| Project/Area Number |
10670252
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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 |
Bacteriology (including Mycology)
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| Research Institution | HAMAMATU UNIVERSITY SCHOOL OF MEDECINE |
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Principal Investigator |
KOIDE Yukio Hamamatsu University School of Medicine, Professor, 医学部, 教授 (30126809)
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| Co-Investigator(Kenkyū-buntansha) |
YOSHIDA Atsushi Hamamatsu University School of Medicine, Assistant Professor, 医学部, 助手 (10242778)
NAGATA Toshi Hamamatsu University School of Medicine, Associate Professor, 医学部, 助教授 (90275024)
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| Project Period (FY) |
1998 – 1999
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| Project Status |
Completed (Fiscal Year 1999)
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| Budget Amount *help |
¥3,100,000 (Direct Cost: ¥3,100,000)
Fiscal Year 1999: ¥1,100,000 (Direct Cost: ¥1,100,000)
Fiscal Year 1998: ¥2,000,000 (Direct Cost: ¥2,000,000)
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| Keywords | DNA vaccine / Cytotoxic T lymphocytes (CTL) / Th1 cells / Intracellular pathogens / Protective immunity / Gene gun / 細胞障害性T細胞 |
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| Research Abstract |
DNA vaccines could have potential advantages over other types of vaccines that the vaccines can induce strong cellular immune responses. Since cytotoxic T lymphocytes (CTL) play an important role in the host immune defense against Listeria monocytogenes, an intracellular bacterium, we investigated the induction of CTL by DNA vaccines encoding LLO91-99, p60 217-225, or p60 449-457, all of which are presented with H-2KィイD1dィエD1. DNA immunization was performed by Helios gene gun system. 0.5mg of gold particles coated with 2μg plasmid DNA were inoculated into the shaved abdominal skin of BALB/c mice. Mice were inoculated three times with plasmids weekly. Two weeks after the last immunization, splenocytes were cultured for 5 days with peptide-pulsed splenocytes. CTL activity was assessed by 51Cr release assay.
Results : 1) Using DNA vaccines encoding LLO91-99, composed of different codons optimized to mammalian cells, we revealed that the codon optimization level of the genes correlates well
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with transfectional efficiency in murine cells, which is concomitantly associated with the induction level of specific CTL responses in the mouse. 2) The optimized LLO91-99 DNA vaccine, appeared to be to induce specific CTL without helper T cells and CpG motif of plasmid. 3) Immunization of each of three DNA vaccines resulted in different level of CTL induction. The relative magnitude of CTL induction can be ranked as follows : LLO91-99 > p60 217-225 >> p60 449-457. Direct frequency analysis of CTL with ELISPOT correlated the level of CTL activities. Furthermore, mixed inoculation of these DNA vaccines failed to surpass dominant LLO91-99 DNA vaccines in protection against listerial challenge.
Conclusion : 1) Optimization of codon usage of DNA vaccine is required for effective T cell responses against L. monocytogenes. 2) DNA vaccine encoding CTL epitope is capable of inducing CTL without helper T cells. 3) DNA vaccine encoding dominant CTL epitope is sufficient for the induction of CTL-mediated protective immunity. Less
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