hnRNPK调控Wnt/β-catenin信号转导通路抑制乳腺癌细胞铁死亡  

hnRNPK regulates Wnt/β-catenin signaling pathway to inhibit ferroptosis in breast cancer

作　　者：马小兰 王娟 石斌 王南 田治翠 曹佳 MA Xiaolan;WANG Juan;SHI Bin;WANG Nan;TIAN Zhicui;CAO Jia(Medical Science Research Institute of General Hospital of Ningxia Medical University,Yinchuan 750004,Ningxia Hui Autonomous Region,China;Department of Surgical OncologyⅡ,General Hospital of Ningxia Medical University,Yinchuan 750004,Ningxia Hui Autonomous Region,China;Department of Pathology,Yinchuan Maternal and Child Health Hospital,Yinchuan 750004,Ningxia Hui Autonomous Region,China;Emergency Department of General Hospital of Ningxia Medical University,Yinchuan 750004,Ningxia Hui Autonomous Region,China;Department of Medical oncologyⅡ,General Hospital of Ningxia Medical University,Yinchuan 750004,Ningxia Hui Autonomous Region,China;Department of Cardiology,Lingwu City People's Hospital,Lingwu 751400,Ningxia Hui Autonomous Region,China)

机构地区：[1]宁夏医科大学总医院医学科学研究院,宁夏银川750004 [2]宁夏医科大学总医院肿瘤外二科,宁夏银川750004 [3]银川市妇幼保健院病理科,宁夏银川750004 [4]宁夏医科大学总医院急诊科,宁夏银川750004 [5]宁夏医科大学总医院肿瘤内二科,宁夏银川750004 [6]灵武市人民医院心血管内科,宁夏灵武751400

出　　处：《中国癌症杂志》2024年第10期931-943,共13页China Oncology

基　　金：宁夏医科大学2024年校级科研项目(XY2024005);宁夏回族自治区重点研发计划项目(2022BEG03124)。

摘　　要：背景与目的:核内不均一核糖核蛋白K(heterogeneous nuclear ribonucleoprotein K,hnRNPK)是一种调控基因表达和蛋白翻译的RNA结合蛋白,已被发现与多种肿瘤恶性进展密切相关,但其在乳腺癌中的作用尚未明确。本研究旨在探讨hnRNPK对乳腺癌细胞铁死亡的影响及其作用机制。方法:基于癌症基因组图谱(The Cancer Genome Atlas,TCGA)和基因表达综合(Gene Expression Omnibus,GEO)数据库,联合生信分析hnRNPK在乳腺癌组织及正常组织中的表达及与临床预后的关系;采用实时定量逆转录聚合酶链反应(real-time quantitative reverse transcription polymerase chain reaction,qRT-PCR)、蛋白质印迹法(Western blot)、免疫组织化学法检测hnRNPK在乳腺癌细胞及组织中的表达;将乳腺癌MDA-MB-231和MCF-7细胞进行siRNA转染,设置分组为对照组(control)、空载体组(NC)、干扰载体组(si-hnRNPK),采用细胞计数试剂盒(cell counting kit-8,CCK-8)和平板克隆实验检测细胞增殖能力;高通量RNA测序(RNA-sequencing,RNA-seq)分析hnRNPK富集的生物学功能及信号转导通路,Western blot及活性氧(reactive oxygen species,ROS)、丙二醛(malondialdehyde,MDA)、谷胱甘肽(glutathione,GSH)、Fe2+试剂盒检测hnRNPK对铁死亡表型的影响,并利用铁死亡抑制剂(ferrostatin-1,Fer-1)检测敲低hnRNPK对铁死亡的回复作用;通过Western blot检测hnRNPK对Wnt/β-catenin通路蛋白表达的影响。结果:数据库结合生物信息学分析结果显示,hnRNPK在乳腺癌组织中表达上调(P<0.01),hnRNPK高表达组患者总生存期OS低于hnRNPK低表达组(P<0.05);hnRNPK在乳腺癌组织及细胞中呈高表达,敲低hnRNPK后乳腺癌细胞增殖能力减弱(P<0.05);RNA-seq分析发现hnRNPK显著富集铁死亡、凋亡、Wnt/β-catenin信号转导通路;敲低hnRNPK通过促进脂质ROS和MDA产生、Fe2+富集促进乳腺癌细胞铁死亡(P<0.05),且联合Fer-1有效抑制敲低hnRNPK对铁死亡的促进作用(P<0.05);hnRNPK下调导致Wnt/β-catBackground and purpose:Heterogeneous nuclear ribonucleoprotein K(hnRNPK)is an RNA special binding protein that participates in regulating the expression of related genes and protein translation.It has been linked to the malignant occurrence and development of various tumors,but its role in breast cancer remains unclear.The aim of this study was to investigate the effects of hnRNPK on ferroptosis in breast cancer cells and the underlying mechanisms.Methods:Based on The Cancer Genome Atlas(TCGA)and Gene Expression Omnibus(GEO)databases,hnRNPK expression in breast cancer tissues and normal tissues and its relationship with clinical prognosis were analyzed by bioinformatics.We detected hnRNPK expression in breast cancer cells and tissues using real-time quantitative reverse transcription polymerase chain reaction(qRT-PCR),Western blot,and immunohistochemistry staining diagnosis methods.MCF-7 and MDA-MB-231 breast cancer cells were transfected with siRNA,and divided into control group(control),empty body group(NC),and interference vector group(si-hnRNPK).Cell proliferation was detected by cell counting kit-8(CCK-8)and plate clone formation assays.RNA-seq analysis was applied to explore potential targeted biological functions and signaling pathways affected by hnRNPK.Additionally,we investigated the impact of hnRNPK on ferroptosis phenotype using Western blot and commercial kits for reactive oxygen species(ROS),malondialdehyde(MDA),glutathione(GSH),and Fe2+,ferroptosis inhibitor(ferrostatin-1,Fer-1)was used to detect the rescue effect of hnRNPK knockdown on ferroptosis.The impact of hnRNPK on the expressions of Wnt/β-catenin pathway-related proteins were determined by Western blot.Results:The bioinformatics analyses indicated hnRNPK was upregulated in breast cancer tissues(P<0.01),and the overall survival of patients in the high expression group was poorer compared with those in the low expression group(P<0.05).hnRNPK was highly expressed in breast cancer tissues and cells,and knocking down hnRNPK weakened the prolife

关 键 词：hnRNPK 乳腺癌 铁死亡 WNT/Β-CATENIN信号转导通路 

分 类 号：R735.3[医药卫生—肿瘤]