title: Targeted-bisulfite sequence analysis of the methylation of CpG islands in genes encoding PNPLA3, SAMM50, and PARVB of patients with non-alcoholic fatty liver disease.
abstract: OBJECTIVE
  The pathogenesis of non-alcoholic fatty liver disease (NAFLD) is affected by epigenetic factors as well as by genetic variation.
  
  
  METHODS
  We performed targeted-bisulfite sequencing to determine the levels of DNA methylation of 4 CpG islands (CpG99, CpG71, CpG26, and CpG101) in the regulatory regions of PNPLA3, SAMM50, PARVB variant 1, and PARVB variant 2, respectively. We compared the levels of methylation of DNA in the livers of the first and second sets of patients with mild (fibrosis stages 0 and 1) or advanced (fibrosis stages 2 to 4) NAFLD and in those of patients with mild (F0 to F2) or advanced (F3 and F4) chronic hepatitis C infection. The hepatic mRNA levels of PNPLA3, SAMM50, and PARVB were measured using qPCR.
  
  
  RESULTS
  CpG26, which resides in the regulatory region of PARVB variant 1, was markedly hypomethylated in the livers of patients with advanced NAFLD. Conversely, CpG99 in the regulatory region of PNPLA3 was substantially hypermethylated in these patients. These differences in DNA methylation were replicated in a second set of patients with NAFLD or chronic hepatitis C. PNPLA3 mRNA levels in the liver of the same section of a biopsy specimen used for genomic DNA preparation were lower in patients with advanced NAFLD compared with those with mild NAFLD and correlated inversely with CpG99 methylation in liver DNA. Moreover, the levels of CpG99 methylation and PNPLA3 mRNA were affected by the rs738409 genotype.
  
  
  CONCLUSIONS
  Hypomethylation of CpG26 and hypermethylation of CpG99 may contribute to the severity of fibrosis in patients with NAFLD or chronic hepatitis C infection.
authors.affiliation: Pharmacogenomics, Kyoto University Graduate School of Medicine, Kyoto, Japan.
authors.forename: Takuya
authors.initials: T
authors.lastname: Kitamoto
authors.affiliation: Pharmacogenomics, Kyoto University Graduate School of Medicine, Kyoto, Japan.
authors.forename: Aya
authors.initials: A
authors.lastname: Kitamoto
authors.affiliation: Department of Gastroenterology and Hepatology, Yokohama City University Graduate School of Medicine, Yokohama, Japan.
authors.forename: Yuji
authors.initials: Y
authors.lastname: Ogawa
authors.affiliation: Department of Gastroenterology and Hepatology, Yokohama City University Graduate School of Medicine, Yokohama, Japan.
authors.forename: Yasushi
authors.initials: Y
authors.lastname: Honda
authors.affiliation: Department of Gastroenterology and Hepatology, Yokohama City University Graduate School of Medicine, Yokohama, Japan.
authors.forename: Kento
authors.initials: K
authors.lastname: Imajo
authors.affiliation: Department of Gastroenterology and Hepatology, Yokohama City University Graduate School of Medicine, Yokohama, Japan.
authors.forename: Satoru
authors.initials: S
authors.lastname: Saito
authors.affiliation: Department of Gastroenterology and Hepatology, Yokohama City University Graduate School of Medicine, Yokohama, Japan.
authors.forename: Masato
authors.initials: M
authors.lastname: Yoneda
authors.affiliation: Laboratory for Mathematics, National Defense Medical College, Tokorozawa, Japan.
authors.forename: Takahiro
authors.initials: T
authors.lastname: Nakamura
authors.affiliation: Department of Gastroenterology and Hepatology, Yokohama City University Graduate School of Medicine, Yokohama, Japan.
authors.forename: Atsushi
authors.initials: A
authors.lastname: Nakajima
authors.affiliation: Pharmacogenomics, Kyoto University Graduate School of Medicine, Kyoto, Japan. Electronic address: kikukoh@kuhp.kyoto-u.ac.jp.
authors.forename: Kikuko
authors.initials: K
authors.lastname: Hotta
mesh_terms: D000185:Actinin / Q000235:genetics* / Q000378:metabolism; D018899:CpG Islands; D004247:DNA / Q000235:genetics*; D019175:DNA Methylation; D005260:Female; D020022:Genetic Predisposition to Disease; D005838:Genotype; D006801:Humans; D008049:Lipase / Q000235:genetics* / Q000378:metabolism; D008099:Liver / Q000378:metabolism* / Q000473:pathology; D008297:Male; D008565:Membrane Proteins / Q000235:genetics* / Q000378:metabolism; D008875:Middle Aged; D065626:Non-alcoholic Fatty Liver Disease / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D010750:Phosphoproteins; D016133:Polymerase Chain Reaction; D011110:Polymorphism, Genetic; D000217:Acyltransferases; D054512:Phospholipases A2, Calcium-Independent
chemical_list: D000185:Actinin; D004247:DNA; D008049:Lipase; D008565:Membrane Proteins; D010750:Phosphoproteins; C486591:PARVB protein, human; C533682:PNPLA3 protein, human; D000217:Acyltransferases; D054512:Phospholipases A2, Calcium-Independent
keywords: DNA methylation; Fibrosis; Next-generation sequencing; Non-alcoholic fatty liver disease; PARVB; PNPLA3; SAMM50