| Project/Area Number |
15390024
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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 |
Biological pharmacy
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| Research Institution | Mukogawa Women's University |
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Principal Investigator |
ICHIKAWA Atsushi Mukogawa Women's University, School of Pharmaceutical Sciences, Professor, 薬学部, 教授 (10025695)
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| Co-Investigator(Kenkyū-buntansha) |
SENMA Masanori Mukogawa Women's University, School of Pharmaceutical Sciences, Professor, 薬学部, 教授 (90098533)
SUGIMOTO Yukihiko Kyoto University, Associate Professor, 薬学研究科, 助教授 (80243038)
TANAKA Satoshi Kyoto University, Lecturer, 薬学研究科, 助手 (40303846)
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| Project Period (FY) |
2003 – 2004
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| Project Status |
Completed (Fiscal Year 2004)
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| Budget Amount *help |
¥15,300,000 (Direct Cost: ¥15,300,000)
Fiscal Year 2004: ¥7,400,000 (Direct Cost: ¥7,400,000)
Fiscal Year 2003: ¥7,900,000 (Direct Cost: ¥7,900,000)
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| Keywords | mast cells / cell adhesion / prostaglandin E2 / Extracellular matrix / cAMP / Epac / Ca2+ / PGE2 / 細胞外マトリックス / PKA / mastocytoma P-815 / Epac-Rapシグナル / EP4サブタイプ / フィブロネクチン / Ca^<2+> / アレルギー |
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| Research Abstract |
The purpose of this project is to determine the mechanism of mast cell adhesion to fibronectin. Mastocytoma P-815 cells were cultured on a plate pre-coated with or without fibronectin in 5%-FCS containing Fisher medium, and incubated with PGE2 for various times at 37℃ in 5% C02-incubator. Obtained results are shown below ;
1.P-815 cells were adhered to fibronectin in response to 10^<-8> M-10^<-6> M PGE2. The adherence activity of PGE2 was appeared through its binding to EP4 receptors expressed on P-815 cells, followed by a stimulated cAMP formation and cAMP-dependent protein kinase A activity. These responses were not demonstrated up to 4-5 hrs after PGE2 addition, indicating involvement of newly formed unknown proteins, which have been searching thereafter.
2.After initiation of PGE2-induced P-815 cell adhesion to fibronectin, being inhibited by PKA inhibitor H-89, adhered P-815 cells did not leak out by H89 treatment. However, PGE2-adhered P-815 cells were easily detached by a decrease of increasing intracellular Ca2+ level by treatment with EGTA,BAPTA/AM, and SKF96365, a Ca2+ channel ROCC inhibitor. Really, adhered P-815 cells were found to consist of high intracellular Ca2+ level.
3.PGE2-activated P-815 cell adhesion to fibronectin were mostly suppressed by incubation with CaM kinase inhibitor.
4.CAMP/PKA-independent signals were extensively searched for, and as a result have determined involvement of Epac signal (exchange protein directly activated by cAMP) as important candidatet molecules.
From these results, we have presented that PGE2-activated P-815 cell adhesion might be comprised of two signals, first during binding initiation phase cAMP/PKA dependent reactions involved in while during late binding maintenance phase Epac reactions did. These results are surely important to resolve mast cell adhesion ability to extra-cellular matrix like ibronectin in local inflammatory tissues.
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