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Assessment of CCN2 Independent Modules Regenerative Capacity on Osteoarthritis and Further Selecting the Most Suitable Among them as a Potential Therapeutic Drug
Author(s) -
Abdelkader Tarek,
Aoyama Eriko,
Nishida Takashi,
Hattori Takako,
Janune Danilo,
Hara Emilio S.,
Ono Mitsuaki,
Tabata Yasuhiko,
Kuboki Takuo,
Kubota Satoshi,
Takigawa Masaharu
Publication year - 2016
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.30.1_supplement.1070.2
Subject(s) - ctgf , cartilage , microbiology and biotechnology , osteoarthritis , matrigel , chemistry , regeneration (biology) , in vivo , growth factor , in vitro , morphogen , ex vivo , biology , anatomy , biochemistry , pathology , medicine , gene , receptor , alternative medicine
CCN family proteins ( C onnective tissue growth factor (CTGF)/CCN2, C ystein rich protein (Cyr61)/CCN1, and N ephroblastoma overexpressed gene (nov)/CCN3) play important roles in differentiation and regeneration of cartilaginous tissues. CCN2/CTGF is known to promote the regeneration of articular cartilage. It's comprised of 4 highly interactive independent modules, each of which was suggested to have its own biological activity. This study aims to assess the effects of these modules independently and their combinations on chondrocytic cells in vitro and damaged cartilage in vivo , especially in relation to full length CCN2. Methods Effect of a single module, or their different combination, was evaluated with human chondrocytic HCS‐2/8 cells. The cellular phenotype was estimated by the gene expression of chondrocytic markers. Proteoglycan synthesis and proliferation were evaluated by [ 35 S]‐sulfate and [ 3 H]‐thymidine incorporation assays, respectively. Physical interaction of 2 modules was kinetically examined by a surface plasmon resonance (SPR) methodology. Cartilage regeneration in vivo was evaluated by using 2 rat models simulating osteoarthritis via surgical or chemical intervention. Angiogenic affinity of independent modules was assessed in vitro by tube formation assay using matrigel with human umbilical endothelial cells (HUVEC) and ex ovo by CAM (chorioallantoic membrane) assay with chick embryo. Results Functional analysis in vitro revealed a biological activity even stronger than the full length CCN2 in a particular module. Interestingly, mixed application of all 4 modules almost reconstructed the bioactivity to the level of the full length. SPR analysis uncovered significant interaction of 2 modules and full‐length CCN2. The result of cartilage regeneration experiments in vivo was also consistent with the findings obtained in vitro . The results of the matrigel assay indicate that the same module has tube formation potential under special circumstances and a similar effect could be observed in the CAM assay ex ovo . Conclusion These results indicate independent bioactivity of each module for cartilage regeneration and suggest unknown means of binding to construct a possible tetramodular complex that is functionally comparable to the full length CCN2. Also the effect of the proposed independent module as a regenerative drug on angiogenesis must be kept in mind and further analyzed as this characteristic might be beneficial for vascular regeneration in spite the fact that it's considered a side effect for a proposed OA treatment.