title: CCN4/WISP-1 positively regulates chondrogenesis by controlling TGF-β3 function.
abstract: The CCN family of proteins plays important roles in development and homeostasis of bone and cartilage. To understand the role of CCN4 in chondrogenesis, human bone marrow stromal cells (hBMSCs) were transduced with CCN4 adenovirus (adCCN4) or siRNA to CCN4 (siCCN4) in the presence or absence of transforming growth factor-β3 (TGF-β3). Overexpression of CCN4 enhanced TGF-β3-induced SMAD2/3 phosphorylation and chondrogenesis of hBMSCs in an in vitro assay using a micromass culture model. On the other hand, knockdown of CCN4 inhibited the TGF-β3-induced SMAD2/3 phosphorylation and synthesis of cartilage matrix in micromass cultures of hBMSCs. Immunoprecipitation-western blot analysis revealed that CCN4 bound to TGF-β3 and regulated the ability of TGF-β3 to bind to hBMSCs. In vivo analysis confirmed there was a significant decrease in the gene expression levels of chondrocyte markers in cartilage samples from Ccn4-knock out (KO) mice, compared to those from wild type (WT) control. In order to investigate the regenerative properties of the articular cartilage in Ccn4-KO mice, articular cartilage defects were surgically performed in the knee joints of young mice, and the results showed that the cartilage was partially repaired in WT mice, but not in Ccn4-KO mice. In conclusion, these results show, for the first time, that CCN4 has a positive influence on chondrogenic differentiation by modulating the effects of TGF-β3.
authors.affiliation: Department of Oral Rehabilitation and Regenerative Medicine, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama, Japan.
authors.forename: Yuya
authors.initials: Y
authors.lastname: Yoshioka
authors.affiliation: Department of Oral Rehabilitation and Regenerative Medicine, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama, Japan; Department of Molecular Biology and Biochemistry, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama, Japan. Electronic address: mitsuaki@md.okayama-u.ac.jp.
authors.forename: Mitsuaki
authors.initials: M
authors.lastname: Ono
authors.affiliation: Department of Oral Rehabilitation and Regenerative Medicine, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama, Japan; Craniofacial and Skeletal Diseases Branch, National Institute of Dental and Craniofacial Research, National Institutes of Health, Bethesda, MD, USA.
authors.forename: Azusa
authors.initials: A
authors.lastname: Maeda
authors.affiliation: Craniofacial and Skeletal Diseases Branch, National Institute of Dental and Craniofacial Research, National Institutes of Health, Bethesda, MD, USA.
authors.forename: Tina M
authors.initials: TM
authors.lastname: Kilts
authors.affiliation: Department of Oral Rehabilitation and Regenerative Medicine, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama, Japan; Department of Biomaterials, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama, Japan.
authors.forename: Emilio Satoshi
authors.initials: ES
authors.lastname: Hara
authors.affiliation: Department of Oral Rehabilitation and Regenerative Medicine, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama, Japan; Advanced Research Center for Oral & Craniofacial Sciences, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama, Japan.
authors.forename: Hany
authors.initials: H
authors.lastname: Khattab
authors.affiliation: Department of Oral Rehabilitation and Regenerative Medicine, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama, Japan.
authors.forename: Junji
authors.initials: J
authors.lastname: Ueda
authors.affiliation: Advanced Research Center for Oral & Craniofacial Sciences, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama, Japan.
authors.forename: Eriko
authors.initials: E
authors.lastname: Aoyama
authors.affiliation: Advanced Research Center for Oral & Craniofacial Sciences, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama, Japan.
authors.forename: Toshitaka
authors.initials: T
authors.lastname: Oohashi
authors.affiliation: Advanced Research Center for Oral & Craniofacial Sciences, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama, Japan.
authors.forename: Masaharu
authors.initials: M
authors.lastname: Takigawa
authors.affiliation: Craniofacial and Skeletal Diseases Branch, National Institute of Dental and Craniofacial Research, National Institutes of Health, Bethesda, MD, USA.
authors.forename: Marian F
authors.initials: MF
authors.lastname: Young
authors.affiliation: Department of Oral Rehabilitation and Regenerative Medicine, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama, Japan.
authors.forename: Takuo
authors.initials: T
authors.lastname: Kuboki
mesh_terms: D000818:Animals; D015415:Biomarkers / Q000378:metabolism; D055511:CCN Intercellular Signaling Proteins / Q000235:genetics / Q000378:metabolism*; D002358:Cartilage, Articular / Q000473:pathology; D002454:Cell Differentiation; D002478:Cells, Cultured; D020219:Chondrogenesis / Q000235:genetics; D005786:Gene Expression Regulation; D006801:Humans; D059630:Mesenchymal Stem Cells / Q000166:cytology; D018345:Mice, Knockout; D010766:Phosphorylation; D011485:Protein Binding; D011518:Proto-Oncogene Proteins / Q000235:genetics / Q000378:metabolism*; D060888:Real-Time Polymerase Chain Reaction; D012038:Regeneration; D015398:Signal Transduction / Q000235:genetics; D051785:Smad Proteins / Q000378:metabolism; D053782:Transforming Growth Factor beta3 / Q000378:metabolism*; D014945:Wound Healing
chemical_list: D015415:Biomarkers; D055511:CCN Intercellular Signaling Proteins; C116396:CCN4 protein, human; C492067:CCN4 protein, mouse; D011518:Proto-Oncogene Proteins; D051785:Smad Proteins; D053782:Transforming Growth Factor beta3
keywords: Articular cartilage; Bone marrow stromal cell (BMSC); CCN4/WISP-1; Chondrogenesis; TGF-β3