| Project/Area Number |
06680708
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| Research Category |
Grant-in-Aid for General Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Research Field |
Cell biology
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| Research Institution | The Tokyo Metropolitan Institute of Medical Science |
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Principal Investigator |
ISHII Ai The Tokyo Metropolitan Institute of Medical Science Dept.of Cell Biology, 細胞生物学研究部門, 研究員 (80124436)
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| Co-Investigator(Kenkyū-buntansha) |
TAYA Choji The Tokyo Metropolitan Institute of Medical Science Dept.of Laboratory animal sc, 実験動物研究部門, 研究員 (90175456)
YONEKAWA Hiromichi The Tokyo Metropolitan Institute of Medical Science Dept.of Laboratory animal sc, 実験動物研究部門, 研究員 (30142110)
MORIYAMA Kenji The Tokyo Metropolitan Institute of Medical Science Dept.of Cell Biology, 細胞生物学研究部門, 研究員 (00250217)
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| Project Period (FY) |
1994 – 1995
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| Project Status |
Completed (Fiscal Year 1995)
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| Budget Amount *help |
¥1,900,000 (Direct Cost: ¥1,900,000)
Fiscal Year 1995: ¥800,000 (Direct Cost: ¥800,000)
Fiscal Year 1994: ¥1,100,000 (Direct Cost: ¥1,100,000)
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| Keywords | HSP70 / Transgenic mouse / Thymus / Negative selection of T cell / Apoptosis / Human disease-model mouse / 胸線 |
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| Research Abstract |
To investigate the role of HSP70 in vivo, we generated transgenic mice which express human HSP70.
1.Construction of BALB/3T3 cells constitutively expressing human HSP70 : Effect of HSP70 on the sensitivity to the TNF or Anti-Fas antibody.
To get some knowledge about the effect of increased level of HSP70, we first transfected BALB/3T3 cells by human HSP70 cDNA expressed under the control of human beta-actin promoter. We got many transfectants which expressed human HSP70. However, we found that the levels of HSP70 expression in those transformants were not so high as determined by, Western blotting analysis. A transfectant named +13 that was expressed at relatively high level was examined for its tolerance to heat (46ﾟC) , TNF and Anti-Fas antibodies. +13 cells were found to be significantly high resistant against not only heat but also the cytotoxicity of TNF or Anti-Fas antibodies. This result suggests that the increased level of HSP70 protects cells from the cell death (apoptosis) indu
… More
ced by various means.
2.Generating transgenic mice expressing human HSP70.
On the other hand, we attempted to generate transgenic mice which express human HSP70 under the control of mouse H-2K promoter or lck promoter. BALB/c mice were used. We generated 2 transgenic founder mice by a DNA fragment constructed to express human HSP70 under the control of H-2K promoter, and generated 3 transgenic founder mice by another DNA construct which expresses HSP70 under lck promotre.
(1) Tg mice with human HSP70 under H-2K promoter
By RT-PCR analysis, expression of the transgene mRNA was detected in all tissues, but human HSP70 protein was not significantly detected by Western blot analysis.
(2) Tg mice with human HSP70 under lck promoter
Expression of human HSP70 protein was detected only in thymus, and the level of human HSP70 expression was the same as that of mouse HSC70. We carried out various characterization in vivo or in vitro of tgand non-tg mice, we did not find any significant difference between tg mice and non-tg mice. Less
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