Gene Expression of Joint Cartilage Preserved under Different Conditions and Treated with Growth Factors

Abstract

Purpose To analyze the gene expression of anabolic and catabolic factors in chondrocyte cultures obtained from cartilage preserved at different temperatures, using different preservation media and treated with growth factors. Materials and methods A culture was made of a young lamb's distal femoral cartilage preserved at 4 °C without a cryopreservative agent, at 4 °C with PBS, at 4 °C with DMEM, at -80 °C without a cryopreservative agent, at –80 °C with 10% glycerol and –80 °C with 10% DMSO. An additional culture was made of osteoarthritic and old cartilage. RT-PCR analyses were conducted of the cultured cells with 10 ng of RNA per reaction and 20 nmols of each of the two primers; the cells were also treated with growth factors (TGF-β, FGF-a, IGF-1, OP-1®). RT-PCR reactions were evaluated by means of electrophoresis in agarose gels. Results The best results were obtained for samples cultured at 4 °C with PBS; in old cartilage a reduction was observed in the expression of all anabolic factors, except agrecan. Cartilage stored at -80 °C, with glycerol or DMSO, expressed a higher amount of catabolic factors. On adding growth factors, an increase was registered in the expression of TGF-βwhile MMP-2 levels remained unchanged, at the same level as those of the control group. The best response was obtained with OP-1®, FGF-a and IGF-1. Conclusion Cartilage stored at -80 °C expresses a higher amount of catabolic factors than cartilage preserved at 4 °C.