ANGPTL2基因启动子区甲基化在骨髓增生异常综合征诊断中的价值

The value of ANGPTL2 gene promoter methylation in the diagnosis of myelodysplastic syndromes

目的探讨骨髓增生异常综合征(MDS)中血管生成素样蛋白2(ANGPTL2)启动子区甲基化状态,以期为MDS的早期诊断提供新的潜在分子标记物。方法收集2018年10月至2023年8月55例MDS患者、14例急性髓系白血病(AML)患者和28例对照者(18例健康供者、6例缺铁性贫血、4例营养性巨幼细胞性贫血)的骨髓血标本,提取DNA和RNA。采用Agena MassARRAY核酸质谱技术,对3组DNA样本进行该基因启动子区甲基化的定量检测,通过实时荧光定量PCR技术检测mRNA的相对表达量。比较各组ANGPTL2基因的甲基化水平和mRNA表达量,采用受试者工作特征(ROC)曲线评估CpG单位诊断的特异度和灵敏度。结果MDS组和AML组ANGPTL2基因启动子区甲基化率分别为10.0%(3.0%,26.0%)、10.5%(1.0%,31.3%),均高于对照组的6.0%(2.0%,15.0%)。对照组ANGPTL2基因mRNA相对表达量显著高于MDS组和AML组。进一步分析不同疾病组CpG单位的甲基化率差异,MDS组与对照组的6个CpG单位、AML组与对照组的4个CpG单位以及MDS组与AML组的3个CpG单位的甲基化率存在明显差异。CpG-20对MDS和AML诊断的灵敏度分别为67.4%和64.6%,特异度分别为100.0%和69.9%,ROC曲线下面积(AUC)最大,分别为0.8483(95%CI:0.7544~0.9421)和0.9063(95%CI:0.7855~1.0000)。结论MDS患者骨髓标本的ANGPTL2基因启动子区存在高甲基化改变,对MDS的分子诊断有重要价值,可作为MDS潜在的早期鉴别诊断标记物。

Objective To investigate the methylation status of the angiopoietin-like protein 2(ANGPTL2)promoter region in myelodysplastic syndrome(MDS)and provide potential molecular markers for early diagnosis of MDS.Methods DNA and RNA were extracted from bone marrow samples of 55 MDS patients from Oct 2018 to Aug 2023,14 acute myeloid leukemia(AML)patients,and 28 controls(18 healthy donors,6 iron-deficiency anemia,and 4 megaloblastic anemia).Agena MassARRAY nucleic acid mass spectrometry was used to quantitatively detect methylation of the gene promoter region in the DNA samples,and real-time quantitative PCR was used to detect relative mRNA expression levels.The methylation level and mRNA expression of the ANGPTL2 gene were compared among different groups,and receiver operating characteristic(ROC)curves were used to evaluate the specificity and sensitivity of CpG unit diagnosis.Results The methylation rates of the ANGPTL2 gene promoter region were 10.0%(3.0%,26.0%)in the MDS group and 10.5%(1.0%,31.3%)in the AML group,which were both higher than that in the control group 6.0%(2.0%,15.0%).The relative mRNA expression of the ANGPTL2 gene was significantly higher in the control group than that in the MDS and AML groups.Further analysis showed that there were significant differences in the methylation rates of six CpG units in the MDS group and four CpG units in the AML group when compared with the control group,as well as three CpG units in the comparison between MDS and AML groups.CpG-20 had a sensitivity of 67.4%and 64.6%,and a specificity of 100.0%and 69.9%for MDS and AML diagnosis,respectively.The area under the ROC curve(AUC)was highest for CpG-20,with values of 0.8483(95%CI:0.7544-0.9421)and 0.9063(95%CI:0.7855-1.0000)for MDS and AML,respectively.Conclusion The ANGPTL2 gene promoter region is highly methylated in bone marrow samples of MDS patients,and has important value for molecular diagnosis and potential early identification of MDS.