Elevated expression of DNMT1 is associated with increased expansion and proliferation of hematopoietic stem cells co-cultured with human MSCs.
- Author:
Moharram AHMADNEJAD
1
;
Naser AMIRIZADEH
;
Roya MEHRASA
;
Ahmad KARKHAH
;
Mahin NIKOUGOFTAR
;
Arezoo OODI
Author Information
- Publication Type:Original Article
- Keywords: DNA methyltransferase 1; HSCs; MSCs; Co-culture; Ex vivo expansion; Cytokines
- MeSH: Apoptosis; Cell Count; Coculture Techniques; Cytokines; DNA; Fetal Blood; Flow Cytometry; Hematopoietic Stem Cells*; Humans*; In Vitro Techniques; Mesenchymal Stromal Cells; Real-Time Polymerase Chain Reaction; RNA, Messenger; Stem Cells; Trypan Blue
- From:Blood Research 2017;52(1):25-30
- CountryRepublic of Korea
- Language:English
- 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. CONCLUSION: 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.
