title: Detection of p16 promoter methylation in premature rats with chronic lung disease induced by hyperoxia.
abstract: BACKGROUND
  The aim of the present study was to investigate p16 promoter methylation in premature rats with chronic lung disease (CLD) induced by hyperoxia.
  
  
  METHODS
  Eighty Wistar rats were randomized into the hyperoxia group (fraction of inspired oxygen [FiO(2)] = 900 mL/L) or the control group (FiO(2) = 210 mL/L), 40 for each group. Semi-nested methylation-specific polymerase chain reaction (sn-MSP) was applied to detect p16 promoter hypermethylation in lung tissues. Additionally, p16 mRNA and protein expression was detected on reverse transcription-polymerase chain reaction (RT-PCR), western blot and the strept actividin-biotin complex method.
  
  
  RESULTS
  Extended exposure to hyperoxia led to increased methylation, and the methylation level reached a peak in the period of maximum pulmonary fibrosis in the hyperoxia group, while the methylation did not occur in the control group. The methylation rates on semi-nested PCR (sn-PCR) and nested-MSP were, respectively, 52.5% and 42.5% in the hyperoxia group. There was no statistically significant difference between the two methods. The p16 mRNA and protein expression was significantly higher in those with p16 promoter hypermethylation than those without.
  
  
  CONCLUSIONS
  Exposure to hyperoxia may induce p16 promoter hypermethylation in lung tissues in premature rats, and methylation risk increases as exposure extends. p16 promoter methylation induced by hyperoxia may be one of the mechanisms for low p16 mRNA and protein expression.
authors.affiliation: Department of Pediatrics, Shengjing Hospital of China Medical University, Shenyang, China.
authors.forename: Xiaohong
authors.initials: X
authors.lastname: Yue
authors.forename: Jianhua
authors.initials: J
authors.lastname: Fu
authors.forename: Xindong
authors.initials: X
authors.lastname: Xue
authors.forename: Hong
authors.initials: H
authors.lastname: Gao
authors.forename: Dongyan
authors.initials: D
authors.lastname: Liu
authors.forename: Zhihong
authors.initials: Z
authors.lastname: Zong
authors.forename: Wei
authors.initials: W
authors.lastname: Wang
authors.forename: Hui
authors.initials: H
authors.lastname: Li
authors.forename: Zhengwei
authors.initials: Z
authors.lastname: Yuan
mesh_terms: D000818:Animals; D000831:Animals, Newborn; D015153:Blotting, Western; D019941:Cyclin-Dependent Kinase Inhibitor p16 / Q000235:genetics* / Q000378:metabolism; D019175:DNA Methylation; D018496:Hyperoxia / Q000150:complications*; D008168:Lung / Q000378:metabolism*; D016133:Polymerase Chain Reaction; D011401:Promoter Regions, Genetic / Q000235:genetics*; D011658:Pulmonary Fibrosis / Q000209:etiology / Q000235:genetics* / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D020133:Reverse Transcriptase Polymerase Chain Reaction
chemical_list: D019941:Cyclin-Dependent Kinase Inhibitor p16; D012333:RNA, Messenger