Spontaneous differentiating primary chondrocytic tissue culture: a model for endochondral ossification

• Published in: Bone (New York, N.Y.) N.Y.), 2002-08, Vol.31 (2), p.333-339
• Main Authors: Reiter, I, Tzukerman, M, Maor, G
• Format: Article
• Language: English
• Subjects: Animals; Biological and medical sciences; Cell Differentiation - physiology; Cells, Cultured; Chondrocytes; Chondrocytes - cytology; Chondrocytes - physiology; Collagen type II; Collagen type X; Core Binding Factors; Core-binding factor α 1; Culture Techniques - methods; Fundamental and applied biological sciences. Psychology; Insulin-like growth factor-1; Mandibular Condyle - cytology; Mandibular Condyle - physiology; Mice; Mice, Inbred ICR; Neoplasm Proteins; Osteogenesis - physiology; Receptor, IGF Type 1 - biosynthesis; Skeleton and joints; Tissue culture; Transcription Factors - biosynthesis; Vertebrates: osteoarticular system, musculoskeletal system
• ISSN: 8756-3282; 1873-2763
• DOI: 10.1016/S8756-3282(02)00823-2

Primary cartilage-derived cell cultures tend to undergo dedifferentiation, acquire fibroblastic features, and lose most of the characteristics of mature chondrocytes. This phenomenon is due mainly to the close matrix-cell interrelationship typical of cartilage tissue, which is vital for the preservation of the cartilaginous features. In this study we present a model for spontaneous redifferentiation of primary chondrocytic culture. Mandibular condyles excised from 3-day-old mice, thoroughly cleaned of all soft tissue, were digested with 0.1% collagenase. These mandibular condyle-derived chondrocytes (MCDC) were cultured under chondrogenesis-supporting conditions; that is, 5 × 10 5 cells/mL were incubated in Dulbecco’s modified Eagle medium supplemented with 100 μg/mL ascorbic acid, 1 mmol/L calcium chloride, 10 mmol/L β-glycerophosphate, 10% fetal calf serum, and antibiotics. Development and growth rates of these cartilage-derived cultures were determined by following morphological and functional changes. MCDC proliferated intensively during the first 24–48 h following plating, showing fibroblast-like (long spindle-shaped) morphology and producing mainly type I collagen. The proliferation rate gradually declined, and the cells developed polygonal shapes and started to produce type II collagen. In the 10–14-day-old cultures, cells began to aggregate in cartilaginous nodules and exhibited positive staining for acidic Alcian blue, type X collagen, and von Kossa. Expression of core-binding factor α 1 increased between 3 and 5 days and declined gradually thereafter. The condylar-derived tissue culture presented here depicts a spontaneous redifferentiation chondrocytic tissue culture that exhibits features of mature chondrocytes typically found in skeletal growth centers. The present study offers a model for primary chondrocytic tissue culture, which might serve as a model for in vitro endochondral ossification.