Department of Surgery, College of Medicine, The Pennsylvania State University, 500 University Drive, MC H 137, Hershey, PA 17033, USA.
Dig Dis Sci. 2012 Dec;57(12):3145-53. doi: 10.1007/s10620-012-2288-z. Epub 2012 Jul 21.
Changes in the methylation status of inflammatory bowel disease (IBD)-associated genes could significantly alter levels of gene expression, thereby contributing to disease onset and progression. We previously identified seven disease-associated DNA methylation loci from intestinal tissues of IBD patients using the Illumina GoldenGate BeadArray assay.
In this study, we extended this approach to identify IBD-associated changes in DNA methylation in B cells from 18 IBD patients [9 Crohn's disease (CD) and 9 ulcerative colitis (UC)]. B cell DNA methylation markers are particularly favorable for diagnosis due to the convenient access to peripheral blood.
We examined DNA methylation profiles of B cell lines using the Illumina GoldenGate BeadArray assay. Disease-associated CpGs/genes with changes in DNA methylation were identified by comparison of methylation profiles between B cell lines from IBD patients and their siblings without IBD. BeadArray data were validated using a bisulfite polymerase chain reaction (PCR)-based restriction fragment length polymorphism (RFLP) method. To verify that observed changes in DNA methylation were not due to virus transformation, we compared specific CpG DNA methylation levels of GADD45A and POMC between B cell lines and matching peripheral blood B lymphocytes from five individuals.
Using this approach with strict statistical analysis, we identified 11 IBD-associated CpG sites, 14 CD-specific CpG sites, and 24 UC-specific CpG sites with methylation changes in B cells.
IBD- and subtype-specific changes in DNA methylation were identified in B cells from IBD patients. Many of these genes have important immune and inflammatory response functions including several loci within the interleukin (IL)-12/IL-23 pathway.
炎症性肠病（IBD）相关基因的甲基化状态的变化可能会显著改变基因表达水平，从而导致疾病的发生和进展。我们之前使用 Illumina GoldenGate BeadArray 分析方法从 IBD 患者的肠道组织中鉴定出了七个与疾病相关的 DNA 甲基化位点。
在这项研究中，我们扩展了这种方法，以鉴定来自 18 名 IBD 患者（9 名克罗恩病[CD]和 9 名溃疡性结肠炎[UC]）B 细胞中与 IBD 相关的 DNA 甲基化变化。由于方便获取外周血，B 细胞 DNA 甲基化标志物特别有利于诊断。
我们使用 Illumina GoldenGate BeadArray 分析方法检查了 B 细胞系的 DNA 甲基化谱。通过比较 IBD 患者和无 IBD 的兄弟姐妹的 B 细胞系之间的甲基化谱，鉴定出与疾病相关的具有 DNA 甲基化变化的 CpG/基因。使用基于亚硫酸氢盐的聚合酶链反应（PCR）-限制性片段长度多态性（RFLP）方法验证了 BeadArray 数据。为了验证观察到的 DNA 甲基化变化不是由于病毒转化引起的，我们比较了来自五个人的 B 细胞系和匹配的外周血 B 淋巴细胞中 GADD45A 和 POMC 的特定 CpG DNA 甲基化水平。
使用这种方法并进行严格的统计分析，我们在 IBD 患者的 B 细胞中鉴定出了 11 个与 IBD 相关的 CpG 位点、14 个 CD 特异性 CpG 位点和 24 个 UC 特异性 CpG 位点，这些位点的甲基化发生了变化。
在 IBD 患者的 B 细胞中鉴定出了与 IBD 相关的和亚型特异性的 DNA 甲基化变化。这些基因中有许多具有重要的免疫和炎症反应功能，包括白细胞介素（IL）-12/IL-23 通路内的多个位点。
