微RNA-429对肾透明细胞癌细胞生物学特征的影响  

Effects of microRNA-429 on the biological characteristics of renal clear cell carcinoma cells

作　　者：康小龙 李云[1] 郭洁 李荣婕 KANG Xiaolong;LI Yun;GUO Jie;LI Rongjie(Department of Urology,Neijiang Hospital of Traditional Chinese Medicine,Neijiang 641000,Sichuan Province,China)

机构地区：[1]内江市中医医院泌尿外科,四川内江641000

出　　处：《新乡医学院学报》2025年第4期277-287,293,共12页Journal of Xinxiang Medical University

摘　　要：目的探讨微RNA(microRNA,miR)-429对肾透明细胞癌细胞生物学特征的影响。方法采用实时荧光定量反转录聚合酶链式反应(qRT-PCR)检测人肾小管上皮细胞HK-2及肾透明细胞癌细胞ACHN、Caki-1、786-O、769-P中miR-429及SLC2A3 mRNA表达水平,其中Caki-1细胞中的miR-429、SLC2A3表达变化最为明显,因此选用Caki-1细胞进行后续实验。将对数生长期Caki-1细胞分为mimic NC组(转染mimic NC)、miR-429 mimic组(转染miR-429 mimic)、inhibitor NC组(转染inhibitor NC)、miR-429 inhibitor组(转染miR-429 inhibitor)、oe-NC组(转染oe-NC)、oe-SLC2A3组(转染oe-SLC2A3)、sh-NC组(转染sh-NC)、sh-SLC2A3组(转染sh-SLC2A3)、mimic NC+oe-NC组(同时转染mimic NC和oe-NC)、miR-429 mimic+oe-NC组(同时转染miR-429 mimic和oe-NC)和miR-429 mimic+oe-SLC2A3组(同时转染miR-429 mimic和oe-SLC2A3)。采用实时荧光定量反转录聚合酶链式反应法检测上述11组Caki-1细胞中miR-429及SLC2A mRNA的表达水平;3-(4,5-二甲基噻唑-2)-2,5-二苯基四氮唑溴盐法检测Caki-1细胞的增殖能力;Transwell实验检测Caki-1细胞的迁移、侵袭能力;双荧光素酶报告实验验证mimic NC组和miR-429 mimic组Caki-1细胞中miR-429与SLC2A3的结合关系;Western blot检测Caki-1细胞中SLC2A3和上皮-间充质转化(EMT)相关蛋白的表达;免疫荧光染色检测Caki-1细胞中EMT相关蛋白的表达。结果肾透明细胞癌细胞系中miR-429的表达显著下调,SLC2A3 mRNA的表达显著上调,过表达miR-429能显著抑制癌细胞的增殖、迁移、侵袭和EMT过程。SLC2A3能促进肾透明细胞癌细胞系EMT进程,并能恢复miR-429过表达对细胞EMT的抑制作用。双荧光素酶报告实验证实miR-429与SLC2A3相互结合,miR-429可靶向抑制SLC2A3。结论miR-429通过靶向抑制SLC2A3的表达,抑制肾透明细胞癌细胞增殖、迁移、侵袭和EMT过程。Objective To investigate the effects of microRNA(miR)-429 on the biological characteristics of clear cell renal cell carcinoma(ccRCC)cells.Methods Real-time quantitative reverse transcription polymerase chain reaction(qRT-PCR)was used to measure miR-429 and solute carrier family 2 member 3(SLC2A3)mRNA levels in renal tubular epithelial cells HK-2 and ccRCC cells ACHN,Caki-1,786-O and 769-P.Since the most significant changes were observed in the expre-ssion of miR-429 and SLC2A3 mRNAs in Caki-1cells,so they were chosen for subsequent tests.Caki-1cells in the logarithmic growth phase were divided into 11 groups:mimic NC(transfected with mimic NC),miR-429 mimic(transfected with miR-429 mimic),inhibitor NC(transfected with inhibitor NC),miR-429 inhibitor(transfected with miR-429 inhibitor),oe-NC(transfected with oe-NC),oe-SLC2A3(transfected with oe-SLC2A3),sh-NC(transfected with sh-NC),sh-SLC2A3(transfected with sh-SLC2A3),mimic NC+oe-NC(co-transfected with mimic NC and oe-NC),miR-429 mimic+oe-NC(co-transfected with miR-429 mimic and oe-NC),and miR-429 mimic+oe-SLC2A3 groups(co-transfected with miR-429 mimic and oe-SLC2A3).qRT-PCR was used to detect the expression levels of miR-429 and SLC2A3 mRNAs in Caki-1 cells of the 11 groups.3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide was used to assess the viability of Caki-1 cell proliferation.Transwell testing was conducted to evaluate Caki-1 cell migration and invasion capabilities.A dual-luciferase reporter assay was performed to validate the targeting relationship between miR-429 and SLC2A3 in Caki-1 cells of mimic NC and miR-429 mimic groups.Western blot and immunofluorescence staining were employed to detect the expression of epithelial-mesenchymal transition(EMT)-related proteins.Results The expression of miR-429 was significantly downregulated while that of SLC2A3 mRNAs was significantly upregulated in ccRCC cell lines.Overexpression of miR-429 significantly inhibited the proliferation,migration,invasion,and EMT process of ccRCC cells.SLC2A3 promoted

关 键 词：微RNA 溶质载体家族2成员3 肾透明细胞癌 上皮间充质转化 

分 类 号：R737.11[医药卫生—肿瘤]