TrxR1基因在脑胶质瘤组织中的表达及其对胶质瘤细胞株U87生物学特征的影响  

Expression of TrxR1 gene in glioma tissues and its influence on the biological characteristics of glioma cell line U87

作　　者：王乐[1] 钟天佑 南阳[1] 任炳成[1] 范志娟[3] 赵立文 钟跃[1] Wang Le;Zhong Tianyou;Nan Yang;Ren Bingcheng;Fan Zhijuan;Zhao Liwen;Zhong Yue(Department ofNeurosurgery,Tianjin Medical University General Hospital,Tianjin 300052,China;Department of Library,Tianjin Medical University General Hospital,Tianjin 300052,China;Department of Laboratory,Tianjin Third Central Hospital,Tianjin 300170,China)

机构地区：[1]天津医科大学总医院神经外科,天津300052 [2]天津医科大学总医院图书馆,天津300052 [3]天津市第三中心医院检验科,天津300170

出　　处：《中华神经外科杂志》2022年第12期1254-1260,共7页Chinese Journal of Neurosurgery

基　　金：国家自然科学基金(81702678);天津市滨海新区卫生和计划生育委员会科技项目(2018BWKY010);天津市卫生健康委员会科技基金(KJ20151)。

摘　　要：目的探究硫氧还蛋白还原酶1(TrxR1)基因在脑胶质瘤组织中的表达情况以及对胶质瘤细胞株U87生物学特征的影响.方法回顾性分析中国脑胶质瘤基因组图谱计划(CGGA)数据库中325例脑胶质瘤患者的胶质瘤样本的转录组数据和临床资料,并利用蛋白质免疫印迹(WB)、实时荧光定量PCR(RT-PCR)和免疫组织化学技术进一步检测TrxR1在正常脑组织及胶质瘤组织中的表达情况.组织标本取自于2010年9月至2018年11月于天津医科大学总医院神经外科行手术治疗的脑胶质瘤和自发性脑出血患者,其中胶质瘤组织标本12例和正常脑组织标本12例.培养胶质瘤细胞株U87,将其分为对照组、无义序列组和siRNA TrxR1转染组(每组n=3).采用RT-PCR、WB和免疫组织化学染色方法检测TrxR1基因的表达情况.采用MTT、Transwell及细胞成球实验检测胶质瘤细胞株U87的增殖活性、侵袭能力及胶质瘤干细胞的致瘤性.采用WB检测敲低TrxR1表达水平后对STAT3表达的影响,以及对STAT3信号通路下游基因表达的影响.结果对CGGA数据库分析显示,TrxR1 mRNA的表达水平随着胶质瘤病理级别的升高而升高(P<0.01),并且高表达组患者的总生存期低于低表达组(P=0.019).WB、RT-PCR和免疫组织化学检测显示,与正常脑组织比较,TrxR1在胶质瘤组织中的表达水平均明显升高(均P<0.01).siRNA TrxR1可有效敲低TrxR1基因的表达,与对照组、无义序列组比较,siRNA TrxR1转染组的TrxR1表达水平在蛋白和mRNA水平中均明显降低(均P<0.05).MTI检测结果显示,敲低TrxR1的表达后,胶质瘤细胞生长明显受到抑制,培养第4天的细胞增殖率仅为对照组的39.3%.Transwell实验结果显示,siRNA TrxR1转染组穿过小室膜的细胞数低于对照组和无义序列组(均P<0.01),仅为对照组的47.0%.细胞成球实验结果显示,与对照组、无义序列组比较,TrxR1 siRNA转染组的成球率均明显降低(均P<0.01).WB检测结果显示,与对照ObjectiveTo investigate the expression of TrxRl gene in glioma tissue and its effect on biological characteristics of glioma cell line U87.MethodsTranscriptome and clinical data of glioma samples from 325glioma patients in the Chinese Glioma Genome Atlas Project(CGGA)database were retrospectively analyzed,and the expression of TrxR1 in normal brain tissue and glioma tissue was further detected by western blot,RT-PCR and immunohistochemical techniques.A total of 12 cases of glioma and 12 cases of normal brain tissues were collected from patients with glioma and spontaneous intracerebral hemorrhage who underwent surgical treatment at the Neurosurgery Department of Tianjin Medical University General Hospital from September 2010 to November 2018.Glioma cell line U87 was cultured and divided into control,nonsense sequence and siRNA TrxRl groups.The expression of TrxRl gene was detected by RTPCR,Western blot and immunohistochemistry.MTT and Transwell assays were used to detect the proliferation and invasion of glioma cell line U87.The spheroidization assay was used to detect the tumorigenicity of U87 cell line.Western blot was used to detect the effect of TrxR1 knockdown on STAT3 expression and the STAT3 signaling pathway related genes.ResultsAnalysis of the CGGA database showed that the expression of TrxR1 mRNA increased with the increase of the pathological grade(WHO grade)of glioma(P<0.01),and the overall survival of patients in the high expression group was lower than that in the low expression group(P=0.O19).Western blot,RT-PCR and immunohistochemical experiments showed that compared with normal brain tissue,the expression of TrxR1 in glioma tissue was significantly increased(all P<0.01).The siRNA TrxRl could effectively downregulate TrxR1 gene expression.Compared with the control and nonsense sequence groups,siRNA TrxRl group significantly reduced TrxR1 expression at both protein and mRNA levels(P<O.05).The results of MTT showed that the growth of glioma cells was significantly inhibited by knockdown of TrxR1 exp

关 键 词：神经胶质瘤 基因表达 硫氧还蛋白还原酶1 细胞增殖 肿瘤侵润 信号转导和转录激活因子3 

分 类 号：R739.41[医药卫生—肿瘤]