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Apoptosis via new serine proteases and its intracellular signaling events.

Research Project

Project/Area Number 15590682
Research Category

Grant-in-Aid for Scientific Research (C)

Allocation TypeSingle-year Grants
Section一般
Research Field Gastroenterology
Research InstitutionSaitama Medical School

Principal Investigator

NAKAYAMA Nobuaki  Saitama Medical School, Department of Medicine, Assistant Professor, 医学部, 講師 (40292015)

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)
Project Period (FY) 2003 – 2004
Project Status Completed (Fiscal Year 2004)
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)
KeywordsApoptosis / HepG2 cells / Ethanol / Acetaminophen / Serine protease
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.

Report

(3 results)
  • 2004 Annual Research Report   Final Research Report Summary
  • 2003 Annual Research Report
  • Research Products

    (8 results)

All 2004 2003 Other

All Journal Article (7 results) Publications (1 results)

  • [Journal Article] Transgenic mice expressing osteopontin in hepatocytes as a model of autoimmune hepatitis.2004

    • Author(s)
      Mochida S, Yoshimoto T, et al.
    • Journal Title

      Biochem Biophys Res Commun 317

      Pages: 114-120

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2004 Final Research Report Summary
  • [Journal Article] Japanese Harbal Medicine Inchin-Ko-To as a Therapeutic drug for Liver Fibrosis2004

    • Author(s)
      Inao M, Mochida S, Matsui A, et al.
    • Journal Title

      Journal of Hepatology 41

      Pages: 584-591

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2004 Annual Research Report 2004 Final Research Report Summary
  • [Journal Article] Transgenic mice expressing osteopontin in hepatocytes as a model of autoimmune hepatitis2004

    • Author(s)
      Mochida S, Yoshimoto T, Mimura S, Inao M, Matsui A, Ohno A, Koh H, Saitoh E, Nagoshi S, Fujiwara K
    • Journal Title

      Biochem Biophys Res Commun 317(1)

      Pages: 114-120

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2004 Final Research Report Summary
  • [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
    • 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
      2004 Final Research Report Summary
  • [Journal Article] 薬剤性肝細胞障害におけるアポトーシスの解析2003

    • Author(s)
      中山 伸朗 他
    • Journal Title

      肝臓 44・Suppl(1)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2004 Final Research Report Summary
  • [Journal Article] Apoptosis in drug-induced liver injuries2003

    • Author(s)
      Nakayama N, Matsui A, Nagoshi S, Mochida S, Nakajima J, Koike T, Fujiwara K
    • Journal Title

      Acta Hepatol Jpn 44 Suppl.1

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2004 Final Research Report Summary
  • [Journal Article] 薬剤性肝細胞障害におけるアポトーシスの解析2003

    • Author(s)
      中山 伸朗
    • Journal Title

      肝臓 44・Suppl(1)

    • Related Report
      2004 Annual Research Report
  • [Publications] 中山 伸朗: "薬剤性肝細胞障害におけるアポトーシスの解析"肝臓. 44・Suppl(1). A90 (2003)

    • Related Report
      2003 Annual Research Report

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Published: 2003-04-01   Modified: 2016-04-21  

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