细胞中miR-6516-5p的表达分别为1.01±0.08和9.91±1.16,与NC组比较,miR-6516-5p组786-O细胞中miR-6516-5p表达明显增加(t=7.63,P<0.01)。上调miR-6516-5p能显著抑制786-O细胞的增殖(P<0.05)。NC组和miR-6516-5p组细胞迁移数分别为(85.65±8.77)个和(28.05±6.20)个,过表达miR-6516-5p可抑制786-O细胞的迁移(t=5.36,P<0.01)。miR-6516-5p的靶基因可能是鸟氨酸脱羧酶1(ODC1),miR-6516-5p能显著抑制野生型ODC1-3′UTR的荧光素酶活性(t=9.83,P<0.01)。上调miR-6516-5p可降低786-O细胞中ODC1 mRNA和蛋白的表达(P<0.01)。结论miR-6516-5p在肾癌细胞系中表达降低,miR-6516-5p通过靶向ODC1抑制肾癌786-O细胞的增殖和迁移,miR-6516-5p可能成为肾癌潜在的分子靶点。Objective:To explore the expression of microRNA(miRNA)-6516-5p in renal cancer cell lines and the molecular mechanisms regulating the proliferation and migration of renal cancer cells.Methods:quantitative real-time polymerase chain reaction(qRT-PCR)was used to detect the expression of miR-6516-5p in renal cancer cell lines and normal proximal renal tubular epithelial cell lines.The liposome method was used to transiently transfect miR-6516-5p mimic and nonsense sequence(NC)into renal cancer cells with the lowest expression of miR-6516-5p,namely miR-6516-5p group and NC group.qRT-PCR was used to detect the expression of miR-6516-5p in transfected cells.CCK-8 and Transwell migration experiment were used to detect the proliferation and migration of transfected cells.Bioinformatics software and dual luciferase gene report experiment were used to predict and verify the regulation of miR-6516-5p on target gene,respectively.qRT-PCR and Western blotting were used to detect the expression of target gene in transfected cells.Measurement data were expressed as mean±standard deviation(±s),t-test was used for comparison between two groups,and one-way analysis of variance was used for comparison between multiple groups.Results:The expression of miR-6516-5p in renal cancer cell lines was significantly lower than that of normal proximal tubular epithelial cells(P<0.01),and the expression of miR-6516-5p in 786-O cells was the lowest(F=27.69,P<0.01).The expression of miR-6516-5p in 786-O cells in NC group and miR-6516-5p group was 1.01±0.08 and 9.91±1.16,respectively.Compared with the NC group,the expression of miR-6516-5p in 786-O cells in the miR-6516-5p group was significantly increased(t=7.63,P<0.01).Up-regulation of miR-6516-5p can significantly inhibit the proliferation of 786-O cells(P<0.05).The migration numbers of NC group and miR-6516-5p group were 85.65±8.77 and 28.05±6.20,respectively.Overexpression of miR-6516-5p could inhibit the migration of 786-O cells(t=5.36,P<0.01).The target gene of miR-6516-5p may be orni

关 键 词：肾肿瘤 miR-6516-5p 鸟氨酸脱羧酶1 细胞增殖 细胞迁移 

分 类 号：R737.11[医药卫生—肿瘤]