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Gene Expression and Distribution of Connective Tissue Growth Factor (CCN2/CTGF) During Secondary Ossification Center Formation

Morihiko Oka, Satoshi Kubota, Seiji Kondo, Takanori Eguchi, Chisa Kuroda, Kazumi Kawata, Shogo Minagi and Masaharu Takigawa

Department of Biochemistry and Molecular Dentistry (MO,SKubota,SKondo,TE,CK,KK,MT) and Department of Occlusal and Oral Functional Rehabilitation (MO,CK,KK), Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama University Dental School, Okayama, Japan

Correspondence to: Masaharu Takigawa, Department of Biochemistry and Molecular Dentistry, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, 2-5-1 Shikata-cho, Okayama 700-8525, Japan. E-mail: takigawa{at}md.okayama-u.ac.jp

CCN2/connective tissue growth factor (CCN2/CTGF) is a critical signaling modulator of mesenchymal tissue development. This study investigated the localization and expression of CCN2/CTGF as a factor supporting angiogenesis and chondrogenesis during development of secondary ossification centers in the mouse tibial epiphysis. Formation of the secondary ossification center was initiated by cartilage canal formation and blood vessel invasion at 7 days of age, and onset of ossification was observed at 14 days. In situ hybridization showed that CCN2/CTGF mRNA was distinctively expressed in the region of the cartilage canal and capsule-attached marginal tissues at 7 days of age, and distinct expression was also observed in proliferating chondrocytes around the marrow space at 14 days of age. Immunostaining showed that CCN2/CTGF was distributed broadly around the expressed cells located in the central region of the epiphysis, where the chondrocytes become hypertrophic and the cartilage canal enters into the hypertrophic mass. Furthermore, an overlapping distribution of metalloproteinase (MMP)9 and CCN2/CTGF was found in the secondary ossification center. These findings suggest that the CCN2/CTGF is involved in establishing epiphyseal vascularization and remodeling, which eventually determines the secondary ossification center in the developing epiphysial cartilage. (J Histochem Cytochem 55:1245–1255, 2007)

Key Words: CCN2 • connective tissue growth factor • secondary ossification center • CCN family • endochondral ossification

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