| Project/Area Number |
09460133
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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 |
Basic veterinary science/Basic zootechnical science
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| Research Institution | HOKKAIDO UNIVERSITY |
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Principal Investigator |
KUWABARA Mikinori Grad.School of Vet.Med., Hokkaido Univ., Pro., 大学院・獣医学研究科, 教授 (10002081)
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| Co-Investigator(Kenkyū-buntansha) |
TAJIMA Motoshi Grad.School of Vet.Med., Hokkaido Univ., Lec., 大学院・獣医学研究科, 講師 (90202168)
KIMURA Kazuhiro Grad.School of Vet.Med., Hokkaido Univ., Asso.Pro., 大学院・獣医学研究科, 助教授 (30192561)
NAGAHATA Hajime Fac.of Vet.Med., Rakuno Gakuen Univ., Pro., 獣医学部, 教授 (10133571)
SYUTO Bun-ei Fac.Agri., Iwate Univ., Pro., 農学部, 教授 (60001533)
INANAMI Osamu Grad.School of Vet.Med., Hokkaido Univ., Asso.Pro., 大学院・獣医学研究科, 助教授 (10193559)
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| Project Period (FY) |
1997 – 1998
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| Project Status |
Completed (Fiscal Year 1998)
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| Budget Amount *help |
¥14,500,000 (Direct Cost: ¥14,500,000)
Fiscal Year 1998: ¥1,900,000 (Direct Cost: ¥1,900,000)
Fiscal Year 1997: ¥12,600,000 (Direct Cost: ¥12,600,000)
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| Keywords | bovine leukocyte adhesion deficiency / BLAD / neutrophil / superoxide / signal transduction / NADPH oxidase / protein kinase / biodefense / 電子スピン共鳴法 / 生体防御機構 / ケミルミネッセンス / インテグリンファミリー / キナーゼ |
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| Research Abstract |
This project was performed to clarify the signal transduction mechanisms for NADPH oxidase activation and phagocytotis by using normal neutrophils and those with bovine leukocyte adhesion deficiency (BLAD) which was genetic deficient in beta2-integrin CR3 corresponding to the receptor of complement iC3b. Various reagents to inhibit NADPH oxidase-related signal transduction were used for this purpose. We found that inhibitors of protein kinase C (PKC), phosphatidyl inositol 3-kinase (PI 3-kinase) and p38 mitogen-activated protein kinase (p38 MAPK) were dose-dependently inhibited superoxide generation from serum-opsonized zymosan (s-OZ)-stimulated neutrophils from BLAD and normal calves, although stimulation of BLAD neutrophils with s-OZ brought about the lower generation of superoxide than that of normal neutrophils with s-OZ.These results indicated that the lack of beta2-integrin CR3 did not influence signal transduction pathways of NADPH oxidase but reduced superoxide production of NA
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DPH oxidase. This reduced NADPH oxidase activity in BLAD was partially recovered by transfusion of CD 18-positive granulocytes to diseased calf. Furthennore, PI 3-kinase and p38 MAPK but not PKC were shown to be required for phagocytotic activity in normal neutrophils. Concerning the intracellular mechanisms of NADPH oxidase activation, the p47phox, one component of NADPH oxidase, is known to be extensively phosphorylated at serines that are located among its C-terminal region and the phosphorylation is a trigger for the activation of NADPH oxidase. Using site-directed mutagenesis of p47phox and p47phox-deficient B cells from human chronic granulomatous disease (CGD), it was shown that the phosphorylation of serines 303/304, 359/370 and possibly serine 379 must take place in order to activate the oxidase. These results seem to be important in not only understanding the signal transduction mechanism of NADPH oxidase activity but also development for therapy of BLAD and p47phox-deficient CGD by the granulocyte or gene transfusion. Less
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