| Project/Area Number |
17590193
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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 |
General physiology
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| Research Institution | Kinki University |
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Principal Investigator |
MATSUO Osamu Kinki University, medicine, Professor, 医学部, 教授 (40030879)
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| Co-Investigator(Kenkyū-buntansha) |
UESHIMA Shigeru Kinki University, Agriculture, Professor, 農学部, 教授 (30193791)
OKADA Kiyotaka Kinki University, medicine, Lecture, 医学部, 講師 (20185432)
KAWAO Naoyuki Kinki University, medicine, Assistant, 医学部, 助手 (70388510)
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| Project Period (FY) |
2005 – 2006
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| Project Status |
Completed (Fiscal Year 2006)
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| Budget Amount *help |
¥3,400,000 (Direct Cost: ¥3,400,000)
Fiscal Year 2006: ¥1,600,000 (Direct Cost: ¥1,600,000)
Fiscal Year 2005: ¥1,800,000 (Direct Cost: ¥1,800,000)
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| Keywords | biotechnology / angiogenesis / proteolysis activity / gene expression regulation / t-PA / t-PAR system / 蛋白分解活性 |
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| Research Abstract |
We have previously demonstrated that tissue-type plasminogen activator receptor (t-PAR) purified from human umbilical vein endothelial cells specifically interacted with t-PA. Following these data, we have produced recombinant t-PAR expressed in E.coli. t-PAR bound to t-PA specifically, which was identified by ligand blot analysis. In the present study, to investigate the physiological role of t-PAR on cell surface, we have stably transfected endothelial cells (ECs) with a myc/His-tagged t-PAR expression plasmid. Expression of both the mRNA and t-PAR/myc/His fusion protein was confirmed by semi-quantitative PCR and western blotting.
The binding ability of t-PA to t-PAR-overexpressed ECs was studied by using IAsys. Constitutive expression of t-PAR increased binding signal of ECs to t-PA compared with control (LacZ-transfected) ECs. The DFP-inactivated t-PA did not give any effect on the interaction to t-PAR. It was suggested that the binding of t-PA to t-PAR on the surface of cells was independent on catalytic activity of t-PA. On the fibrinolytic activity using chromogenic assay, t-PAR-overexpressed ECs showed marked plasminogen activator (PA) activity after the addition of t-PA. This t-PAR-induced PA activity could be inhibited by the addition of antibodies against t-PA, suggesting that t-PAR might concentrate t-PA on the surface of cell and enhanced the fibrinolytic properties around cells. Furthermore, we have demonstrated that t-PAR promoted proliferation of EC after the addition of t-PA. Thus, t-PAR could mediate proliferation of ECs via signal transduction pathway.
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