Primary culture and phenotyping of murine chondrocytes.
The culture of chondrocytes is one of the most powerful tools for exploring the intracellular and molecular features of chondrocyte differentiation and activation. However, chondrocytes tend to dedifferentiate into fibroblasts when they are subcultured, which is a major problem. This protocol, involving primary cultures to limit dedifferentiation, describes two different methods for culturing chondrocytes of different anatomical origins (articular and costal chondrocytes, both of which represent hyaline cartilage) from mice. Mice are of particular interest for cellular and molecular studies, as many tools suitable for use in mice are available. In addition, rapid development of transgenic and gene-targeted mice provides powerful instruments for biological studies. The protocol can be divided into four stages: isolation of cartilage (15 min per animal), isolation of chondrocytes (2 h extended overnight), seeding of chondrocytes (1 h 30 min) and growth in culture (6 d). To obtain confluency of chondrocytes using this protocol takes 7 d. Methods for phenotyping chondrocytes are also provided.
Gosset
Marjolaine
M
Paris Universitas-Université Pierre et Marie Curie Paris VI, Department of Physiology and Physiopathology, Centre National de la Recherche Scientifique (Unité Mixte de Recherche 7079), 7 quai St-Bernard, Paris 75252 Cedex 5, France.
Berenbaum
Francis
F
Thirion
Sylvie
S
Jacques
Claire
C
D000818:Animals; D002356:Cartilage / Q000166:cytology; D018929:Cell Culture Techniques; D002454:Cell Differentiation; D019902:Chondrocytes / Q000166:cytology*; D003470:Culture Media; D051379:Mice; D010641:Phenotype
D003470:Culture Media