Background: Alcohol use disorder (AUD) is a complex condition associated with adverse health consequences that affect millions of individuals worldwide. Epigenetic modifications, including DNA methylation (5 mC), have been associated with AUD and other alcohol-related traits. Epigenome-wide association studies (EWAS) have identified differentially methylated genes associated with AUD in human peripheral and brain tissue. More recently, epigenetic studies of AUD have also evaluated DNA hydroxymethylation (5 hmC) in the human brain. However, most of the epigenetic work in postmortem brain tissue has examined bulk tissue. In this study, we investigated neuronal-specific 5 mC and 5 hmC alterations at CpG sites associated with AUD in the human orbitofrontal cortex (OFC).Methods: Neuronal nuclei from the OFC were evaluated in 34 human postmortem brain samples (10 AUD, 24 non-AUD). Reduced representation oxidative bisulfite sequencing was used to assess 5 mC and 5 hmC at the genome-wide level. Differential 5 mC and 5 hmC were evaluated using the methylKit R package and significance was set at false discovery rate < 0.05 and differential methylation > 2. Functional enrichment analyses were performed, and gene-level convergence was evaluated in an independent dataset that assessed 5 mC and 5 hmC of AUD in bulk cortical tissue.Results: We identified 417 5 mC and 363 5hmC significant differential CpG sites associated with AUD, with 59% in gene promoters. Some of the identified genes have been previously implicated in alcohol consumption, including SYK, DNMT3A for 5 mC, GAD1, DLX1, DLX2, for 5 hmC and GATA4 in both. Convergence with a previous AUD 5 mC and 5 hmC study was observed for 28 genes. We also identified 5 and 35 differential regions for 5 mC and 5 hmC, respectively. Lastly, GWAS enrichment analysis showed an association with AUD for differential 5 mC genes.Discussion: This study reveals neuronal-specific methylome and hydroxymethylome dysregulation associated with AUD, identifying both previously reported and potentially novel gene associations with AUD. Our findings provide new insights into the epigenomic dysregulation of AUD in the human brain.

中文翻译：

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神经元特异性甲基化组和羟甲基化组分析揭示了与酒精使用障碍相关的显着位点

背景：酒精使用障碍（AUD）是一种复杂的疾病，与影响全球数百万人的不良健康后果相关。表观遗传修饰，包括 DNA 甲基化 (5 mC)，与 AUD 和其他酒精相关性状有关。全表观基因组关联研究 (EWAS) 已在人类外周和脑组织中鉴定出与 AUD 相关的差异甲基化基因。最近，AUD 的表观遗传学研究还评估了人脑中的 DNA 羟甲基化 (5 hmC)。然而，死后脑组织的大多数表观遗传学工作都是检查大块组织。在本研究中，我们研究了人类眶额皮质 (OFC) 中与 AUD 相关的 CpG 位点的神经元特异性 5 mC 和 5 hmC 变化。方法：在 34 个人类死后脑样本（10 AUD、24非澳元）。使用简化代表性氧化亚硫酸氢盐测序在全基因组水平评估 5 mC 和 5 hmC。使用methylKit R包评估差异5 mC和5 hmC，并且将显着性设定为错误发现率＜1。 0.05且差异甲基化> 2. 进行了功能富集分析，并在独立数据集中评估了基因水平收敛，该数据集评估了大块皮质组织中 AUD 的 5 mC 和 5 hmC。 结果：我们确定了 417 个 5 mC 和 363 个 5hmC 与 AUD 相关的显着差异 CpG 位点，其中 59% 为基因启动子。一些已识别的基因先前已与饮酒有关，包括 SYK、5 mC 的 DNMT3A、5 hmC 的 GAD1、DLX1、DLX2 以及两者中的 GATA4。观察到 28 个基因与之前 AUD 5 mC 和 5 hmC 研究趋同。我们还分别鉴定了 5 mC 和 5 hmC 的 5 个和 35 个差异区域。最后，GWAS 富集分析显示差异 5 mC 基因与 AUD 相关。讨论：这项研究揭示了与 AUD 相关的神经元特异性甲基化组和羟甲基化组失调，识别了先前报道的和潜在的新基因与 AUD 的关联。我们的研究结果为人脑中 AUD 的表观基因组失调提供了新的见解。