Evaluation of chondrocyte micromass culture for the study of cartilage degradation
© BioMed Central Ltd 2005
Received: 11 January 2005
Published: 17 February 2005
An understanding of the mechanisms involved in cartilage degradation that occur in rheumatoid arthritis and osteoarthritis is essential for the development of treatments to block disease progression. Culturing chondrocytes to synthesize a cartilage matrix would ideally provide a cartilage matrix of consistent composition that would reproducibly respond to different stimuli. When cultured in monolayers, chondrocytes convert to a fibroblastic morphology and gradually lose their native phenotype, no longer able to synthesize type II collagen and aggrecan. However, when chondrocytes are cultured at a high density in the presence of growth factors, the chondrocytes regain their native phenotype and native round morphology .
To optimise the culture conditions for chondrocyte micromass synthesis of matrix components, and to evaluate the response of the culture to cytokine stimulation by measuring the degradation of matrix components.
Chondrocytes extracted from bovine nasal cartilage were passaged several times to produce substantial cell stocks. Harvested cells were resuspended at 2 × 107 cells/ml and seeded in the centre of each well of a 48-well plate (20 μl/well). Cultures were maintained in a serum-free medium with or without ITS (insulin, transferrin and selenium), ascorbic acid, and transforming growth factor beta 1 (TGFβ1) for 14–21 days. Histological analysis of the micromass cultures examined the presence of proteoglycans and type II collagen. The effects of cytokine stimulation with IL-1, oncostatin M (OSM), and the combination of both cytokines on matrix degradation were tested by analysing proteoglycan and collagen release. Collagenase activity was measured by bioassay.
The chondrocyte micromass cultures synthesized a matrix that contained proteoglycan and type II collagen. The micromass culture may serve as a good model for studying proteoglycan but not collagen degradation.
This work was funded by GlaxoSmithKline.