造血系细胞特异蛋白1基因在脑胶质瘤中的表达及其意义

The expression and clinical significance of hematopoietic cell specific protein-1 gene in gliomas

目的分析造血系细胞特异蛋白1(HCLS1)基因在脑胶质瘤中的表达特征及其意义。方法收集中国脑胶质瘤基因组图谱(CGGA)计划数据库的两套mRNA测序数据及其临床资料(CGGA325数据集325例,CGGA693数据集693例)。分析HCLS1在不同世界卫生组织(WHO)病理学分级、异柠檬酸脱氢酶(IDH)突变状态、2016年中枢神经系统肿瘤分类、O6-甲基鸟嘌呤DNA甲基转移酶(MGMT)启动子甲基化状态的脑胶质瘤患者中的表达特征。根据HCLS1表达量的中位数,将患者分为高表达组和低表达组,绘制Kaplan-Meier生存曲线,采用log-rank检验比较不同HCLS1表达水平患者的生存差异。采用单因素和多因素Cox回归分析判断脑胶质瘤患者总生存期的影响因素。收集2007年1月至2015年10月首都医科大学附属北京天坛医院神经外科收治的脑胶质瘤患者的脑胶质瘤样本和生存信息(实验组1共27例,实验组2共17例),实验组1采用实时荧光定量聚合酶链式反应(qRT-PCR)检测HCLS1的表达量,实验组2采用免疫组织化学染色检测HCLS1蛋白的表达情况,并计算免疫组织化学染色评分。采用生物信息学方法分析HCLS1的生物学功能。结果在CGGA325数据集中,HCLS1的表达量随WHO病理学级别(Ⅱ~Ⅳ级)的升高而增加(F=30.29,P<0.001);HCLS1在IDH野生型中的表达量高于IDH突变型(t=8.51,P<0.001);依据2016年的中枢神经系统肿瘤分类,3组低级别脑胶质瘤患者中,HCLS1表达量在IDH突变+1p/19q联合缺失组最低(F=23.61,P<0.001),而两组高级别脑胶质瘤患者中,HCLS1在IDH野生组的表达高于IDH突变组(P<0.001);MGMT启动子非甲基化者的HCLS1表达量高于甲基化者(P=0.001);HCLS1高表达组患者的总生存期短于低表达组(P<0.001)。HCLS1的上述表达特征在CGGA693数据集中得到了验证。多因素Cox回归分析结果显示,HCLS1表达量是影响脑胶质瘤患者总生存期的独立危险因素(HR=1.951,95%CI:1.307~2.910,P=0.001)。qRT-PCR结果显示,HCLS1的表达量随WHO病理学级别的升高而增加(F=11.11,P<0.001)。免疫组织化学染色评分结果显示,HCLS1蛋白在WHOⅡ~Ⅳ级组织样本中的评分依次为(1.33±0.21)分、(3.40±0.87)分及(7.67±0.88)分,组间差异有统计学意义(F=21.88,P<0.001)。实验组1和2中,HCLS1基因和蛋白高表达患者的总生存期均短于低表达者(均P<0.05)。生物信息学分析提示,HCLS1可能与脑胶质瘤细胞的免疫反应有关。结论HCLS1的表达量与脑胶质瘤的恶性程度及患者的生存期有关,可作为判断患者预后的潜在指标。

Objective To analyze the expression characteristics and clinical significance of hematopoietic cell specific protein-1(HCLS1)gene in gliomas.Methods Two mRNA sequencing datasets and clinical data were collected from the Chinese Glioma Genome Atlas(CGGA)(325 cases in the CGGA325 dataset,693 cases in the CGGA693 dataset).The expression characteristics of HCLS1 were analyzed in different World Health Organization(WHO)pathological grades,isocitrate dehydrogenase(IDH)mutation status,2016 central nervous system tumor classification,O6-methylguanine DNA methyltransferase(MGMT)promoter status.Using the median HCLS1 expression level as a cutoff,patients were divided into high and low expression groups.Kaplan-Meier survival analysis and log-rank test were used to compare the survival significance between stratified groups.Univariate and multivariate Cox regression analysis were used to assess prognostic factors for overall survival of patients with glioma.Glioma samples and survival information were collected from glioma patients admitted to Department of Neurosurgery,Beijing Tiantan Hospital,Capital Medical University from January 2007 to October 2015(27 cases in group 1 and 17 cases in group 2).Quantitative real-time fluorescent polymerase chain reaction(qRT-PCR)was used to detect the expression of HCLS1 in group 1.Immunohistochemical staining was used to detect the expression of HCLS1 protein in group 2,and Immunohistochemical staining scores were calculated.Bioinformatics methods were used to analyze the biological functions of HCLS1.Results In the CGGA325 dataset,the expression of HCLS1 increased along with grade progression(F=30.29,P<0.001);the expression level of HCLS1 in the IDH wildtype group was significantly higher than that in the IDH mutant group(t=8.51,P<0.001).In the 2016 classification of central nervous system tumors,HCLS1 expression was the lowest in lower grade gliomas with IDH mutation and 1p/19q codeletion among three lower grade groups(F=23.61,P<0.001),while HCLS1 expression was higher in high grade gliomas with IDH wildtype(P<0.001);the expression of HCLS1 was higher in the MGMT promoter unmethylated group(P=0.001).Patients who had higher HCLS1 expression had a significantly shorter survival time than those who had lower HCLS1 expression(P<0.001).Similar results were shown in the CGGA693 dataset.Multivariate Cox regression analysis showed that the expression of HCLS1 was an independent hazard factor affecting the overall survival of patients with glioma(HR=1.951,95%CI:1.307-2.910,P=0.001).The results of qRT-PCR confirmed that the expression of HCLS1 increased along with grade progression(F=11.11,P<0.001).Immunohistochemical staining results showed that the scores of HCLS1 protein in WHOⅡ-Ⅳgrade samples were 1.33±0.21,3.40±0.87,7.67±0.88,and the difference among groups was statistically significant(F=21.88,P<0.001).The overall survival of patients with high HCLS1 gene and protein expression were shorter than that of patients with low HCLS1 gene and protein expression in group 1 and 2(both P<0.05).HCLS1 might be related to the immune response of glioma cells based on the bioinformatics analysis.Conclusion The expression level of HCLS1 is related to the malignant degree of glioma and the survival of the patient,and can be used as a potential indicator for judging the prognosis of the patient.