title: Generation and Analysis of Cartilage-Specific CCN2 Overexpression in Transgenic Mice.
abstract: Recent progress in gene-editing technology has provided a strong impact for improved our understanding of molecular functions in living organisms. Here we describe our method to generate transgene-overexpressing mouse models, which method involves the use of tissue-specific promoters for analyzing a certain molecule (s) in special tissues. The protocol described in this chapter uses the Col2a1 promoter-enhancer, which is known for driving specific and strong transgene expression in cartilage and is based on several of our studies showing a positive role of the connective tissue growth factor (CCN2) in cartilage-bone development and maintenance of articular cartilage. These mice show strongly accelerated endochondral ossification resulting in enhanced bone elongation, as well as resistance to age-related articular degeneration. This protocol also describes how to analyze the molecular mechanisms of these phenomena by use of chondrocytes isolated from CCN2-overexpressing cartilage.
authors.affiliation: Department of Biochemistry & Molecular Dentistry, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, 2-5-1, Shikata-cho, Kita-ku, Okayama, 700-8525, Japan. hottorit@md.okayama-uc.ac.jp.
authors.forename: Takako
authors.initials: T
authors.lastname: Hattori
authors.affiliation: Department of Orthodontics and Dentofacial Orthopedics, Osaka University Graduate School of Dentistry, Osaka, Japan.
authors.forename: Shinsuke
authors.initials: S
authors.lastname: Itoh
authors.affiliation: Advanced Research Center for Oraland Craniofacial Sciences, Okayama University Dental School/Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, 2-5-1, Shikata-cho, Kita-ku, Okayama, 700-8525, Japan. takigawa@md.okayama-u.ac.jp.
authors.forename: Masaharu
authors.initials: M
authors.lastname: Takigawa
mesh_terms: D000818:Animals; D002358:Cartilage, Articular / Q000378:metabolism*; D019902:Chondrocytes / Q000378:metabolism; D020219:Chondrogenesis / Q000235:genetics; D055513:Connective Tissue Growth Factor / Q000235:genetics*; D015870:Gene Expression; D023061:Gene Order; D017930:Genes, Reporter; D005822:Genetic Vectors / Q000235:genetics; D005838:Genotype; D051379:Mice; D008822:Mice, Transgenic; D009928:Organ Specificity / Q000235:genetics; D011401:Promoter Regions, Genetic; D001616:beta-Galactosidase / Q000235:genetics
chemical_list: D055513:Connective Tissue Growth Factor; D001616:beta-Galactosidase
keywords: Aging; Cartilage; Col2a1 promoter-enhancer; Development; Endochondral ossification; Histochemistry; IGF; LacZ staining; Skeletal preparation; Transgenic