Microarray analysis of the chondrocytic cell line T/C-28a2 under dynamic fluid shear

Abstract

The behavior of the chondrocytic cell line T/C-28a2 under shear flow was examined using a 32,448 element microarray. A parallel plate flow chamber was used to generate a shear stress level of 20 dyn/cm/sup 2/ for 1.5 or 24 hours (h), after which gene regulation was measured. Microarray analysis revealed differentially regulated genes affecting proliferation/differentiation, extracellular matrix/cytoskeleton, and inflammation at both time points. A ribonuclease protection assay was performed on a subset of genes to confirm the data obtained from the microarray. However, the cyclooxygenase-2 (COX-2) gene, which plays a role in the prostaglandin production in inflamed tissues and the synovium of rheumatoid arthritis (RA) patients, was studied further. Western hybridization revealed that COX-2 protein is present at 24 h, but not at 6 or 12 h. Also, immunofluorescence microscopy shows that COX-2 protein localizes in the cytosol after 24 h of shear, and is not present after 1.5 h. By examining the overall gene expression profiles of chondrocytes under different conditions of dynamic fluid shear, new insights on the pathogenesis of cartilage related diseases such as rheumatoid arthritis might be generated.