1999 Fiscal Year Final Research Report Summary
Molecular cloning of the receptors for chondrosarcoma-derived chondrocyte growth factor Ecogenin/CTGF
| Project/Area Number |
09671893
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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 |
Functional basic dentistry
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| Research Institution | Okayama University |
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Principal Investigator |
NAKANISHI Tohru Okayama University Dental School, Associate Professor, 歯学部, 助教授 (30243463)
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| Co-Investigator(Kenkyū-buntansha) |
OHYAMA Kazumi Okayama University Dental School, Research Assistant, 歯学部, 教務員 (00253021)
HATTORI Takako Okayama University Dental School, Instructor, 歯学部, 助手 (00228488)
TAKIGAWA Masaharu Okayama University Dental School, Professor, 歯学部, 教授 (20112063)
INOUE Miho Okayama University Dental School, Research Assistant, 歯学部, 教務員 (20271059)
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| Project Period (FY) |
1997 – 1999
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| Keywords | differential display / connective tissue growth factor (CTGF) / crosslink / affinity chromatography / MAP kinase / proteoglycan / integrin / tyrosine kinase |
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| Research Abstract |
(1) Connective tissue growth factor (CTGF) was cloned from a chondrocyte-derived chondrocytic cell line, HCS-2/8 by differential display-PCR.
(2) Recombinant CTGF (rCTGF) stimulated the proliferation, maturation and differentiation of chondrocytes.
(3) Two types of receptors for CTGF were found on a chondrocyte-derived chondrocytic cell line, HCS-2/8. The receptor with high affinity was supposed to be cell adhesion molecules including integrins. The receptor with low affinity was supposed to be extracellular matrix compounds including proteoglycans.
(4) The inhibitory experiments using signal inhibitors showed that intercellular signal transduction in HCS-2/8 cells caused by the stimulation of CTGF was mediated by MAP kinase-pathways including MEK and ERK.
(5) CTGF-binding proteins were purified from membrane fractions and cytoplasmic fractions of HCS-2/8 cells with CTGF-conjugated affinity chromatography. As a result, four binding proteins (34, 44, 66 kDa from membrane fractions 50 kDa from cytoplasmic fractions) were purified from HCS-2/8 cells. The expression of these four proteins were regulated by the stimulation of CTGF, suggesting that they were functionally associated with CTGF.
(6) The cDNA of tyrosine kinase-type receptors were cloned from HCS-2/8 cells using degenerate primers corresponding to the consensus sequences between tyrosine kinase-domains. The sequence analysis of these cDNA revealed that there were several types of tyrosine kinase-receptors including novel receptors in HCS-2/8 cells. The expression of these receptors were regulated by the stimulation of CTGF. In addition, CTGF-producing cells showed high level of expression of these receptors. These results suggest that they were functionally associated with CTGF.
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