Abstract | BACKGROUND: METHODS: Liquid chromatography mass spectrometry/mass spectrometry (LC-MS/MS) was used to assess 5-methyldeoxycytidine (5-mC), 5-hydroxymethyldeoxycytidine (5-hmC), 5-formyldeoxycytidine (5-fC) and 5-carboxyldeoxycytidine (5-caC) quantitatively in tumorous and non-tumorous regions of GCs; [D2]-5-hmC was used as an internal standard. Expression levels of the genes TET1, TET2, TET3, TDG, IDH1 and IDH2 were measured using a real-time reverse transcription polymerase chain reaction (RT-PCR) and were compared to the clinical attributes of each case. Using HEK293T cells the effects of introducing plasmids containing full-length TET1, TET2, and TET3 and 7 variants of the TET2 catalytic domain were evaluated in terms of their effect on cytosine demethylation. RESULTS: LC-MS/MS showed that 5-hmC was significantly decreased in tumorous portions. 5-mC was also moderately decreased in tumors, while 5-fC and 5-caC were barely detectable. The expressions of TET1, TET2, TET3, TDG and IDH2, but not IDH1, were notably decreased in GCs, compared with the adjacent non- tumor portion. TET1 expression and the 5-hmC levels determined using LC-MS/MS had a significantly positive correlation and TET1 protein had a greater effect on the increase in 5-hmC than TET2 and TET3 in HEK293T cells. CONCLUSIONS: The loss of 5-hmC and the down-regulation of TET1-3, TDG and IDH2 were found in GCs. The loss of 5-hmC in GCs was mainly correlated with the down-regulation of TET1.
|
Authors | Chunping Du, Nobuya Kurabe, Yoshitaka Matsushima, Masako Suzuki, Tomoaki Kahyo, Ippei Ohnishi, Fumihiko Tanioka, Shogo Tajima, Masanori Goto, Hidetaka Yamada, Hong Tao, Kazuya Shinmura, Hiroyuki Konno, Haruhiko Sugimura |
Journal | Gastric cancer : official journal of the International Gastric Cancer Association and the Japanese Gastric Cancer Association
(Gastric Cancer)
Vol. 18
Issue 3
Pg. 516-25
(Jul 2015)
ISSN: 1436-3305 [Electronic] Japan |
PMID | 25098926
(Publication Type: Journal Article, Research Support, Non-U.S. Gov't)
|
Chemical References |
- 5-formyl-2'-deoxycytidine
- 5-hydroxymethyl-2'-deoxycytidine
- DNA-Binding Proteins
- Enzymes
- Proto-Oncogene Proteins
- Deoxycytidine
- 5-hydroxymethylcytosine
- 5-Methylcytosine
- Cytosine
- 5-methyldeoxycytidine
- Mixed Function Oxygenases
- TET1 protein, human
- TET3 protein, human
- IDH2 protein, human
- Isocitrate Dehydrogenase
- IDH1 protein, human
- Dioxygenases
- TET2 protein, human
|
Topics |
- 5-Methylcytosine
(analogs & derivatives)
- Aged
- Chromatography, Liquid
- Cytosine
(analogs & derivatives, analysis, metabolism)
- DNA-Binding Proteins
(genetics)
- Deoxycytidine
(analogs & derivatives, analysis, metabolism)
- Dioxygenases
(genetics)
- Enzymes
(genetics, metabolism)
- Female
- Gene Expression Regulation, Neoplastic
- HEK293 Cells
- Humans
- Isocitrate Dehydrogenase
(genetics)
- Male
- Middle Aged
- Mixed Function Oxygenases
- Polymorphism, Single Nucleotide
- Proto-Oncogene Proteins
(genetics)
- Stomach Neoplasms
(enzymology, genetics, metabolism)
- Tandem Mass Spectrometry
|