人胶质瘤中HIF1A和MAGED4的表达及调控关系初探

A preliminary study on the expressions and regulatory relationships of HIF1A and MAGED4 in human glioma

目的探讨缺氧诱导因子1A(HIF1A)和黑色素瘤相关抗原D4(MAGED4)在人胶质瘤中的表达情况,并分析HIF1A对MAGED4表达的影响。方法通过基因表达谱交互式分析(GEPIA)数据库收集人胶质瘤组织样本数据681例,正常脑组织样本数据207例,比较其MAGED4 mRNA和HIF1A mRNA表达水平。通过中国胶质瘤基因组图谱(CGGA)数据库中下载693例胶质瘤患者的mRNA-seq_693数据集及相应的临床信息,分析MAGED4 mRNA和HIF1A mRNA表达的相关性,以及二者与患者临床特征指标的关联性。应用氯化钴模拟体外缺氧环境,通过实时荧光定量聚合酶链式反应(RT-qPCR)和蛋白免疫印记(WB)实验探讨沉默胶质瘤细胞HIF1A表达对MAGED4表达变化的影响。结果基于GEPIA数据库的分析结果显示,低级别胶质瘤(LGG)和胶质母细胞瘤(GBM)组织的HIF1A mRNA和MAGED4 mRNA水平均高于正常脑组织,差异有统计学意义(P<0.05)。基于CGGA数据库的分析结果显示,MAGED4 mRNA表达水平与HIF1A mRNA表达水平呈正相关(r=0.322,P=0.000)。MAGED4 mRNA高表达组年龄<40岁、异柠檬酸脱氢酶(IDH)突变型的人数比例大于MAGED4 mRNA低表达组,差异有统计学意义(P<0.05),两组性别、肿瘤类型、世界卫生组织(WHO)肿瘤分级,O6-甲基鸟瞟-DNA甲基转移酶(MGMT)启动子甲基化状态、1p/19q共缺失情况比较差异无统计学意义(P>0.05)。将HIF1A siRNA转染至U87-MG和U251细胞,并在缺氧条件下培养48 h,RT-qPCR和WB实验结果显示,HIF1A下调后,MAGED4的表达水平也显著下降。结论HIF1A和MAGED4均高表达于胶质瘤,且两者呈正相关。胶质瘤细胞中HIF1A可调控MAGED4的表达。

Objective To investigate the expressions of hypoxia-inducible factor 1A(HIF1A)and melanoma-associated antigen D4(MAGED4)in human glioma,and to analyze the effect of HIF1A on the expression of MAGED4.Methods Gene Expression Profiling Interactive Analysis(GEPIA)database was used to collect the data from 681 human glioma samples and the data from 207 normal brain tissue samples to compare the expression levels of MAGED4 mRNA and HIF1A mRNA.The mRNA-seq_693 data set and the corresponding clinical information of 693 glioma patients were downloaded from Chinese Glioma Genome Atlas(CGGA)database.The correlation between the expressions of MAGED4 mRNA and HIF1A mRNA,and the correlation between the two expressions and the clinical characteristics of the patients were analyzed.The hypoxic environment in vitro was simulated by using cobalt chloride,and the effect of silencing HIF1A expression in glioma cells on the expression of MAGED4 was investigated by reverse transcription-quantitative real-time polymerase chain reaction(RT-qPCR)and Western blot(WB)experiments.Results The analysis results based on the GEPIA database showed that the levels of HIF1A mRNA and MAGED4 mRNA in the low-grade glioma(LGG)and the glioblastoma(GBM)tissues were higher than those in the normal brain tissues,and the differences were statistically significant(P<0.05).The analysis results based on the CGGA database showed that the MAGED4 mRNA expression level was positively correlated with the HIF1A mRNA expression level(r=0.322,P=0.000).The proportion of the patients with age<40 years and isocitrate dehydrogenase(IDH)mutation in the high MAGED4 mRNA expression group was greater than that in the low MAGED4 mRNA expression group,and the difference was statistically significant(P<0.05).There were no significant differences in gender,tumor type,World Health Organization(WHO)tumor grade,O6-methylguanie-DNA methyltransferase(MGMT)promoter methylation status and 1p/19q co-deletion between the two groups(P>0.05).U87-MG and U251 cells were transfected with HIF1A siRNA and were cultured for 48 hours under the condition of hypoxia.The results of RT-qPCR and WB experiments showed that the expression level of MAGED4 also decreased significantly after HIF1A was down-regulated.Conclusion Both HIF1A and MAGED4 are highly expressed in gliomas,and their expressions are positively correlated.HIF1A can regulate the expression of MAGED4 in glioma cells.