2004 Fiscal Year Final Research Report Summary
Analysis of pathogen recognition by TLR4 and RP105 and their application to immunothrapy
| Project/Area Number |
13470073
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| Research Category |
Grant-in-Aid for Scientific Research (B)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Immunology
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| Research Institution | Saga University (2004)
佐賀医科大学 (2001-2003) |
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Principal Investigator |
KIMOTO Masao Saga University, Medicine, Professor, 医学部, 教授 (40153225)
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| Co-Investigator(Kenkyū-buntansha) |
NAGASAWA Kouhei Saga University, Medicine, Professor, 医学部, 教授 (00108721)
MIYAKE Kensuke The University of Tokyo, The Institute of Medical Science, Professor, 医科学研究所, 教授 (60229812)
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| Project Period (FY) |
2001 – 2004
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| Keywords | MD-2 / recombinant protein / bacterial expression / CD14 / LBP / ligand / LPS / cytokine |
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| Research Abstract |
1.Analysis of the TLR and RP105 molecular structure : We obtained highly purified form of recombinant bacterial products of MD-2,CD14 and LBP. Using these purified proteins, we analyzed thier critical structures for binding to LPS. We also revealed the differential binding of CD14 and MD-2 to LPS.
2.Analysis of TLR4 and RP105 ligands : We found that MD-2 directly binds to LPS. On the other hand, MD-1 was found not to bind LPS directly, although LPS can signal through RP105/MD-1 molceule. We compared the LPS binding pattern between MD-1 and MD-2 using their site-specific mutant molecules.
3.Role of TLR4 and RP105 in immunological diseases : We found increased frequency of RP105-negative B cells in SLE patients. Culture of RP105-negative B cells resulted in the secretion of IgG and IgM, the production of these were augmented by SAC or IL-6 stimulation. RP105-negative B cells from SLE patients produced dsDNA antibodies. These results suggest that RP105-negative B cells are in an activated state and possibly involved in the production of auto-antibodies.
4.Manipulation of TLR4 and RP105 signal transduction : We found alternative spliced forms of MD-2 by RT-PCR. Such alternative form of MD-2 molecules may potentially function as dominant negative molecules for TLRA/MD-2 signaling, and may function as a potential tool for the manipulation of LPS signaling. We also obtained monoclonal antibodies against TLRA/MD-2. This mAb was found to transmit activation signals through TLRA/MD-2 and resulted in the secretion of pro-inflammatory cytokines.
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