Wnt/β-catenin信号通路在miR-21介导的非小细胞肺癌顺铂耐药性中的作用  

Role of Wnt/β-catenin signaling pathway in miR-21-mediated cisplatin resistancein non-small cell lung cancer

作　　者：郑传会 林莉[1] 汪霞 王乃刚 ZHENG Chuanhui;LIN Li;WANG Xia;WANG Naigang(Department of Pathology,The Second Affiliated Hospital of Nanjing Medical University,Nanjing 210011;Department of Critical Care Medicine,Heze Hospital,Shandong Provincial Hospital,Heze municipal Hospital,Heze 274000,China)

机构地区：[1]南京医科大学第二附属医院病理科,江苏南京210011 [2]山东省立医院菏泽医院菏泽市立医院重症医学科,山东菏泽274000

出　　处：《西安交通大学学报(医学版)》2025年第2期238-248,共11页Journal of Xi’an Jiaotong University(Medical Sciences)

基　　金：2023年江苏省自然科学基金项目(No.yl2021 lh01)。

摘　　要：目的探究microRNA-21-5p(miR-21)在非小细胞肺癌顺铂(cisplatin,CDDP)耐药性中的作用和机制。方法通过RT-qPCR检测非小细胞肺癌患者癌组织和癌旁组织中miR-21的表达。用非小细胞肺癌细胞系H1299和H1975构建非小细胞肺癌CDDP耐药细胞H1299/CR和H1975/CR,CCK-8检测各组细胞在CDDP处理时的细胞活性,流式细胞术分析细胞凋亡。双荧光素酶检测H1299细胞中miR-21与PTEN的作用关系,RT-qPCR检测miR-21表达,Western blotting检测PTEN和PTEN下游Wnt/β-catenin信号通路蛋白表达。用H1299和H1299/CR构建荷瘤裸鼠模型,验证miR-21对非小细胞肺癌CDDP治疗敏感性的影响。结果非小细胞肺癌患者癌组织miR-21的表达显著高于癌旁组织(P<0.001)。与H1299和H1975细胞相比,耐药细胞H1299/CR和H1975/CR中miR-21表达显著升高(P<0.05)。H1299/CR和H1975/CR细胞经过inhibitor转染后用不同浓度的CDDP处理,CCK-8检测结果显示,CDDP≥12μmol/L时,与inhibitor NC组相比,miR-21 inhibitor组细胞活性显著降低(P<0.001);流式细胞仪检测结果显示,与未处理组相比,CDDP组和miR-21 inhibitor组细胞凋亡率无显著变化,CDDP+miR-21 inhibitor组细胞凋亡率显著升高(P<0.001)。H1299和H1975细胞转染miR-21模拟分子(miR-21 mimic)后,用不同浓度的CDDP处理细胞时,CDDP≥12μmol/L,与miR-21 mimic组相比,mimic-NC组的细胞活性显著降低(P<0.05)。Target scan网站预测显示miR-21与PTEN的3′-UTR区域结合。双荧光素酶检测表明miR-21和PTEN的蛋白表达存在靶向调控关系。PTEN过表达(PTEN-OE)与miR-21 mimic共转染H1299细胞,Western blotting检测结果显示,与mimic-NC组相比,miR-21 mimic组PTEN表达降低,p-β-catenin(Ser552、Ser675)和β-catenin表达升高;与miR-21 mimic组相比,miR-21 mimic+H1299/PTEN组PTEN表达升高,p-β-catenin(Ser552、Ser675)和β-catenin水平降低。流式细胞术检测结果显示,mimic-NC组细胞经过CDDP处理后细胞凋亡率显著升高(P<0.05)。使用H1299构建的荷瘤裸�Objective To investigate the role and mechanism of microRNA-21-5p(miR-21)in cisplatin(CDDP)resistance in non-small cell lung cancer.Methods The expression of miR-21 in cancer tissues and paracancerous tissues of non-small cell lung cancer patients was detected by real-time fluorescence quantitative PCR.Non-small cell lung cancer CDDP-resistant cells H1299/CR and H1975/CR were constructed using non-small cell lung cancer cell lines H1299 and H1975.CCK-8 was used to detect the cell activity of each group of cells under CDDP treatment,and apoptosis was analyzed by flow cytometry.Dual luciferase was used to detect the role of miR-21 in relation to PTEN in H1299 cells,RT-qPCR was used to detect the miR-21 level,and Western blotting was used to detect the protein expression level of PTEN and PTEN downstream Wnt/β-catenin signaling pathway.H1299 and H1299/CR were used to construct a hormonal nude mouse model to verify the effect of miR-21 on the sensitivity of CDDP treatment in non-small cell lung cancer.Results The expression of miR-21 was significantly higher in cancer tissues than in adjacent normal tissues(P<0.001).The expression of miR-21 in drug-resistant cells H1299/CR and H1975/CR was significantly elevated compared to that in H1299 and H1975 cells(P<0.05).After transfection with miR-21 inhibitor,cell viability in the inhibitor group was significantly lower than in the inhibitor NC group when treated with CDDP≥12μmol/L(P<0.001).Flow cytometry analysis showed that while the apoptosis rate in the CDDP and miR-21 inhibitor groups did not significantly differ from that in the untreated group,the apoptosis rate in the CDDP+miR-21 inhibitor group was significantly higher(P<0.001).In contrast,after transfection with miR-21 mimic,the cell viability of the mimic-NC group was significantly reduced compared to the miR-21 mimic group when treated with CDDP≥12μmol/L(P<0.05).TargetScan predicted that miR-21 could bind to the 3′-UTR region of PTEN.Dual-luciferase reporter assays confirmed that miR-21 directly targeted

关 键 词：MIR-21 非小细胞肺癌 顺铂 顺铂耐药性 PTEN WNT/Β-CATENIN通路 

分 类 号：R734.2[医药卫生—肿瘤]