Immature murine articular chondrocytes in primary culture: a new tool for investigating cartilage

Abstract

Many genetically modified animal models are providing new keys for unlocking the pathophysiology of cartilage degradation. To produce a tool for cellular and molecular studies in genetically engineered murine models, we defined the optimal culture conditions for primary cultures of articular chondrocytes from newborn mice (C57Bl/6). To determine whether the cultured cells exhibited the typical articular chondrocyte phenotype, we examined several morphological, biochemical, and functional features. The cells had the typical chondrocyte morphology, with a rounded or polygonal shape. Immunolocalization studies showed high levels of type II collagen and aggrecan expression, together with sulfated glycosaminoglycan accumulation. Type II collagen and aggrecan expression decreased with passaging. In contrast, type I collagen expression was low in primary cultures and high after four passages, indicating a fibroblast phenotype. To evaluate the functional integrity of our cultured cells, we evaluated their ability to produce prostaglandin E2 (PGE2) and nitric oxide (NO) in response to the catabolic cytokine interleukin (IL)-1beta (10 ng/ml). Production of both PGE2 and NO increased significantly as compared to untreated controls. In addition, IL-1beta induced COX-2 expression by the cultured cells, as shown by Western blotting. Since functional and molecular parameters can be measured readily in mice, the immature murine articular chondrocyte (iMAC) model described here should prove a powerful tool for research, particularly as many transgenic and knockout mouse strains are available, even if iMACs are not optimal substitutes for human chondrocytes.