| Project/Area Number |
15590994
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Hematology
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| Research Institution | The University of Tokyo |
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Principal Investigator |
TOJO Arinobu The University of Tokyo, The Institute of Medical Science, Associate Professor, 医科学研究所, 助教授 (00211681)
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| Co-Investigator(Kenkyū-buntansha) |
SODA Yasushi The University of Tokyo, The Institute of Medical Science, Research Associate, 医科学研究所, 助手 (00361618)
ASANO Shigetaka The University of Tokyo, The Institute of Medical Science, Professor, 医科学研究所, 教授 (50134614)
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| Project Period (FY) |
2003 – 2004
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| Project Status |
Completed (Fiscal Year 2004)
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| Budget Amount *help |
¥3,500,000 (Direct Cost: ¥3,500,000)
Fiscal Year 2004: ¥1,300,000 (Direct Cost: ¥1,300,000)
Fiscal Year 2003: ¥2,200,000 (Direct Cost: ¥2,200,000)
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| Keywords | leukemic stem cell / telomerase / hTERT / lentiviral vector / promoter / flow cytometry / AML / hTERTプロモーター / Venus |
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| Research Abstract |
We analyzed the promoter activity of the human telomerase reverse transcriptase (hTERT) gene in a single cell basis to gain insight into the hierarchy in replicative potential of acute myeloid leukemia (AML) cells. A self-inactivating lentiviral vector, in which a variant of yellow fluorescence protein (Venus) was expressed from the 1.2-kb upstream region of the hTERT gene, was constructed and introduced into 293T cells together with packaging plasmids. The resulting hTERT promoter reporter virus was used to infect primary blast cells from 19 AML patients. The expression of Venus was monitored by flow cytometry as well as under fluorescence microscopy. hTERT promoter activity displayed by the fluorescence intensity in the flow cytogram correlated well with endogenous telomerase activity. In 19 cases, hTERT promoter activity revealed considerable patient-to-patient variation, regardless of stimulation by colony-stimulating factors, but its distribution was rather broad and appeared as a single peak or two narrowly separated peaks, suggesting that hTERT expression occurs along a continuous gradient in the whole population. AML cells in S/G_2/M phase had greater promoter activity than those in G_0/G_1 phase. These results suggest heterogeneous hTERT promoter activity in individual AML cells and are compatible with hierarchy of replicative potential in AML.
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