CMTM3通过NF-κB信号通路调控胶质母细胞瘤U251细胞增殖和迁移  

CMTM3 regulates proliferation and migration of glioblastoma U251 cells via the NF-κB signaling pathway

作　　者：姜利时 滕伟 仇文进 凌园果 石学平 龙妮娅 出良钊 刘健 Jiang Lishi;Teng Wei;Qiu Wenjin;Ling Yuanguo;Shi Xueping;Long Niya;Chu Liangzhao;Liu Jian(Clinical Medical College of Guizhou Medical University,Guiyang 550000,China;Department of Neurosurgery,Affiliated Hospital of Guizhou Medical University,Guiyang 550000,China)

机构地区：[1]贵州医科大学临床医学院,贵阳550000 [2]贵州医科大学附属医院神经外科,贵阳550000

出　　处：《中华肿瘤杂志》2025年第2期136-148,共13页Chinese Journal of Oncology

基　　金：贵州省科技计划项目(2016-2905);贵州省卫生健康委员会科学技术基金(gzwkj2022090)。

摘　　要：目的探讨趋化素样因子样含MARVEL跨膜结构域3(CMTM3)对胶质母细胞瘤(GBM)细胞增殖和迁移的影响及可能机制。方法利用癌症基因组图谱(TCGA)和基因型-组织表达(GTEx)数据库中的CMTM3表达数据,分析CMTM3在GBM组织中的差异表达及其对GBM患者预后的影响。通过对取自2022年11月3日至2023年3月15日贵州医科大学附属医院手术治疗的11例GBM患者的GBM组织和癌旁组织样本进行CMTM3的免疫组化染色和蛋白含量测定、对GBM细胞系U87、U251、LN229和人胶质细胞系NHA进行CMTM3的实时荧光定量聚合酶链反应(RT-qPCR)、Western blot检测加以验证。利用U251细胞构建沉默和过表达CMTM3的稳转细胞株(sh-CMTM3组和OE-CMTM3组),采用细胞计数试剂盒8(CCK-8)实验检测CMTM3表达对细胞增殖能力的影响,采用流式细胞术检测CMTM3表达对细胞周期的影响,采用Transwell实验检测CMTM3表达对细胞迁移能力的影响,采用生信分析方法、Western blot检测、核因子κB(NF-κB)激活-核转运实验以及NF-κB通路抑制剂吡咯烷二硫代甲酸铵(PDTC)来验证CMTM3对NF-κB通路的影响。最后通过裸鼠皮下成瘤实验观察CMTM3表达对U251细胞在体内生长的影响。结果生物信息学分析显示,CMTM3在GBM组织中高表达,CMTM3高表达组GBM患者的总生存率和无病生存率均低于CMTM3低表达组(P值分别为0.010和0.032)。11例GBM组织样本中,有10例CMTM3蛋白的表达量分别高于相应患者的癌旁组织,这10例GBM组织样本中CMTM3蛋白的表达量总体为0.44±0.09,高于癌旁组织中CMTM3蛋白的表达量(0.12±0.02;t=6.69,P<0.001)。只有1例GBM组织组织中CMTM3蛋白的表达量为0.15±0.01,其相应癌旁组织中CMTM3蛋白的表达量为0.17±0.08,差异无统计学意义(t=0.32,P=0.750)。RT-qPCR检测结果显示,人胶质细胞NHA中CMTM3 mRNA的表达量为1.0±0.1,GBM细胞U87、LN229和U251中CMTM3 mRNA的表达量分别为2.1±0.3、3.4±0.5和3.7±0.8,均高于NHA细胞(1.0�Objective To explore the effects and potential mechanisms of chemokine-like factor-like MARVEL transmembrane domain-containing Protein 3(CMTM3)on the proliferation and migration of glioblastoma(GBM)cells.Methods Using CMTM3 expression data from The Cancer Genome Atlas(TCGA)and Genotype-Tissue Expression(GTEx)databases,we analyzed the differential expression of CMTM3 in GBM tissues and its impact on the prognosis of GBM patients.Immunohistochemical staining and protein content determination of CMTM3 was performed on GBM and adjacent non-cancerous tissue samples from 11 GBM patients who underwent surgical treatment at the Affiliated Hospital of Guizhou Medical University between November 3,2022 and March 15,2023.Additionally,the expression of CMTM3 was validated in GBM cell lines U87,U251,LN229,and the human astrocyte(NHA)cell line using real-time quantitative polymerase chain reaction(RT-qPCR)and Western blot analyses.Stable cell lines with silenced and overexpressed CMTM3(sh-CMTM3 group and OE-CMTM3 group)were constructed using U251 cells.The effect of CMTM3 expression on cell proliferation was assessed using the Cell Counting Kit-8(CCK-8)assay.Flow cytometry was employed to examine the impact of CMTM3 expression on the cell cycle.Transwell assays were conducted to evaluate the influence of CMTM3 expression on cell migration.Bioinformatics analysis,Western blotting,NF-κB activation-nuclear translocation assays,and the NF-κB pathway inhibitor pyrrolidine dithiocarbamate ammonium(PDTC)were used to validate the effect of CMTM3 on the NF-κB pathway.Finally,a subcutaneous tumorigenesis assay in nude mice was performed to observe the impact of CMTM3 expression on the in vivo growth of U251 cells.Results Bioinformatics analysis revealed that CMTM3 is highly expressed in GBM tissues.Patients with a high CMTM3 expression had lower overall survival(OS)and disease-free survival(DFS)rates compared with those with a low CMTM3 expression(with P values of 0.010 and 0.032,respectively).Among the 11 GBM pathological specimens,

关 键 词：胶质母细胞瘤 趋化素样因子样含MARVEL跨膜结构域3 核因子ΚB 细胞增殖 细胞迁移 

分 类 号：R73[医药卫生—肿瘤]