title: Elevated expression of DNMT1 is associated with increased expansion and proliferation of hematopoietic stem cells co-cultured with human MSCs.
abstract: BACKGROUND
  Mesenchymal stem cells (MSCs) play an important role in hematopoietic stem cell (HSC) maintenance, proliferation, and apoptosis. DNA methyltransferase 1 (DNMT1) is considered an essential factor in the maintenance of HSCs in mammalian cells. Therefore, this study was conducted to evaluate the mRNA expression level of DNMT1 during cord blood (CB)-HSC ex vivo expansion with MSCs.
  
  
  METHODS
  Ex vivo cultures of CB-HSCs were performed in three culture conditions for 7 days: cytokines, cytokines with MSCs, and only MSCs. Total and viable cell numbers were counted after 5 and 7 days using trypan blue stain, and the stem cell percentage was then evaluated by flow cytometry. Moreover, in vitro colony-forming unit assay was carried out to detect clonogenic potential of HSCs at days 0 and 7 using MethoCult H4434. Finally, DNMT1 mRNA expression level was evaluated by real-time polymerase chain reaction.
  
  
  RESULTS
  Maximum CB-CD34+cell expansion was observed on day 7 in all the three cultures. After 7 days, ex vivo expansion of CB-CD34+cells indicated a significant decrease in DNMT1 expression in the cytokine cultures, whereas in the two co-culture conditions DNMT1 expression was increased. A significant difference between the number of CD34+and CD34-cells in the cytokine co-culture system was observed.
  
  
  CONCLUSIONS
  These data indicated that an elevated expression of DNMT1 is associated with increased expansion and proliferation of HSCs co-cultured with human MSCs. Hence, DNMT1 may be a potential factor in the maintenance of expanded HSCs co-cultured with human MSCs.
authors.affiliation: Blood Transfusion Research Center, High Institute for Research and Education in Transfusion Medicine, Tehran, Iran.
authors.forename: Moharram
authors.initials: M
authors.lastname: Ahmadnejad
authors.affiliation: Blood Transfusion Research Center, High Institute for Research and Education in Transfusion Medicine, Tehran, Iran.
authors.forename: Naser
authors.initials: N
authors.lastname: Amirizadeh
authors.affiliation: Blood Transfusion Research Center, High Institute for Research and Education in Transfusion Medicine, Tehran, Iran.
authors.forename: Roya
authors.initials: R
authors.lastname: Mehrasa
authors.affiliation: Cellular and Molecular Biology Research Center, Health Research Institute, Babol University of Medical Sciences, Babol, Iran.; Student Research Committee, School of Medicine, Babol University of Medical Sciences, Babol, Iran.
authors.forename: Ahmad
authors.initials: A
authors.lastname: Karkhah
authors.affiliation: Blood Transfusion Research Center, High Institute for Research and Education in Transfusion Medicine, Tehran, Iran.
authors.forename: Mahin
authors.initials: M
authors.lastname: Nikougoftar
authors.affiliation: Blood Transfusion Research Center, High Institute for Research and Education in Transfusion Medicine, Tehran, Iran.
authors.forename: Arezoo
authors.initials: A
authors.lastname: Oodi
keywords: Co-culture; Cytokines; DNA methyltransferase 1; Ex vivo expansion; HSCs; MSCs