Institute for Science and Technology in Medicine; Keele University; Guy Hilton Research Centre; Staffordshire, UK; Haywood Rheumatology Centre; Haywood Hospital; Staffordshire, UK.
Haywood Rheumatology Centre; Haywood Hospital; Staffordshire, UK.
Epigenetics. 2013 Nov;8(11):1188-97. doi: 10.4161/epi.26265. Epub 2013 Sep 4.
Multiple reports now describe changes to the DNA methylome in rheumatoid arthritis and in many cases have analyzed methylation in mixed cell populations from whole blood. However, these approaches may preclude the identification of cell type-specific methylation, which may subsequently bias identification of disease-specific changes. To address this possibility, we conducted genome-wide DNA methylation profiling using HumanMethylation450 BeadChips to identify differences within matched pairs of T-lymphocytes and B-lymphocytes isolated from the peripheral blood of 10 healthy females. Array data were processed and differential methylation identified using NIMBL software. Validation of array data was performed by bisulfite pyrosequencing. Genome-wide DNA methylation was initially determined by analysis of LINE-1 sequences and was higher in B-lymphocytes than matched T-lymphocytes (69.8% vs. 65.2%, P ≤ 0.01). Pairwise analysis identified 679 CpGs, representing 250 genes, which were differentially methylated between T-lymphocytes and B-lymphocytes. The majority of sites (76.6%) were hypermethylated in B-lymphocytes. Pyrosequencing of selected candidates confirmed the array data in all cases. Hierarchical clustering revealed perfect segregation of samples into two distinct clusters based on cell type. Differentially methylated genes showed enrichment for biological functions/pathways associated with leukocytes and T-lymphocytes. Our work for the first time shows that T-lymphocytes and B-lymphocytes possess intrinsic differences in DNA methylation within a restricted set of functionally related genes. These data provide a foundation for investigating DNA methylation in diseases in which these cell types play important and distinct roles.
现在有多项报告描述了类风湿关节炎中 DNA 甲基化组的变化，并且在许多情况下，分析了来自全血的混合细胞群中的甲基化。然而，这些方法可能会排除对细胞类型特异性甲基化的识别，这可能随后会影响对疾病特异性变化的识别。为了解决这个问题，我们使用 HumanMethylation450 BeadChips 进行了全基因组 DNA 甲基化谱分析，以鉴定 10 名健康女性外周血中分离的 T 淋巴细胞和 B 淋巴细胞匹配对之间的差异。使用 NIMBL 软件处理阵列数据并鉴定差异甲基化。通过亚硫酸氢盐焦磷酸测序验证阵列数据。通过分析 LINE-1 序列初步确定了全基因组 DNA 甲基化，B 淋巴细胞中的甲基化程度高于匹配的 T 淋巴细胞（69.8%比 65.2%，P ≤ 0.01）。成对分析鉴定出 679 个 CpG，代表 250 个基因，它们在 T 淋巴细胞和 B 淋巴细胞之间存在差异甲基化。大多数位点（76.6%）在 B 淋巴细胞中呈高甲基化。对选定候选者的焦磷酸测序在所有情况下均证实了阵列数据。层次聚类显示，基于细胞类型，样品完美地分为两个不同的簇。差异甲基化基因显示出与白细胞和 T 淋巴细胞相关的生物学功能/途径的富集。我们的工作首次表明，在一组功能相关基因中，T 淋巴细胞和 B 淋巴细胞具有内在的 DNA 甲基化差异。这些数据为研究这些细胞类型在重要和独特作用的疾病中的 DNA 甲基化提供了基础。
