Department of Medicine, Biomedical Genetics Section, Boston University School of Medicine, Boston, MA, United States; Department of Genetics & Genomics, Boston University School of Medicine, Boston, MA, United States; Genome Science Institute, Boston University School of Medicine, Boston, MA, United States.
Mental Health Research Center, Department of Psychiatry, Iran University of Medical Sciences, Tehran, Iran.
Schizophr Res. 2014 Feb;152(2-3):373-80. doi: 10.1016/j.schres.2013.12.007. Epub 2014 Jan 8.
Dysfunctional serotonin signaling has been linked to the pathogenesis of autism, obsessive compulsive disorder, mood disorders and schizophrenia. While the hypo-activity of serotonin signaling is involved in the pathogenesis of depression, anxiety and obsessive compulsive disorder; LSD, an agonist of serotonin type 2 receptor (5-HTR2A) induces psychosis. Therefore, anxiety and depressive disorders are treated by SSRIs which inhibit serotonin transporter (5-HTT) while psychotic disorders are controlled by drugs that block serotonin and/or dopamine receptors. Since genetic polymorphisms and epigenetic dysregulation of 5-HTT are involved in the pathogenesis of mental diseases, we analyzed DNA methylation of 5-HTT promoter in post-mortem brains and saliva samples of patients with schizophrenia (SCZ) and bipolar disorder (BD) to evaluate its potential application as a diagnostic and/or therapeutic biomarker in SCZ and BD.
Whole genome DNA methylation profiling was performed for a total of 24 samples (including two saliva samples) using the Illumina 27K (for 12 samples) and 450K DNA methylation array platform (for another 12 samples), followed by bisulfite sequencing to identify candidate CpGs for further analysis. Quantitative methylation specific PCR (qMSP) was used to assess the degree of CpG methylation of 5-HTT promoter in 105 post-mortem brains (35 controls, 35 SCZ and 35 BD) and 100 saliva samples (30 controls, 30 SCZ, 20 BD and 20 first degree relatives of SCZ or BD). The U133 2.0 Plus Human Transcriptome array for a total of 30 post-mortem brain samples (each group 10) followed by quantitative real-time PCR was used to study 5-HTT expression in 105 post-mortem brain samples.
The qMSP analysis for 5-HTT promoter region showed DNA hypermethylation in post-mortem brain samples of SCZ patients (30%), particularly in drug free patients (60%, p=0.04). Similarly, there was a trend for DNA hypermethylation in antipsychotic free BD patients (50%, p=0.066). qMSP analysis of DNA extracted from the saliva samples also exhibited hypermethylation of 5-HTT promoter in patients with SCZ (30%, p=0.039), which was more significant in drug naïve SCZ patients (>50%, p=0.0025). However, the difference was not significant between the controls and unaffected first degree relatives of patients with SCZ (p=0.37) and versus patients using antipsychotic drugs (p=0.2). The whole genome transcriptome analysis of post-mortem brain samples showed reduced expression of 5-HTT in SCZ compared to the control subjects (50%, p=0.008), confirmed by quantitative real-time PCR analysis (40%, p=0.035) which was more significant in drug free SCZ patients (~70%, p=0.022).
A correlation between reduction in 5-HTT expression and DNA hypermethylation of the 5-HTT promoter in drug naïve SCZ patients suggests that an epigenetically defined hypo-activity of 5-HTT may be linked to SCZ pathogenesis. Furthermore, this epigenetic mark in DNA extracted from saliva can be considered as one of the key determinants in a panel of diagnostic and/or therapeutic biomarkers for SCZ.
功能失调的血清素信号与自闭症、强迫症、情绪障碍和精神分裂症的发病机制有关。虽然血清素信号的低活性与抑郁症、焦虑症和强迫症的发病机制有关；但 LSD，一种血清素 2 型受体（5-HTR2A）激动剂，会引起精神病。因此，SSRIs 通过抑制血清素转运蛋白（5-HTT）来治疗焦虑症和抑郁症；而抗精神病药物则通过阻断血清素和/或多巴胺受体来控制精神病。由于 5-HTT 的遗传多态性和表观遗传失调与精神疾病的发病机制有关，我们分析了精神分裂症（SCZ）和双相情感障碍（BD）患者死后大脑和唾液样本中的 5-HTT 启动子的 DNA 甲基化，以评估其作为 SCZ 和 BD 的潜在诊断和/或治疗生物标志物的应用。
使用 Illumina 27K（用于 12 个样本）和 450K DNA 甲基化阵列平台（用于另外 12 个样本）对总共 24 个样本（包括两个唾液样本）进行全基因组 DNA 甲基化谱分析，随后进行亚硫酸氢盐测序以鉴定候选 CpG 进行进一步分析。定量甲基特异性 PCR（qMSP）用于评估 5-HTT 启动子在 105 个死后大脑（35 个对照、35 个 SCZ 和 35 个 BD）和 100 个唾液样本（30 个对照、30 个 SCZ、20 个 BD 和 20 个 SCZ 或 BD 的一级亲属）中的 CpG 甲基化程度。使用总共 30 个死后大脑样本的 U133 2.0 Plus 人类转录组阵列（每组 10 个），随后进行定量实时 PCR，研究了 105 个死后大脑样本中的 5-HTT 表达。
qMSP 分析 5-HTT 启动子区域显示 SCZ 患者死后大脑样本中的 DNA 过度甲基化（约 30%），特别是在未使用抗精神病药物的患者中（约 60%，p=0.04）。同样，抗精神病药物未治疗的 BD 患者的 DNA 过度甲基化也有趋势（约 50%，p=0.066）。qMSP 分析从唾液样本中提取的 DNA 也显示 SCZ 患者的 5-HTT 启动子过度甲基化（约 30%，p=0.039），在未使用药物的 SCZ 患者中更为显著（>50%，p=0.0025）。然而，在 SCZ 患者的对照组和未受影响的一级亲属之间（p=0.37）以及与使用抗精神病药物的患者之间（p=0.2），差异不显著。死后大脑样本的全基因组转录组分析显示，与对照组相比，SCZ 患者的 5-HTT 表达减少（约 50%，p=0.008），定量实时 PCR 分析进一步证实（约 40%，p=0.035），在未使用药物的 SCZ 患者中更为显著（约 70%，p=0.022）。
在未使用药物的 SCZ 患者中，5-HTT 表达减少与 5-HTT 启动子的 DNA 过度甲基化之间存在相关性，这表明 5-HTT 的表观遗传定义低活性可能与 SCZ 的发病机制有关。此外，从唾液中提取的 DNA 中的这种表观遗传标记可以被认为是 SCZ 诊断和/或治疗生物标志物的关键决定因素之一。
