免疫相关性血细胞减少症患者外周血CD4＋T细胞DNA甲基化异常的分析  

Abnormal DNA methylation in CD4 ＋ T cells from patients with immune related pancytopenia： a preliminary study

作　　者：任悦[1] 付蓉[1] 王化泉[1] 刘惠[1] 王一浩[1] 齐薇薇[1] 陶景莲[1] 邵宗鸿[1] 

机构地区：[1]天津医科大学总医院血液科,300052

出　　处：《中华医学杂志》2014年第28期2165-2168,共4页National Medical Journal of China

基　　金：国家自然科学基金（81170472、81370607）;天津市自然科学基金（14JCYBJC25400）;天津市抗癌重大专项攻关计划（12ZCDZSY17900、12ZCDZSY18000）;天津市卫生行业重点攻关项目（11KG135）;天津市卫生行业科研专项（201202017）

摘　　要：目的 观察免疫相关性血细胞减少症（IRP）患者CD4＋T细胞基因组DNA总体甲基化水平及甲基化相关调控基因的表达水平,探讨甲基化在IRP发病中的作用.方法 选取2012年12月至2013年12月天津医科大学总医院血液科确诊的30例IRP患者（15例初治患者,15例恢复患者）及15名健康对照,以免疫磁珠法分选CD4＋T细胞,应用ELISA法测定外周血CD4＋T细胞DNA总体甲基化水平;采用反转录PCR （RT-PCR）技术检测CD4＋T细胞中甲基化相关调控基因：DNA甲基化转移酶（DNMT）和甲基结合蛋白（MBD）的mRNA表达水平,并进行相关性分析.结果 IRP患者外周血CD4＋T细胞DNA总体甲基化水平初治组和恢复组均明显低于对照组（3.525％±2.046％和4.790％±1.471％比10.101％±3.449％,均P＜0.05）;初治组DNMT3b mRNA表达水平显著低于对照组（1.332±0.785比2.077±1.059,P＜0.05）;初治组、恢复组MBD2 mRNA表达水平显著高于对照组（2.999±1.601和2.055±1.576比0.581±0.247,均P＜0.05）;初治组MBD4 mRNA表达水平显著高于对照组（2.736±2.719比1.167±1.006,P＜0.05）.IRP患者外周血CD4＋T细胞DNMT3b mRNA表达水平与DNA总体甲基化水平呈正相关（r=0.569,P＜0.01）;MBD2 mRNA表达水平与CD4＋T细胞基因组DNA甲基化水平和Th1/Th2水平均呈负相关（r=-0.763,P＜0.01;r=-0.652,P＜0.05）.CD4＋T细胞甲基化水平与CD5＋B细胞比例呈负相关（r=-0.439,P ＜0.05）.结论 IRP患者外周血CD4＋T细胞基因组DNA低甲基化和甲基化相关调控基因表达异常可能与IRP发病有关.Objective To explore the global DNA methylation and the expression of regulatory genes for methylation in CD4 ＋ T cells of the patients with immune related pancytopenia （IRP） and explore the role of methylation in the pathogenesis of IRP.Methods Thirty IRP patients （untreated,n =15 ;remission,n =15） and 15 healthy donors as controls were enrolled from December 2012 to December 2013.CD4 ＋ T cells were sorted by immunomagnetic separation.The global DNA methylation was tested with enzyme-linked immunosorbent assay （ELISA）.The mRNA levels of DNA methylation-related regulating genes,DNA methyltransferases （DNMTs） and methylated CpG binding proteins （MBDs）,were measured by real-time quantitative-polymerase chain reaction （RT-PCR）.Results The level of global DNA methylation in peripheral blood CD4 ＋ T cells of untreated IRP patients （3.525％ ± 2.046％） and remission patients （4.790％ ± 1.471％）were significantly lower than that of normal controls （10.101％ ± 3.449％） respectively （both P ＜ 0.05）.DNMT3b mRNA level of untreated IRP patients （1.332 ± 0.785） wassignificantly lower than that of normal controls （2.077 ± 1.059） in CD4 ＋ T cells （P ＜ 0.05）.The mRNA expression of MBD2 was significantly higher in CD4 ＋ T cells from untreated and remission IRP patients （2.999 ± 1.601,2.055 ± 1.576） than that in controls（0.581 ±0.247） （both P ＜0.05）.The MBD4 mRNA level was significantly higher in CD4 ＋ T cells from untreated IRP patients （2.736 ± 2.719） compared to that in normal controls （1.167 ± 1.006） （P ＜ 0.05）.DNMT3b mRNA expression and CD4 ＋ T cell DNA methylation to be positive correlated within IRP patients （r =0.569,P ＜ 0.01）.The MBD2 mRNA expression was negatively correlated with CD4 ＋ T cell DNA methylation and the ratio of Th1/Th2 （r =-0.763,P ＜ 0.01; r =-0.652,P ＜ 0.05）.The global methylation of CD4 ＋ T cells negatively related to the ratio of CD5 ＋ B cells （r =-0.439,P ＜ 0.05）.Con

关 键 词：全血细胞减少 DNA甲基化 免疫相关性血细胞减少症 DNA甲基转移酶 甲基结合蛋白 

分 类 号：R55[医药卫生—血液循环系统疾病]