Department of Pediatrics, Ditmanson Medical Foundation Chia-Yi Christian Hospital, Chia-Yi, Taiwan; Min-Hwei Junior College of Health Care Management, Tainan, Taiwan.
Department of Biomedical Sciences and Institute of Molecular Biology, National Chung Cheng University, Chia-Yi, Taiwan.
J Microbiol Immunol Infect. 2020 Dec;53(6):986-995. doi: 10.1016/j.jmii.2020.06.004. Epub 2020 Jun 26.
BACKGROUND/PURPOSE: Allergen-specific immunotherapy (SIT) is now considered curative to allergic diseases such as asthma. Mechanistically, our previous work showed DNA hypermethylation of cytokine genes, in T-helper cells, in allergic asthmatic children treated with allergen-SIT. In this study, we extended to work to assess possible changes in the DNA methylomes of peripheral blood mononuclear cells (PBMCs), isolated from mite allergen-SIT asthmatic children, to explore further the underlying methylation changes.
Thirteen allergic asthmatic children who received Der p-SIT, 12 non-SIT allergic asthmatic controls, and 12 healthy controls were enrolled. Bisulfite-converted DNA from Der p-stimulated PBMCs was analyzed using Human Methylation 450 k BeadChip. Pyrosequencing and quantitative real-time PCR were used to validate the DNA methylation levels and the gene expression of individual samples.
We identified 108 significantly differentially methylated regions (DMRs) unique to Der p-treated PBMCs, with 53 probes linked to demethylated DMRs, and 55 probes linked to methylated DMRs. Three associated genes (BCL6, HSPG2, and HSP90AA1), of selected DMRs, were subjected to bisulfite pyrosequencing. Of these, BCL6 showed significant hypomethylation, while HSPG2 and HSP90AA1 were hypermethylated in SIT group, compared to the AA group. Furthermore, SIT group had significantly higher gene expression of BCL6 and lower gene expression of HSPG2. KEGG pathway analysis further revealed DMR genes involved in ECM-receptor interactions, asthma, and antigen processing and presentation pathways.
Several DNA regions showed DNA methylation altered by Der p specific immunotherapy, indicating desensitization-associated methylomes. Genes belonging to these SIT-altered pathways may represent therapeutic targets for better clinical management of asthma.
背景/目的：变应原特异性免疫治疗（SIT）目前被认为可治愈哮喘等过敏性疾病。从机制上讲，我们之前的研究表明，接受变应原-SIT 治疗的过敏性哮喘儿童的辅助性 T 细胞中的细胞因子基因发生了 DNA 超甲基化。在这项研究中，我们进一步评估了从尘螨变应原-SIT 哮喘儿童分离的外周血单个核细胞（PBMC）的 DNA 甲基组谱是否可能发生变化，以进一步探索潜在的甲基化变化。
纳入 13 例接受 Der p-SIT 的过敏性哮喘儿童、12 例非 SIT 过敏性哮喘对照者和 12 例健康对照者。使用 Human Methylation 450 k BeadChip 分析 Der p 刺激的 PBMC 中的经亚硫酸氢盐转化的 DNA。使用焦磷酸测序和实时定量 PCR 验证个体样本的 DNA 甲基化水平和基因表达。
我们鉴定了 108 个独特的、Der p 处理的 PBMC 中差异甲基化区域（DMR），其中 53 个探针与去甲基化 DMR 相关，55 个探针与甲基化 DMR 相关。在选定的 DMR 中，有 3 个相关基因（BCL6、HSPG2 和 HSP90AA1）进行了亚硫酸氢盐焦磷酸测序。其中，与 AA 组相比，SIT 组的 BCL6 表现出显著的低甲基化，而 HSPG2 和 HSP90AA1 则表现出高甲基化。此外，SIT 组的 BCL6 基因表达显著升高，HSPG2 基因表达显著降低。KEGG 通路分析进一步揭示了 DMR 基因参与 ECM-受体相互作用、哮喘和抗原加工与呈递途径。
一些 DNA 区域的 DNA 甲基化被 Der p 特异性免疫治疗改变，表明脱敏相关的甲基组。属于这些 SIT 改变途径的基因可能代表治疗哮喘的潜在治疗靶点，以更好地进行临床管理。
