Effects of CTGF/Hcs24, a product of a hypertrophic chondrocyte-specific gene, on the proliferation and differentiation of chondrocytes in culture.
Recently, we cloned a messenger RNA (mRNA) predominantly expressed in chondrocytes from a human chondrosarcoma-derived chondrocytic cell line, HCS-2/8, by differential display PCR and found that its gene, named hcs24, was identical with that of connective tissue growth factor (CTGF). Here we investigated CTGF/Hcs24 function in the chondrocytic cell line HCS-2/8 and rabbit growth cartilage (RGC) cells. HCS-2/8 cells transfected with recombinant adenoviruses that generate CTGF/Hcs24 sense RNA (mRNA) proliferated more rapidly than HCS-2/8 cells transfected with control adenoviruses. HCS-2/8 cells transfected with recombinant adenoviruses that generate CTGF/Hcs24 sense RNA expressed more mRNA of aggrecan and type X collagen than the control cells. To elucidate the direct action of CTGF/Hcs24 on the cells, we transfected HeLa cells with CTGF/Hcs24 expression vectors, obtained stable transfectants, and purified recombinant CTGF/Hcs24 protein from conditioned medium of the transfectants. The recombinant CTGF/Hcs24 effectively promoted the proliferation of HCS-2/8 cells and RGC cells in a dose-dependent manner and also dose dependently increased proteoglycan synthesis in these cells. In addition, these stimulatory effects of CTGF/Hcs24 were neutralized by the addition of anti-CTGF antibodies. Furthermore, the recombinant CTGF/Hcs24 effectively increased alkaline phosphatase activity in RGC cells in culture. Moreover, RT-PCR analysis revealed that the recombinant CTGF/Hcs24 stimulated gene expression of aggrecan and collagen types II and X in RGC cells in culture. These results indicate that CTGF/Hcs24 directly promotes the proliferation and differentiation of chondrocytes.
Nakanishi
T
T
Department of Biochemistry and Molecular Dentistry, Biodental Research Center, Okayama University Dental School, Japan.
Nishida
T
T
Shimo
T
T
Kobayashi
K
K
Kubo
T
T
Tamatani
T
T
Tezuka
K
K
Takigawa
M
M
D000256:Adenoviridae / Q000235:genetics; D000818:Animals; D015153:Blotting, Western; D002356:Cartilage / Q000166:cytology; D002454:Cell Differentiation / Q000187:drug effects; D002455:Cell Division / Q000187:drug effects; D002478:Cells, Cultured; D019902:Chondrocytes / Q000166:cytology* / Q000648:ultrastructure; D003094:Collagen / Q000096:biosynthesis; D055513:Connective Tissue Growth Factor; D004273:DNA, Neoplasm / Q000096:biosynthesis; D006133:Growth Substances / Q000235:genetics* / Q000494:pharmacology*; D006801:Humans; D017874:Immediate-Early Proteins; D036341:Intercellular Signaling Peptides and Proteins; D011509:Proteoglycans / Q000096:biosynthesis; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics; D011817:Rabbits; D011994:Recombinant Proteins / Q000494:pharmacology; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014162:Transfection
C526317:CCN2 protein, human; D004273:DNA, Neoplasm; D006133:Growth Substances; D017874:Immediate-Early Proteins; D036341:Intercellular Signaling Peptides and Proteins; D011509:Proteoglycans; D012333:RNA, Messenger; D011994:Recombinant Proteins; D055513:Connective Tissue Growth Factor; D003094:Collagen