| Project/Area Number |
12307017
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| Research Category |
Grant-in-Aid for Scientific Research (A)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Pediatrics
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| Research Institution | KYOTO UNIVERSITY |
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Principal Investigator |
NAKAHARA Tatsutoshi Graduate School of Medicine KYOTO UNIVERSITY Professor, 医学研究科, 教授 (20110744)
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| Co-Investigator(Kenkyū-buntansha) |
YORIFUJI Tohru Graduate School of Medicine KYOTO UNIVERSITY Associated Professor, 医学研究科, 助手 (60220779)
KATAMURA Kenji Graduate School of Medicine KYOTO UNIVERSITY Associated Professor, 医学研究科, 講師 (40185806)
HEIKE Toshio Graduate School of Medicine KYOTO UNIVERSITY Associated Professor, 医学研究科, 助教授 (90190173)
林 英蔚 京都大学, 医学研究科, 助手 (40283600)
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| Project Period (FY) |
2000 – 2001
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| Project Status |
Completed (Fiscal Year 2001)
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| Budget Amount *help |
¥41,370,000 (Direct Cost: ¥37,500,000、Indirect Cost: ¥3,870,000)
Fiscal Year 2001: ¥16,770,000 (Direct Cost: ¥12,900,000、Indirect Cost: ¥3,870,000)
Fiscal Year 2000: ¥24,600,000 (Direct Cost: ¥24,600,000)
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| Keywords | embryonic stem cells / hematopoietic stem cells / stromal cell line / mouse / regenarated medicine / culture / transplantation / molecular cloning / 中胚葉細胞 / Flk1 / AGM領域 |
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| Research Abstract |
Recent studies on the developing mouse embryo have shown that long-term repopulating hematopoietic stem cells (LTR-HSC) are first identified in the aorta-gonad-mesonephros (AGM) region at 10 to 10.5 days post coitum (dpc), prior to such activity being observed in yolk sac (YS) and fetal liver, suggesting that the AGM region at 10 to 11 dpc provides a microenvironment suitable for the development of LTR-HSC. These observations prompted us to establish stromal cell lines from the AGM region at 10 to 11 dpc, which would stimulate generation of hematopoietic stem cells from more early stage of embryo or YS. We succeeded the establishment of a stromal cell line (A9) from the AGM region of 10.5 dpc mouse embryo. When co-cultured with the A9 for 4 days, both the cells from YS and intraembryonic paraaortic splanchnopleures (P-Sp) at 8 to 8.5 dpc generated LTR-HSC capable of reconstituting definitive hematopoiesis in adult mice. No stromal cell lines other than A9 could support generation of LTR-HSC from YS and P-Sp as early as 8.5 dpc. These findings suggest that precursors with the potential to generate definitive LTR-HSC appear independently in YS and intraembryonic P-Sp and that A9 expresses some molecules that facilitate generation of LTR-HSC from precursors. We started the molecular cloning of the molecules which might deeply contribute stem cell-genesis. In preliminary experiment, we have examined whether A9 could support generation of LTR-HSC from murine embryonic stem (ES) cells. Flk-1^+ VE-cadherin^- PECAM-1^- cells sorted from differentiation culture of ES cells were co-cultured with A9 cells and then transplanted into lethally irradiated adult mice. Although donor (ES cells) type hematopoietic cells failed to be detected by FACS analysis, polymerase chain reaction of bone marrow cells 6 months after transplantation revealed donor type message, suggesting that possibility of the generation of LTR-HSC from ES cells using A9 or some molecules expressing A9 cells.
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