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1991 Fiscal Year Final Research Report Summary

Mechanism of Opioid Peptide Production in Cultured Fibroblasts of The Tooth Pulp.

Research Project

Project/Area Number 01480432
Research Category

Grant-in-Aid for General Scientific Research (B)

Allocation TypeSingle-year Grants
Research Field Functional basic dentistry
Research InstitutionOsaka University

Principal Investigator

KUDO Teruo  Osaka University Faculty of Dentistry, assist. Prof., 歯学部, 講師 (10028805)

Co-Investigator(Kenkyū-buntansha) MAEDA Sadaaki  Osaka University Faculty of Dentistry, Research Instructor, 歯学部, 助手 (00135732)
YONEHARA Norifumi  Osaka University Faculty of Dentistry, Research Instructor, 歯学部, 助手 (70124534)
SAITO Kihachi  Osaka University Faculty of Dentistry, Assoc. Prof., 歯学部, 助教授 (40110788)
Project Period (FY) 1989 – 1991
KeywordsTooth Pulp / Fibroblasts / Bradykinin / Des-Arg^9-Bradykinin / Arginine / Enkephalins / Cathepsin B / Protein Phosphorylation
Research Abstract

The present study was aimed to investigate the mechanism of enkephalin (EK) production induced b. y bradykinin (BK) in dental pulp of the rat, using subcellular fractions or cultured fibroblasts of the rat incisor pulp tissue. In experiments using subcellular fractions, it was found that (1) the en2yme activity of EK production was localized in lysosomal fraction and (2) EK precursor proteins were conatined in microsomal and supernatant fractions. Thereafter, EK producing enzyme was isolated from the lysosomal fraction and purified. (3) lt was demonstrated that the en2yme was a cysteine proteinase, cathepsin B (M. W. ; 23, 600). On the other hand, EK precursor protein was isolated from the microsomal and supernatant fractions. and purified. (4) lt was certified that a common protein (M. W. ; 58, 000) in both fractions was a major precursor protein. (5) The purified cathepsin B was not activated by BK but the enzyme in entire lysosomal structure was activated by it. In experiments using … More pulpal cells, fibroblasts were easily cultured from the pulp tissue. (6) lt was immunocytochemically or immunochemically demonstrated that the cells contained EKs and their precursor protein. (7) lncubation of the cells with BK, des-Arg9-BK, or arginine (Arg) increased cathepsin B activity of the cells, thereby EKs were produced. (8) The actions of these agents was dependent on concentration of intra- and extracelluar Ca^<++>. In experiments using the fibroblasts with various enzyme inhibitors and antagonist, (9) it was found that the actions of BK and des-Arg^9-BK were mediated through B_1-receptor coupled with an islet-activating protein (IAP)-sensitive GTP-binding protein (G-protein), followed by activation of phospholipase C (PLC), a key en2yme in cathepsin B activation. The activation of PLC induced an elevation of intracellular Ca^<++> concentration, followed by activation of various protein kinases, such as protein kinase (PK) C, Ca^<++>/calmodulin-dependent PK and CGMPdependent PK. It was also found that (10) IAP-sensitive G-protein was not involved in activation of cathepsin B by Arg, but Arg was uptaken into the cells to form kyotorphin (KTP), an EK releaser, which acted on KTP receptor of cell membrane, resulting in activation of cathepsin B, and in addition, to form nitric oxide (NO), an activator of guanylate cyclase, resulting in production of CGMP. In conclusion, it was considered that BK action involved in cathepsin B activation of the fibroblasts was due to a combination effect of des-Arg^9-BK and Arg which were produced by an enzymatic degradation of BK by which various protein phosphorylation steps were induced. Less

  • Research Products

    (22 results)

All Other

All Publications (22 results)

  • [Publications] Kudo,T.: "Inflammatory pain and its regulation in the tooth pulp." J.Clin.Expt1.Med.(IGAKU NO AYUMI). 161(2). (1992)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Zhu,B.-F.,Kudo,T.,Maeda,S.,and Inoki,R.: "Protein kinases involved in the activation of cathepsin B in fibroblasts of the tooth pulp by bradykinin." Pain Research. 7. (1992)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Zhu,B.-F.,Kudo,T.,Maeda,S.,and Inoki,R.: "Involvement of GTP-binding proteins in activation of cathepsin B by bradykinin in fibroblasts of the tooth pulp." Pain Research. 7. (1992)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Kudo,T.,Zhu,B.-F.,and Inoki,R.: "Ca^<++>-dependency of bradykinin-stimulated enkephalin production in fibroblasts of the tooth pulp." Pain Research. 6. 105-110 (1991)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Wei,E.-Q.,Kudo,T.,and Inoki,R.: "subcellular distribution of enkephalin precursor proteins in rat dental pulp and the usefulness as a substrate for enkephalin-producing enzymes." Life Sciences. 48. 667-674 (1991)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Wei,E.-Q.,Kudo,T.,and Inoki,R.: "Pharmacological and biochemical study on the mechanism of enkephalin production in rat dental pulp." J.Osaka Univ.Dent.Sch.30. 8-30 (1990)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Inoki,R.,Kudo,T.,and Olgart,L.(eds): "Dynamic Aspects of Dental Pulp." Chapman and Hall,London,508 (1990)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Wei, E. -Q., Bian, R. -L., Kudo, T. and Lnoki, R.: "Interaction between bradykinin and enkephalins in the rat dental pulp. (in chinese)" Acta Pharmacol. Sinica. 10. 500-503 (1989)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Kudo, T., Wei, E. -Q. and Inoki, R.: "Activation of calcium ion-dependent proteinases by bradykinin in dental pulp of the rat." Adv. Exp. Med. and Biol.247B. 627-631 (1989)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Kudo, T., Wei, E. -Q., Zhu, B. -F. and Inoki, R.: "Involvement of lysosomal enzyme in enkephalin production in dental pulp of the rat." Dentistry in Japan. 26. 41-44 (1989)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Kudo, T., Wei, E. -Q., Zhu, B. -F. and Inoki, R.: "Stabilizing effect of prostaglandin E_2 on lysosomal membrane of the rat dental pulp." J. Osaka Univ. Dent. Soc.35. 13-21 (1990)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Kudo, T., Wei, E. -Q. and Inoki, R.: "Characteristics of BANA-degrading enzyme in the tooth pulp activated by bradykinin." Progr. Clin. Biol. Res.328. 275-278 (1990)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Kudo, T., Wei, E. -Q. and Inoki, R.: "Is cathepsin B an enkephalin processing enzyme in the tooth pulp?" Progr. Clin. Biol. Res.328. 271-274 (1990)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Inoki, R. and Kudo, T.: "Bradykinin and enkephalins." Dynamic Aspects of Dental Pulp.edited by Inoki, R., Kudo, T. and Olgart, L., Chapman and Hall, London. 385-401 (1990)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Kudo. T., Wei, E. -Q. and Inoki, R.: "Proteases in inflammation." ibid.445-470 (1990)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Kudo, T., Wei, E. -Q., Zhu, B. -F. and Inoki, R.: "Enkephalin production in tooth pulp cells by cathepsin B." New Leads in Opioid Research, edited by J. M. Van Ree, et al.Excerpta Medica, Amsterdam. 27-29 (1990)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Wei, E. -Q., Kudo, T. and Inoki, R.: "Pharmacological and biochemical study on the mechanism of enkephalin production in rat dental pulp." J. Osaka Univ. Dent. Sch.30. 8-30 (1990)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Wei, E. -Q., Kudo, T. and Inoki, R.: "Subcellular distribution of enkephalin precursor proteins in rat dental pulp and the usefulness as a substrate for enkepnhalin-producing enzymes." Life Sci.48. 667-674 (1991)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Kudo, T., Zhu, B. -F. and Inoki, R.: "Ca^<++> dependency of bradykinin-stimulated enkephalin production in fibroblasts of the tooth pulp." Pain Res.6. 105-110 (1991)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Zhu, B. -F., Kudo, T., Maeda, S. and Inoki, R.: "Involvement of GTP-binding proteins in activation of cathepsin B by bradykinin in fibroblasts of the tooth pulp." Pain Res.7. (1992)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Zhu, B. -F., Kudo. T., Maeda, S. and Inoki, R.: "Protein kinases involved in the activation of cathepsin B in fibroblasts of the tooth pulp by bradykinin." Pain Res.7. (1992)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Kudo, T.: "Inflammatory pain and its regulation in the tooth pulp." J. Clin. Exptl. Med.161(2). (1992)

    • Description
      「研究成果報告書概要(欧文)」より

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Published: 1993-03-16  

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