ISOLATED CHONDRONS - A VIABLE ALTERNATIVE FOR STUDIES OF CHONDROCYTE METABOLISM IN-VITRO

Objective: To develop and test a simple enzymatic procedure for isolat ing chondrons, which consist of the chondrocytes and their surrounding pericellular microenvironment. Design: Chondrons were obtained by dig esting adult human articular cartilage with a mixture of dispase and c ollagenase. Chondrons and chondrocytes were cultured in alginate beads , immunofluorescence labeled and examined by confocal microscopy. Resu lts: Comparison of freshly isolated chondrons with cryostat sections o f cartilage revealed that type VI collagen, type ii collagen and aggre can were retained, but fibronectin and a unique chondroitin sulfate ep itope recognized by the antibody, 7D4, were lost. Comparison of enzyma tic and mechanical homogenization methods revealed subtle changes in c hondron morphology and retention of fibronectin in mechanically isolat ed chondrons. Average yield of enzyme-isolated chondrons was slightly lower than that of chondrocytes isolated by pronase and collagenase di gestion, but was much greater than that reported for mechanically isol ated chondrons. Enzyme-isolated chondron viability was greater than 80 % 1 day after isolation, and continued to be above 80% through 7 weeks of alginate bead culture. Viability of isolated chondrocytes was init ially greater than 80% but fell to 60-80% with time in culture. Chondr ons and isolated chondrocytes had a similar division rate except osteo arthritic chondrons were significantly slower after 2 weeks in culture . Cell division was more rapid for nonosteoarthritic chondrons than fo r osteoarthritic ones. Conclusions: Enzymatic isolation of chondrons i s relatively simple, gives better yield and viability than mechanical isolation, but comparable yield and viability of traditional chondrocy te isolation. Enzymatic chondron isolation allows the effect of the in vivo-formed pericellular matrix on chondrocyte metabolism to be studi ed in vitro.