CCN family 2/connective tissue growth factor (CCN2/CTGF) promotes osteoclastogenesis via induction of and interaction with dendritic cell-specific transmembrane protein (DC-STAMP).
CCN family 2/connective tissue growth factor (CCN2/CTGF) promotes endochondral ossification. However, the role of CCN2 in the replacement of hypertrophic cartilage with bone is still unclear. The phenotype of Ccn2 null mice, having an expanded hypertrophic zone, indicates that the resorption of the cartilage extracellular matrix is impaired therein. Therefore, we analyzed the role of CCN2 in osteoclastogenesis because cartilage extracellular matrix is resorbed mainly by osteoclasts during endochondral ossification. Expression of the Ccn2 gene was upregulated in mouse macrophage cell line RAW264.7 on day 6 after treatment of glutathione S transferase (GST) fusion mouse receptor activator of NF-κB ligand (GST-RANKL), and a combination of recombinant CCN2 (rCCN2) and GST-RANKL significantly enhanced tartrate-resistant acid phosphatase (TRACP)-positive multinucleated cell formation compared with GST-RANKL alone. Therefore, we suspected the involvement of CCN2 in cell-cell fusion during osteoclastogenesis. To clarify the mechanism, we performed real-time PCR analysis of gene expression, coimmunoprecipitation analysis, and solid-phase binding assay of CCN2 and dendritic cell-specific transmembrane protein (DC-STAMP), which is involved in cell-cell fusion. The results showed that CCN2 induced and interacted with DC-STAMP. Furthermore, GST-RANKL-induced osteoclastogenesis was impaired in fetal liver cells from Ccn2 null mice, and the impaired osteoclast formation was rescued by the addition of exogenous rCCN2 or the forced expression of DC-STAMP by a retroviral vector. These results suggest that CCN2 expressed during osteoclastogenesis promotes osteoclast formation via induction of and interaction with DC-STAMP.
Nishida
Takashi
T
Department of Biochemistry and Molecular Dentistry, Okayama University Graduate School of Medicine, Dentistry, and Pharmaceutical Sciences, Okayama, Japan.
Emura
Kenji
K
Kubota
Satoshi
S
Lyons
Karen M
KM
Takigawa
Masaharu
M
D000818:Animals; D002356:Cartilage / Q000378:metabolism; D002460:Cell Line; D055513:Connective Tissue Growth Factor / Q000378:metabolism*; D017931:DNA Primers / Q000235:genetics; D003712:Dendrites / Q000378:metabolism*; D005109:Extracellular Matrix / Q000378:metabolism; D005786:Gene Expression Regulation; D005982:Glutathione Transferase / Q000378:metabolism; D008099:Liver / Q000166:cytology; D008297:Male; D008565:Membrane Proteins / Q000378:metabolism*; D051379:Mice; D009419:Nerve Tissue Proteins / Q000378:metabolism*; D010010:Osteoclasts / Q000166:cytology*; D053245:RANK Ligand / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction
C526318:CCN2 protein, mouse; C499636:DC-STAMP protein, mouse; D017931:DNA Primers; D008565:Membrane Proteins; D009419:Nerve Tissue Proteins; D053245:RANK Ligand; C505689:Tnfsf11 protein, mouse; D055513:Connective Tissue Growth Factor; D005982:Glutathione Transferase