1999 Fiscal Year Final Research Report Summary
Study on regulation of inflammatory response by new mooclonal antibody against leukocytes.
| Project/Area Number |
10672058
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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 |
Biological pharmacy
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| Research Institution | Tokyo University of Pharmacy and Life Science,School of Pharmacy |
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Principal Investigator |
YADOMAE Toshiro Tokyo University of Pharmacy and Life Science, School of Pharmacy, Professor, 薬学部, 教授 (40057310)
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| Co-Investigator(Kenkyū-buntansha) |
ADACHI Yoshiyuki Tokyo University of Pharmacy and Life Science, School of Pharmacy, Associate Professor, 薬学部, 講師 (60222634)
OHNO Naohito Tokyo University of Pharmacy and Life Science, School of Pharmacy, Associate Professor, 薬学部, 助教授 (80152213)
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| Project Period (FY) |
1998 – 1999
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| Keywords | macrophage / CD14 / monoclonal antibody / cytokine / nitric oxide / STAT / MAPK / raft |
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| Research Abstract |
1998 : We have established an anti-CD14 mAb named 4Cl against murine macrophages. 4Cl can bind to thioglycolate-elicited peritoneal macrophages, bone marrow-derived macrophages and casein-induced peritoneal neutrophils. Immunostaining with 4Cl was inhibited by treatment of the cells with phosphatidylinositol specific phospholipase C, suggesting that the antigen is GPI-anchored. Immunoprecipitates from biotin-labeled RAW264.7 cell lysate with 4Cl were around 55 kDa and were visualized with rmC5-3, the only commercially available anti-murine CD14 mAb. 4Cl positively stained COS7 cells transfected with an expression vector containing cDNA of murine CD14. Pretreatment of macrophages with 4Cl reduced LPS-mediated production of TNFα, IL-6, and nitrite. The binding of FITC-LPS to RAW264.7 cell was blocked by pretreatment with 4Cl but not with rmC5-3. Pretreatment of cells with unlabeled 4Cl mAb but not unlabeled rmC5-3 reduced binding of FITC-4Cl. These results suggest that the 4Cl epitope on murine CD14 plays an important role in LPS binding and is distinct from the rmC5-3 epitope.
1999 : In order to explore a function of CD14, we had prepared new monoclonal antibody against murine CD14, named 4Cl. Addition of 4Cl to macrophage culture, NO production induced by LPS or by combination of IFNγ and IL-6 was significantly reduced. The inhibitory effect of 4Cl on cytokines-mediated NO production was also observed by using peritoneal macrophages from C3H/HeJ mice. Inhibition of iNOS expression was also observed in 4Cl-treatment. Pretreatment with 4Cl did not inhibit tyrosine phosphorylation of STAT1 and STAT3, but p38 MAP kinase. Preincubation of macrophages with methyl β-cyclodextrin to disturb raft formation, inhibitory effect of 4Cl on cytokine-mediated NO production was abrogated. These results suggest that cross-linking of CD14 by 4Cl may impair signal transduction via altering micromembrane environment around cytokine receptors.
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Research Products
(2 results)
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[Publications] Adachi, Y. Satokawa, C., Saeki, M., Ohno, N., Tamura, H., Tanaka, S., Yadomae, T.: "Inhibition by a CD14 monoclonal antibody of lipopolysaccharide binding to murine macrophages"J. Endotoxin Res.. 5. 139-146 (1999)
Description
「研究成果報告書概要(和文)」より
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[Publications] Adachi,Y. Satokawa,C., Saeki,M., Ohno,N., Tamura,H., Tanaka,S., Yadomae,T.: "Inhibition by a CD14 Monoclonal Antibody of Lipopolysaccharide Binding to Murine Macrophages"J.Endotoxin Res.. 5(3). 139-146 (1999)
Description
「研究成果報告書概要(欧文)」より
