| Project/Area Number |
10557224
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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 | 展開研究 |
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| Research Field |
Biological pharmacy
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| Research Institution | Showa University |
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Principal Investigator |
MURAKAMI Makoto School of Pharmaceutical Sciences, Showa University Assistant Professor, 薬学部, 助教授 (60276607)
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| Co-Investigator(Kenkyū-buntansha) |
SHIMBARA Satoko School of Pharmaceutical Sciences, Showa University Assistant, 薬学部, 助手 (60266161)
NAKATANI Yoshihito School of Pharmaceutical Sciences, Showa University Lecturer, 薬学部, 講師 (80266163)
YANOSHITA Rei School of Pharmaceutical Sciences, Showa University Assistant, 薬学部, 助手 (00224915)
CHIBA Shinsuke Japan energy Co., 研究員
KUWATA Hiroshi School of Pharmaceutical Sciences, Showa University Assistant, 薬学部, 助手 (80286864)
厚味 厳一 昭和大学, 薬学部, 助手 (70276608)
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| Project Period (FY) |
1998 – 2000
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| Project Status |
Completed (Fiscal Year 2000)
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| Budget Amount *help |
¥5,200,000 (Direct Cost: ¥5,200,000)
Fiscal Year 2000: ¥1,200,000 (Direct Cost: ¥1,200,000)
Fiscal Year 1999: ¥1,500,000 (Direct Cost: ¥1,500,000)
Fiscal Year 1998: ¥2,500,000 (Direct Cost: ¥2,500,000)
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| Keywords | Phosholipase A_2 / Mast cell / Degranulation / Eicosanoid / Arachidonic acid / Phosphatidycoline / PAF / AVRL-2 / 分化誘導 / 非IgE型刺激 |
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| Research Abstract |
In this study, we found that mast cells express all known members of the group II subfamily of secretory phospholipase A_2 (sPLA_2) isozymes and those having heparin affinity markedly enhance the exocytotic response. Rat mastocytoma RBL-2H3 cells transfected with heparin-binding (sPLA_2-IIA,-V and-IID), but not heparin-non-binding (sPLA_2-IIC), enzymes released more granule-associated markers (β-hexosaminidase and histamine) than mock-or cytosolic PLA_2α (cPLA_2α)-transfected cells after stimulation with IgE and antigen. Site-directed mutagenesis of sPLA_2-IIA and-V revealed that both the catalytic and heparin-binding domains are essential for this function. Confocal laser and electron microscopic analyses revealed that sPLA_2-IIA, which was stored in secretory granules in unstimulated cells, accumulated on the membranous sites where fusion between the plasma membrane and granule membranes occurred in activated cells. These results suggest that the heparin-binding sPLA_2s bind to the perigranular membranes through their heparin-binding domain, and lysophospholipids produced in situ by their enzymatic action may facilitate the ongoing membrane fusion. In contrast to the redundant role of sPLA_2-IIA,-IID and-V in the regulation of degranulation, only sPLA_2-V had the ability to markedly augment IgE/antigen-stimulated immediate production of lipid mediators such as prostaglandin D_2, leukotriene C_4 and platelet-activating factor (PAF), which reached a level comparable to that elicited by cPLA_2α. Analyses using sPLA_2 mutants as well as immunocytochemical studies demonstrated that the lipid mediator-producing ability of sPLA_2-V, and sPLA_2-X if artificially overexpressed, was attributed to their high binding ability, relative to other sPLA_2 isozymes, to phosphatidylcholine-rich plasma membrane surfaces.
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