| Project/Area Number |
06807025
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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 |
寄生虫学(含医用動物学)
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| Research Institution | Saitama Medical School |
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Principal Investigator |
HIRAYAMA Kenji Saitama Medical School, Professor, 医学部, 教授 (60189868)
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| Co-Investigator(Kenkyū-buntansha) |
堀 栄太郎 埼玉医科大学, 医学部, 教授 (30049762)
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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 |
¥2,000,000 (Direct Cost: ¥2,000,000)
Fiscal Year 1995: ¥1,000,000 (Direct Cost: ¥1,000,000)
Fiscal Year 1994: ¥1,000,000 (Direct Cost: ¥1,000,000)
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| Keywords | Trypanosoma cruzi / HLA / Peptide vaccine / Cysteine protease / Cytotoxic T cell / CD8 molecule / Genetic variation / Protective immunity / 防御免疫 / トランスジェニックマウス / トリパノソーマ / 抗原ペプチド |
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| Research Abstract |
Analysis of the protective immunity against infection with Trypanosoma cruzi (TC) in the experimental animalmodel using a HLA-B35 transgenic mouse (HLA-B35-TG).
CD8 positive cytotoxic T cells are known to be major effector cells to kill the TC infected host cells. Those T cells recognize the invaders antigen with their own HLA-class lmolecules. Using chronically infected HLA-B35-TG,we tried to identify the T cell epitopes that HLA molecule presents to the CD8 cytotoxic T cells. Using the information of the binding motif of HLA-B35, we selected and synthesized 61 different peptides (8mer to 10 mer) from TC cysteine protease. After the binding assay, 5 peptides were chosen for in vitro stimulation experiments. Two months after the infection with TC,HLA-B35-TG mice were sacrificed to get the splenocytes. One million cells in 1 ml of culture medium were incubated with 10muM of each peptide and 30Units of IL-2. Ten dyas later, re-stimulation was done by adding irradiated peptide pulsed autologous splenocytes and IL-2. Totally 2wks. later, the viable cells (T cells) were examined for their cytotoxic activity against peptide pulsed target cells (HLA-B35 Transfectant) by Chromium 51 release assay. None of the peptides stimulated the cytotoxic T cells. though we repeated the experiments twice. Main reasons why there were no effective stimulation were considered as follows.
(1) HLA-B35 might not be ideal for the analysis of immune response against the cysteine protease antigen due to row affintiy.
(2) Human CD8 might have been essential to complete the HLA-peptide-T cell interaction.
2. Search for the major T cell antigens recognized by human T cells from the patients in the endemic area. We have produced T cell clones reactive to epimastigote antigen. Using a T cell clone, 55KDa protein were purified whose molecular weight is almost the same as the cysteine protease. The amino acid sequencing are now on going study.
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