To analyze the gene expression of osteoarthritic chondrocytes cultured in alginate after stimulation with interleukin (IL)-1beta.
Chondrocytes were isolated from osteoarthritic cartilage obtained during total knee replacement by sequential enzymatic digestion. After suspension in alginate, cells were cultured with and without 100 pg/ml IL-1beta. Quantitative RT-PCR reaction was used to estimate the messenger RNA (mRNA) of three different metabolites [tumor-necrosis-factor-stimulated gene 6 (TSG-6), stromelysin-1 (MMP-3) and aggrecan (AGG)].
After having shown the precision of quantitative PCR, this method allowed us to detect IL-1beta-induced changes in mRNA of TSG-6, MMP-3 and AGG. MMP-3 was found to be the most abundant transcript, IL-1beta induced a 12-fold upregulation of MMP-3 levels compared to control, and 7-fold of TSG-6. The AGG transcript level, indicating anabolic events, was found to be downregulated by between 2- and 3-fold.
In our culture system, the response of osteoarthritic chondrocytes to IL-1beta is preserved. Therefore, this system might be helpful for further investigation of the influences of drugs, cytokines and growth factors, for example, on the metabolism of chondrocytes at the level of gene transcription as the most basic level of regulation.