1999 Fiscal Year Final Research Report Summary
Functional coupling of arachidonate metabolizing enzymes
| Project/Area Number |
09557213
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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 |
応用薬理学・医療系薬学
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| Research Institution | Kitasato University |
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Principal Investigator |
OH-ISHI Sachiko Kitasato University, Professor, 薬学部, 教授 (70050416)
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| Co-Investigator(Kenkyū-buntansha) |
KUDO Ichiro Shouwa University, Professor, 薬学部, 教授 (30134612)
NARABA Hiroaki Kitasato University, Schhol of Parmac. Sci., Assistant Professor, 薬学部, 助手 (90296517)
UENO Akinori Kitasato University, Schhol of Parmac. Sci., Associate Professor, 薬学部, 助教授 (00112657)
KAMIMURA Takashi Teijin Co. Ltd., Institute of Biomedical Research, Senior Scientist, 生物医学研究所, 主任研究員
MURAKAMI Makoto Kitasato University, Schhol of Parmac. Sci., Associate Professor, 薬学部, 助教授 (60276607)
IKEDA Yuri Kitasato University, Schhol of Parmac. Sci., Assiatant Professor (10306657)
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| Project Period (FY) |
1997 – 1999
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| Keywords | Arachidonate Metabolites / Prostaglandins / PGE2 / Cyclooxygenase / PDE synthase / Inducible enzyme / Macrophages |
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| Research Abstract |
Prostaglandin E2 is known as one of the inflammatory mediators and appearing in the exudates of late phase of inflammation. We investigated functional coupling of the enzymes working in the arachidonate cascade to produce PGE2 by using rat peritonreal macrophages, when stimulated with lipopolysaccharide of E Coli Rat macrophages produced TXB2 and PGD2 at the initial phase of incubation, and the levels of these PGs did not change throughout the incubation upto 24 hr. On the contrary, PGE2 level increased with time and became 50-60 fold of the initial phase This increase was suppressed by COX-2 selective inhibitors, such as NS-398, to the initial level. When exogenous arachidonic acid was added to the stimulated macrophages, conversion to PGE2 peaked at 12 hr and the activity declined at 24 hr. However, Western blot of COX-2 indicates that COX-2 level increased upto 24 hr of incubation. Then we measured PGE synthase activity in the macrophage lysates by assessing the conversion of PGH2 to PGE2. This PGE synthase activity increased with incubation time and peaked around 12hr. Thus we conclude that an increase in inducible PGE synthase activity may be responsible for the increased PGE2 production in the LPS stimulated macrophages in addition to the induction of COX-2. We also cloned cDNAs of PGE synthases from rat and mouse macrophages and characterized the enzymes transfected into HEK 293 cells, respectively. We also constructed primers for Northern blot analysis of PGE synthases in mouse and rats. Northern blot analysis for PGE synthases of rat and mouse macrophages expressed that the PGE synthases in macrophages from both species were inducible enzyme to couple COX-2 and produce huge amount of PGE2.
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[Publications] Murata T, Ushikubi F, Matsuoka T, Hirata M, Yamasaki A, Sugimoto Y, Ichikawa A , Aze Y, Tanaka T, Yoshida N, Ueno A, Oh-ishi S, Narumiya S: "Altered pain perception and inflammatory response in mice lacking prostacyclin receptor"Nature. 388(6643). 678-682 (1997)
Description
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