Demethylating agent, 5-azacytidine, reverses differentiation of embryonic stem cells

The de novo methylation activity is essential for embryonic development as well as embryonic stem (ES) cell differentiation, where the intensive and extensive DNA methylation was detected. In this study, we investigated the effects of a demethylating agent, 5-azacytidine (5-AzaC), on differentiated...

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Bibliographic Details
Published in:Biochemical and biophysical research communications 2004-10, Vol.323 (1), p.86-90
Main Authors: Tsuji-Takayama, Kazue, Inoue, Toshiya, Ijiri, Yoshihiro, Otani, Takeshi, Motoda, Ryuichi, Nakamura, Shuji, Orita, Kunzo
Format: Article
Language:English
Subjects:
5-Azacytidine
Alkaline Phosphatase - metabolism
Animals
Azacitidine - pharmacology
Cell Differentiation - drug effects
Cell Line
Demethylation
Differentiation
DNA Methylation
Embryo, Mammalian - cytology
Embryo, Mammalian - drug effects
Enzyme Inhibitors - pharmacology
Flow Cytometry
H19
Methylation
Mice
Polymerase Chain Reaction
Reverse Transcriptase Polymerase Chain Reaction
RNA, Long Noncoding
RNA, Untranslated - metabolism
SSEA-1
Stem Cells - cytology
Stem Cells - drug effects
Sulfites - pharmacology
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Summary:The de novo methylation activity is essential for embryonic development as well as embryonic stem (ES) cell differentiation, where the intensive and extensive DNA methylation was detected. In this study, we investigated the effects of a demethylating agent, 5-azacytidine (5-AzaC), on differentiated ES cells in order to study the possibility of reversing the differentiation process. We first induced differentiation of ES cells by forming embryoid bodies, and then the cells were treated with 5-AzaC. The cells showed some undifferentiated features such as stem cell-like morphology with unclear cell-to-cell boundary and proliferative responsiveness to LIF. Moreover, 5-AzaC increased the expressions of ES specific markers, SSEA-1, and alkaline phosphatase activity as well as ES specific genes, Oct4, Nanog, and Sox2. We also found that 5-AzaC demethylated the promoter region of H19 gene, a typical methylated gene during embryonic differentiation. These results indicate that 5-AzaC reverses differentiation state of ES cells through its DNA demethylating activity to differentiation related genes.
ISSN:0006-291X
1090-2104
DOI:10.1016/j.bbrc.2004.08.052