Apoptosis via new serine proteases and its intracellular signaling events.
| Project/Area Number |
15590682
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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 |
Gastroenterology
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| Research Institution | Saitama Medical School |
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Principal Investigator |
NAKAYAMA Nobuaki Saitama Medical School, Department of Medicine, Assistant Professor, 医学部, 講師 (40292015)
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| Co-Investigator(Kenkyū-buntansha) |
MATSUI Atsushi DSaitama Medical School, Department of Medicine, Assistant Professor, 医学部, 講師 (40260484)
MOCHIDA Satishi Saitama Medical School, Department of Medicine, Professor, 医学部, 教授 (20219968)
NAGOSHI Sumiko Saitama Medical School, Department of Medicine, Associate Professor, 医学部, 助教授 (50306271)
LNAO Mie Saitama Medical School, Department of Medicine, Assistant Professor, 医学部, 講師 (70286037)
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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 |
¥3,400,000 (Direct Cost: ¥3,400,000)
Fiscal Year 2004: ¥1,400,000 (Direct Cost: ¥1,400,000)
Fiscal Year 2003: ¥2,000,000 (Direct Cost: ¥2,000,000)
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| Keywords | Apoptosis / HepG2 cells / Ethanol / Acetaminophen / Serine protease |
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| Research Abstract |
We used HepG2 cells as a model system for human liver cells. We investigated intracellular signaling events of acetaminophen-induced apoptosis in HepG2 cells. Acetaminophen induces apoptosis in HepG2 cells in a dose- and time-dependent manner. It was efficiently inhibited by the caspase inhibitor N-benzyloxycarbonyl-Asp-Glu-Val-fluoromethylketone (zVAD-fink) and the serine protease inhibitor N-p-tosyl-_L-lysine chloromethyl ketone (TLCK). Loss of the mitochondrial transmembrane potential (ΔΨm) reached a plateau after treatment for 24 h. TLCK could inhibit reduction of ΔΨm partially. For measurement of serine protease activity in cell lysates, Boc-Glu-Lys-Lys-MCA was selected as the best substrate among various peptidyl-MCA substrates because of its high sensitivity and specificity. Affinity chromatography was carried out in order to purify serine proteases from acetaminophen-treated HepG2 cells. We could not obtain any fraction with high protease activity by affinity chromatography on SBTI-immobilized Sepharose. Lysine-immobilized Sepharose was next utilized, because plasmin-specific peptidyl-MCA substrates showed high protease activity in acetaminophen-treated cell extract. One fraction of high protease activity was obtained and submitted to SDS-PAGE and silver staining. These data suggest that acetaminophen-induced apoptosis in liver cells is initiated activation of TLCK-sensitive serine proteases and that these proteases might mimic plasminogen structurally.
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Report
(3 results)
Research Products
(8 results)
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[Journal Article] Japanese herbal medicine Inchin-ko-to as a therapeutic drug for liver fibrosis2004
Author(s)
Inao M, Mochida S, Matsui A, Eguchi Y, Yulutuz Y, Wang Y, Naiki K, Kakinuma T, Fujimori K, Nagoshi S, Fujiwara K
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Journal Title
J Hepatol 41(4)
Pages: 584-591
Description
「研究成果報告書概要(欧文)」より
Related Report
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