miR-29a-3p靶向TRAF5抑制骨肉瘤细胞顺铂耐药  

miR-29a-3p inhibits cisplatin resistance of osteosarcoma cells by targeting TRAF5

作　　者：匡健 董文刚 丁家云 谢晴 刘文毅 周志坚 刘军 KUANG Jian;DONG Wengang;DING Jiayun;XIE Qing;LIU Wenyi;ZHOU Zhijian;LIU Jun(Center for Diseases Control and Prevention,Southern Theater Command,Guangdong Guangzhou 510603,China;Department of Emergency Surgery,Shaanxi Provincial People's Hospital,Shaanxi Xi'an 710068,China)

机构地区：[1]南部战区疾病预防控制中心,广东广州510603 [2]陕西省人民医院急诊外科,陕西西安710068

出　　处：《现代肿瘤医学》2025年第4期576-582,共7页Journal of Modern Oncology

基　　金：广东省基础与应用基础研究基金资助项目(编号:2020A1515010299,2020A1515010931)。

摘　　要：目的:探究miR-29a-3p通过调控肿瘤坏死因子受体相关因子5(tumor necrosis factor receptor associated factor-5,TRAF5)的表达对骨肉瘤细胞顺铂耐药的影响。方法:采用real-time PCR实验检测骨肉瘤细胞系中miR-29a-3p表达水平,对骨肉瘤143B细胞施加顺铂药物处理,检测细胞随着药物作用时间、浓度的增加miR-29a-3p表达丰度的变化。对转染miR-29a-3p mimics的143B细胞施加顺铂处理,而后利用流式细胞术、Western blotting实验检测其对细胞凋亡的影响,并通过CCK-8实验检测细胞增殖的变化。利用生物信息学网站TargetScan预测得到miR-29a-3p的靶基因TRAF5,通过双荧光素酶报告基因实验检测miR-29a-3p与TRAF5的靶向关系,而后利用real-time PCR和Western blotting实验检测miR-29a-3p与TRAF5的表达水平。结果:real-time PCR实验结果显示相比于成骨细胞系hFOB1.19,miR-29a-3p在骨肉瘤细胞系中低表达,且在143B细胞中表达水平最低。顺铂处理143B细胞后,miR-29a-3p表达呈时间、浓度依赖性下调。对转染miR-29a-3p mimics的143B细胞施加顺铂处理后,流式细胞术结果显示miR-29a-3p能够促进顺铂处理后的143B细胞凋亡,CCK-8实验结果表明miR-29a-3p能够抑制顺铂处理后的143B细胞增殖,Western blotting实验结果表明miR-29a-3p上调使顺铂处理后的143B细胞凋亡相关基因表达显著改变。生物信息学预测TRAF5是miR-29a-3p的靶基因,双荧光素酶报告基因实验证实miR-29a-3p可靶向抑制TRAF5。过表达miR-29a-3p后,143B细胞TRAF5表达水平下调。结论:miR-29a-3p可能通过下调TRAF5的表达抑制143B细胞顺铂耐药。Objective:To explore the effect of miR-29a-3p on cisplatin resistance of osteosarcoma cells by regulating the expression of tumor necrosis factor receptor associated factor-5(TRAF5).Methods:Real-time PCR assay was used to detect the level of miR-29a-3p in osteosarcoma cell lines,then miR-29a-3p level in 143B cells was detected by real-time PCR assay after administration of cisplatin to osteosarcoma 143B cells to inspect that whether miR-29a-3p changed along the administration time and concentration or not.After transfection of miR-29a-3p mimics into cisplatin-administered 143B cells,flow cytometry and Western blotting assays were carried out to investigate the effects of miR-29a-3p on apoptosis of 143B cells,and CCK-8 assay was carried out to study the effects of miR-29a-3p on proliferation of 143B cells.Online bioinformatics prediction through TargetScan website and dual luciferase reporter assays were performed to testify the targeting role of miR-29a-3p on TRAF5.Real-time PCR and Western blotting detected the expression level of miR-29a-3p and TRAF5.Results:Real-time PCR results showed that miR-29a-3p expression was aberrantly downregulated in osteosarcoma cell lines compared with osteoblast cell line hFOB1.19,and was the lowest in 143B cells.After administration of cisplatin,miR-29a-3p level was gradually downregulated dependent on increment of cisplatin concentration and action time in osteosarcoma 143B cells.After transfection of miR-29a-3p mimics into cisplatin-administered 143B cells,flow cytometry result demonstrated that miR-29a-3p improved cisplatin-administered 143B cells apoptosis,and CCK-8 assays results demonstrated that miR-29a-3p inhibited cisplatin-administered 143B cells proliferation.Western blotting result showed that the upgrading of miR-29a-3p significantly changed the expression of apoptosis-related genes of 143B cells after cisplatin treatment.Bioinformatics predicts that TRAF5 was the target gene of miR-29a-3p.Dual luciferase reporter assay confirmed that miR-29a-3p can target the TRAF5

关 键 词：骨肉瘤 miR-29a-3p TRAF5 顺铂耐药 

分 类 号：R738.1[医药卫生—肿瘤]