实时定量PCR检测骨髓增生异常综合征患者多个基因甲基化水平  

Detection of methylation levels of multi-genes by real-time PCR in patients with myelodysplastic syndrome

作　　者：王玉春[1] 杜欣[1] 耿素霞[1] 李明莹[1] 翁建宇[1] 陆泽生[1] 钟立业[1] 邓程新[1] 赖沛龙[1] 黄欣[1] 

机构地区：[1]广东省人民医院肿瘤中心血液科,广州510632

出　　处：《中华血液学杂志》2011年第4期254-258,共5页Chinese Journal of Hematology

基　　金：“十一五”国家科技支撑计划项目（2008BA161802）;广东省中医药局项目（2009116）

摘　　要：目的 了解骨髓增生异常综合征（MDS）患者p15、CDH1、DAPK和HICI基因的甲基化情况,探讨其与临床特征的关系.方法 通过实时荧光定量PCR方法检测52例MDS、38例初诊AML患者和46名正常人骨髓或外周血4种基因的甲基化水平,并分析MDS患者基因的甲基化水平与其临床和实验室特征的关系.结果 MDS患者骨髓p15、CDH1、DAPK和HICI基因的甲基化水平分别为16.23±21.69、6.59±9.39、0.14±0.11和7.81±9.70,外周血为14.96±20.16、6.00±9.26、0.12±0.14和6.74±9.72,骨髓与外周血4种基因甲基化水平差异无统计学意义（P值均＞0.05）.MDS-RAEB Ⅰ/Ⅱ患者骨髓或外周血p15及CDH1基因的甲基化水平（分别为14.70±18.17和6.61±8.79）较MDS-RCUD/RARS/5q-（分别为1.99±1.59,1.23±1.14）、MDS-RCMD（分别为3.02±3.42和1.53±2.06）和正常对照组（分别为1.69±1.82和1.01±1.12）显著升高（P值均＜0.05）;各分型之间DAPK基因甲基化水平差异均无统计学意义;而HIC1基因甲基化水平仅在RAEB Ⅰ/Ⅱ（9.16±11.95）与对照组（2.49±2.26）差异有统计学意义（P=0.042）.不同分型组间骨髓样本4种基因（p15、CDH1、DAPK和HICI）甲基化水平差异有统计学意义（P值分别为0.001、0.003、0.039、0.023）;不同分型组间外周血样本p15及DAPK基因的甲基化水平差异有统计学意义（P值分别为0.013、0.006）.p15基因甲基化水平与IPSS分型、原始细胞百分数、血小板计数有相关性;CDH1基因甲基化水平与IPSS分型、原始细胞百分数有相关性;DAPK基因甲基化水平仅与年龄有相关性;HIC1基因甲基化水平与各项指标均无相关性.结论 p15和CDH1基因是MDS中较为特征性出现的高度甲基化基因,HIC1甲基化水平从低危至高危组有升高趋势,DAPK基因虽然在各分型中均能检测到甲基化,但水平较低,而四种基因的甲基化水平与MDS疗效或预后的关系有待于进一步研究.Objective To analyze the promoter methylation levels of p15、 CDH1、 DAPK and HICI genes of patients with myelodysplastic syndrome （MDS） and explore the relationship between the level of methylation and clinical features. Methods DNA methylation levels of p15、CDH1 、DAPK and HICI in peripheral blood （PB） or bone marrow （BM） samples from 52 MDS patients were detected by real-time quantitative PCR. The correlation of the methylation level with clinical features and hematological findings was analyzed. 38 de novo AML patients and 46 normal individuals served as controls. Results The methylation levels of p15 、CDH1 、DAPK and HICI were 16.23 ± 21.69,6.59 ±9.39,0. 14 ±0. 11 and 7.81 ± 9.70 in BM,and 14.96 ± 20.16,6.00 ± 9.26,0.12 ± 0. 14 and 6.74 ± 9.72 in PB, respectively from 18 MDS patients,and the difference between BM and PB was not statistically significant （P 〉0.05）. The methylation levels of p15 （14.70 ± 18.17） and CDH1 （6.61 ±8.79） genes in high risk （RAEB Ⅰ / Ⅱ） MDS were significantly higher than in low risk （RCMD/RARS/5q -, p15： 1.99 ± 1.59, CDH1： 1.23 ± 1.14 and RCMD, p15：3.02 ±3.42, CDH1：1.53 ±2.06） MDS or control （p15：1.69 ± 1.82, CDH1： 1.01 ± 1.12） （P 〈0.05）.The methylation levels of DAPK gene had no difference among subtypes of MDS, and that of HIC1 gene only differed between RAEB Ⅰ / Ⅱ （9.16 ± 11.95） and control （2.49 ± 2.26） （P = 0. 042）. The difference of methylation levels of p15 、CDH1 、DAPK and HICI in BM was statistically significant among subtypes of MDS （P=0. 001, 0. 003 ,0. 039,0. 023 , respectively）. And so did of p15 and DAPK in PB （P=0. 013, 0.006,respectively）. The methylation level of p15 and CDH1 was significantly correlated with IPSS classification and blasts percentage in BM. Conclusions p15 and CDH1 genes are special hepermethylation genes in MDS.Methylation level of HIC1 gene showed an upward tendency from low risk to high risk MDS.

关 键 词：骨髓增生异常综合征 DNA甲基化 聚合酶链反应 

分 类 号：R551.3[医药卫生—血液循环系统疾病]