title: DNA methylation status of NKX2-5, GATA4 and HAND1 in patients with tetralogy of fallot.
abstract: BACKGROUND
  NKX2-5, GATA4 and HAND1 are essential for heart development, however, little is known regarding their epigenetic regulation in the pathogenesis of tetralogy of fallot (TOF).
  
  
  METHODS
  Methylation levels were measured in three regions of NKX2-5 (M1: -1596 bp ~ -1374 bp, M2: -159 bp ~ 217 bp and M3: 1058 bp ~ 1524 bp), one region of GATA4 (M: -392 bp ~ 107 bp) and three regions of HAND1 (M1: -887 bp ~ -414 bp, M2: -436 bp ~ 2 bp and M3: 37 bp ~ 398 bp) using the Sequenom MassARRAY platform. QRT-PCR was used to analyze NKX2-5 and HAND1 mRNA levels in the right ventricular myocardium of TOF patients.
  
  
  RESULTS
  TOF patients had a significantly higher NKX2-5_M3 median methylation level than controls (41.65% vs. 22.18%; p = 0.0074; interquartile range [IQR]: 30.46%-53.35%, N = 30 and 20.07%-24.31%, N = 5; respectively). The HAND1_M1 median methylation level was also significantly higher in TOF patients than controls (30.05% vs. 17.54%; p = 0.0054; IQR: 20.77%-40.89%, N = 30 and IQR: 14.69%-20.64%; N = 6; respectively). The methylation statuses of NKX2-5_M1, NKX2-5_M2, GATA4_M, HAND1_M2 or HAND1_M3 were not significantly different in TOF patients compared to controls. The methylation values for NKX2-5_M3 were negatively correlated with mRNA levels (r = - 0.463, p = 0.010, N = 30) and there was a significant association between HAND1_M1 methylation status and mRNA levels (r = - 0.524, p = 0.003, N = 30) in TOF patients.
  
  
  CONCLUSIONS
  Aberrant methylation statuses of the NKX2-5 gene body and HAND1 promoter regions are associated with the regulation of gene transcription in TOF patients and may play an important role in the pathogenesis of TOF.
authors.forename: Wei
authors.initials: W
authors.lastname: Sheng
authors.forename: Yanyan
authors.initials: Y
authors.lastname: Qian
authors.forename: Huijun
authors.initials: H
authors.lastname: Wang
authors.forename: Xiaojing
authors.initials: X
authors.lastname: Ma
authors.forename: Ping
authors.initials: P
authors.lastname: Zhang
authors.forename: Lianwei
authors.initials: L
authors.lastname: Diao
authors.forename: Quan
authors.initials: Q
authors.lastname: An
authors.forename: Long
authors.initials: L
authors.lastname: Chen
authors.affiliation: Children Hospital of Fudan University, Shanghai 201102, China. duanma@yahoo.cn.
authors.forename: Duan
authors.initials: D
authors.lastname: Ma
authors.forename: Guoying
authors.initials: G
authors.lastname: Huang
mesh_terms: D001483:Base Sequence; D051792:Basic Helix-Loop-Helix Proteins / Q000235:genetics*; D016022:Case-Control Studies; D002675:Child, Preschool; D019175:DNA Methylation; D044127:Epigenesis, Genetic; D005260:Female; D050991:GATA4 Transcription Factor / Q000235:genetics*; D000072576:Homeobox Protein Nkx-2.5; D018398:Homeodomain Proteins / Q000235:genetics*; D006801:Humans; D007223:Infant; D008297:Male; D008969:Molecular Sequence Data; D009206:Myocardium / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D015203:Reproducibility of Results; D013771:Tetralogy of Fallot / Q000235:genetics*; D014157:Transcription Factors / Q000235:genetics*
chemical_list: D051792:Basic Helix-Loop-Helix Proteins; D050991:GATA4 Transcription Factor; D000072576:Homeobox Protein Nkx-2.5; D018398:Homeodomain Proteins; D012333:RNA, Messenger; D014157:Transcription Factors; C484170:GATA4 protein, human; C095552:NKX2-5 protein, human; C097341:helix-loop-helix protein, eHAND