| Project/Area Number |
09672056
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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 |
Surgical dentistry
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| Research Institution | Nagasaki University |
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Principal Investigator |
MATSUO Takemitsu The First Department of Oral and Maxillofacial Surgery, Nagasaki University, Assistant Professor, 歯学部・附属病院, 講師 (90190412)
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| Co-Investigator(Kenkyū-buntansha) |
NAITO Mariko Department of Oral bacteriology, Nagasaki University, Instructor, 歯学部, 助手 (20244072)
OHARA Naoya Department of Oral bacteriology, Nagasaki University, Assistant Professor, 歯学部, 助教授 (70223930)
松本 壮吉 長崎大学, 歯学部, 助手 (30244073)
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| Project Period (FY) |
1997 – 2000
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| Project Status |
Completed (Fiscal Year 2000)
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| Budget Amount *help |
¥3,300,000 (Direct Cost: ¥3,300,000)
Fiscal Year 2000: ¥500,000 (Direct Cost: ¥500,000)
Fiscal Year 1999: ¥700,000 (Direct Cost: ¥700,000)
Fiscal Year 1998: ¥800,000 (Direct Cost: ¥800,000)
Fiscal Year 1997: ¥1,300,000 (Direct Cost: ¥1,300,000)
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| Keywords | BCG / antigenic proteins / macrophage / phagosome / cytoplasm / release / MHC class I / anti-tumor effect / 抗原物質 / MPB70 / 抗腫癌効果 / DNA結合蛋白質 / リコンビナントBCG |
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| Research Abstract |
Mycobacterium bovis BCG (BCG) secrete different antigenic proteins. The amount and anti-tumor effects depend on BCG substrains. BCG has not only a direct anti-tumor effects but also indirect effects mediated macrophages. In order to study for an anti-cancer immuno-chemotherapy by using recombinant BCG, first we studied expression mechanisms, regulatory factors, and DNA binding proteins. Secondly we examined direct anti-tumor effects. Finally we investigated indirect anti-tumor effects mediated macrophages.
BCG substrain Tokyo (BCG Tokyo) secrete a large amount of the MPB70 protein, whereas BCG substrain Pasteur secrete little. The MPB70 genes in two substrains showed exactly the same sequence, and regulated transcriptional stage. We confirmed DNA binding proteins by gel shift assay, then extracted a 20 kDa protein, and confirmed a single strand DNA binding protein.
We prepared culture filtrate (CF) and cell lysate (Ly) by BCG Tokyo, then measured anti-tumor effects. The cell growth was measured by the human squamous cell carcinoma cell line (SCC-25) and human gingival fibroblast cell line (HGF). CF and Ly dose-dependently inhibited the cell growth of SCC-25. The inhibition effect of CF was higher than that of Ly and the anti-tumor effect of CF was also greater.
For an indirect anti-tumor effects mediating macrophages, we examined the secreted proteins of BCG released to the cytoplasm from the phagosome by BCG infected macrophages. The MPB70 protein was shown only the phagosomal fraction. We suppose the MPB70 protein released to the cytoplasm from the phagosome. Same result was observed for BCG infected SCC-25. Usually, bacteria were phagocyted by macrophage and fragmented into peptides in phagosome. These peptides bound to MHC class II molecules. But it was thought that the proteins released to the cytoplasm are fragmented peptides and bound to MHC class I molecules. According to exam, possibility was suggested to make CTLs activate much more.
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