title: [mRNA expression and methylation status of p15 promoter in mouse bone marrow cells exposed to 1,4-benzoquinone].
abstract: OBJECTIVE
  To detect the expression and the CpG island methylation status of tumor suppressor gene p15 after exposure to 1,4-benzoquinone (1,4-BQ) in primary cultivated C57BL/6J mouse bone marrow cells in vitro.
  
  
  METHODS
  The mouse bone marrow cells were isolated in vitro. The effect of 0, 0.1, 1, 5, 10, 20, and 40 µmol/L 1,4-BQ on cell viability (CKK-8) was detected. 0, 0.1, 1, 10 µmol/L 1,4-BQ were used to intoxicate the mouse bone marrow cells for 24 h; Real-time PCR was employed to analyze the mRNA expression level of p15; The bisulfite sequencing PCR (BSP) was used to look into the methylation status of CpG islands in p15 promoter region.
  
  
  RESULTS
  1,4-BQ exhibited dose-dependent toxicity to mouse bone marrow cells, and the LC(50) was 8.3 µmol/L (95%CI: 4.6 - 10.6 µmol/L). The mRNA expression of p15 in 10 µmol/L group was only equivalent to 43% of control group. Compared with control group, the decrease of p15 mRNA expression in1 and 10 µmol/L concentration were obvious, and the differences had statistical significance (P < 0.05 or P < 0.01). BSP sequencing results were same between the exposure groups and control group, the 56 CpG sites on CpG islands remained in the state of unmethylated.
  
  
  CONCLUSIONS
  mRNA expression of p15 gene decreases after exposure to 1,4-BQ, but the CpG islands methylation status in promoter is not affected, suggesting that methylation does not participate in 1,4-BQ-mediated p15 gene expression decrease, other effect mechanisms still need to be investigated.
authors.affiliation: School of Environmental Science and Public Health, Wenzhou Medical College, Wenzhou 325035, China.
authors.forename: Jin-feng
authors.initials: JF
authors.lastname: Tian
authors.forename: Pei
authors.initials: P
authors.lastname: Cao
authors.forename: Xiu-yuan
authors.initials: XY
authors.lastname: Yu
authors.forename: Chun-hua
authors.initials: CH
authors.lastname: Peng
authors.forename: Xin-jun
authors.initials: XJ
authors.lastname: Yang
authors.forename: Hong-tao
authors.initials: HT
authors.lastname: Yan
mesh_terms: D000818:Animals; D001483:Base Sequence; D016227:Benzoquinones / Q000633:toxicity*; D001854:Bone Marrow Cells / Q000378:metabolism*; D002478:Cells, Cultured; D018899:CpG Islands; D050762:Cyclin-Dependent Kinase Inhibitor p15 / Q000235:genetics*; D019175:DNA Methylation; D004781:Environmental Exposure; D051379:Mice; D008810:Mice, Inbred C57BL; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000235:genetics
chemical_list: D016227:Benzoquinones; D050762:Cyclin-Dependent Kinase Inhibitor p15; D012333:RNA, Messenger; C004532:quinone