title: DNA hypermethylation of serotonin transporter gene promoter in drug naïve patients with schizophrenia.
abstract: BACKGROUND
  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.
  
  
  METHODS
  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.
  
  
  RESULTS
  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).
  
  
  CONCLUSIONS
  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.
authors.affiliation: 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.
authors.forename: Hamid Mostafavi
authors.initials: HM
authors.lastname: Abdolmaleky
authors.affiliation: Mental Health Research Center, Department of Psychiatry, Iran University of Medical Sciences, Tehran, Iran.
authors.forename: Shabnam
authors.initials: S
authors.lastname: Nohesara
authors.affiliation: Mental Health Research Center, Department of Psychiatry, Iran University of Medical Sciences, Tehran, Iran.
authors.forename: Mohammad
authors.initials: M
authors.lastname: Ghadirivasfi
authors.affiliation: 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.
authors.forename: Arthur W
authors.initials: AW
authors.lastname: Lambert
authors.affiliation: Mental Health Research Center, Department of Psychiatry, Iran University of Medical Sciences, Tehran, Iran.
authors.forename: Hamidreza
authors.initials: H
authors.lastname: Ahmadkhaniha
authors.affiliation: 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.
authors.forename: Sait
authors.initials: S
authors.lastname: Ozturk
authors.affiliation: 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.
authors.forename: Chen Khuan
authors.initials: CK
authors.lastname: Wong
authors.affiliation: Metrowest CNS Research Center, Psychiatric Clinical Trials, Pharmacogenomics, Natick, MA, United States.
authors.forename: Rahim
authors.initials: R
authors.lastname: Shafa
authors.affiliation: Arian Salamat Counseling and Nursing Cervices Center, Tehran, Iran.
authors.forename: Ashraf
authors.initials: A
authors.lastname: Mostafavi
authors.affiliation: 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; Department of Pathology and Laboratory Medicine, Boston University School of Medicine, Boston, MA, United States; Genome Science Institute, Boston University School of Medicine, Boston, MA, United States.
authors.forename: Sam
authors.initials: S
authors.lastname: Thiagalingam
mesh_terms: D000328:Adult; D001714:Bipolar Disorder / Q000235:genetics / Q000473:pathology; D001921:Brain / Q000378:metabolism*; D016022:Case-Control Studies; D019175:DNA Methylation / Q000235:genetics*; D004252:DNA Mutational Analysis; D005260:Female; D006801:Humans; D008297:Male; D008875:Middle Aged; D011180:Postmortem Changes; D011401:Promoter Regions, Genetic / Q000235:genetics*; D012463:Saliva; D012559:Schizophrenia / Q000235:genetics* / Q000473:pathology; D050486:Serotonin Plasma Membrane Transport Proteins / Q000235:genetics*
chemical_list: D050486:Serotonin Plasma Membrane Transport Proteins
keywords: Brain; DNA methylation; Saliva; Schizophrenia; Serotonin transporter