title: [Oct4 methylation in induced differentiation of bone mesenchymal stem cells].
abstract: OBJECTIVE
  To investigate the methylation Oct4 in orientation induced differentiation in bone marrow mesenchymal stem cells
  
  
  METHODS
  Mice BMSCs were isolated and purified from bone marrow by adherent culture,and then identified by morphology and immunocytochemistry.Mouse osteoblastic cells were cultured by bone fragments inoculation,and then identified by alkaline phosphatase(AKP)staining and alizarin red staining.BMSCs were induced to differentiate into osteoblasts in vitro. Indirect immunofluorescence staining and reverse transcription polymerase chain reaction(RT PCR)were used to detect the expressions of Oct4 in BMSCs before and after induction.The methylation status of Oct4 gene in mouse BMSCs was explored by a methylation specific PCR before and after induction
  
  
  RESULTS
  The isolated mice BMSCs massively proliferated in vitro and formed cell colones with uniform morphology.Positive expressions of CD29,cKit,and CD44 and negative expression of CD34 were found in the isolated cells.After 10 days[DK]'[DK] induction,both AKP and the alizarin red were positive in cells and osteoblastic cells isolated from mice skull bones.The indirect immunoinfluorescence staining and RT-PCR also showed that the Oct4 expression in the directed differentiation of mouse BMSCs was down-regulated.The CpG island of Otc4 gene promoter in mouse BMSCs became methylated during the induced differentiation.
  
  
  CONCLUSIONS
  Mice BMSCs and osteoblasts were successfully cultured in vitro in this studyOct4 may be involved in the maintenance of adult stem cell pluripotency.The down regulated expression of Oct4 gene in mouse BMSCs during the directed differentiation may contribute to the methylation of CpG island in Otc4 gene promoter.
authors.affiliation: Department of Histology and Embryology, Shihezi University, Shihezi, Xinjiang, China.
authors.forename: Ju
authors.initials: J
authors.lastname: Wang
authors.forename: Yu-hao
authors.initials: YH
authors.lastname: Fu
authors.forename: Wei-shan
authors.initials: WS
authors.lastname: Wang
authors.forename: Duan-ming
authors.initials: DM
authors.lastname: Wang
authors.forename: Zong-yao
authors.initials: ZY
authors.lastname: Zhou
mesh_terms: D000818:Animals; D001854:Bone Marrow Cells / Q000166:cytology*; D002454:Cell Differentiation; D002478:Cells, Cultured; D018899:CpG Islands; D019175:DNA Methylation; D059630:Mesenchymal Stem Cells / Q000166:cytology*; D051379:Mice; D050814:Octamer Transcription Factor-3 / Q000378:metabolism*; D010006:Osteoblasts / Q000166:cytology*; D011401:Promoter Regions, Genetic
chemical_list: D050814:Octamer Transcription Factor-3; C494174:Pou5f1 protein, mouse