To determine the effects of recombinant equine IL-1beta and a number of antiinflammatory compounds on the expression and activity of inducible nitric oxide synthase (iNOS) in cultured equine chondrocytes.
RT-PCR methods were used to amplify a portion of the equine iNOS message to prepare an RNA probe. Northern blot analysis was used to quantify the expression of iNOS in first passage cultures of equine articular chondrocytes propagated in the presence or absence of recombinant equine interleukin-1beta (reIL-1beta), dexamethasone (DEX), polysulfated glycosaminoglycan (PSGAG), hyaluronan (HA), and phenylbutazone (PBZ), each at concentrations of 10 and 100 microg/ml. Nitrite concentrations in conditioned media of similarly treated cells were used to quantify iNOS activity.
Recombinant equine IL-1beta increased the expression of iNOS in a dose-dependent manner. This result was paralleled by an increased concentration of nitrite in the culture media of reIL-1beta-treated cells. DEX and PSGAG significantly reduced iNOS gene expression and media supernatant nitrite concentrations in cytokine-stimulated cultures. HA and PBZ had no consistent effect on the expression of iNOS and did not significantly influence nitrite content of conditioned media.
NO is considered an important mediator in the pathophysiologic processes of arthritis and an inducible NOS is expressed by equine chondrocytes. Pre-translational regulation of the iNOS gene by DEX and PSGAG appears to contribute to the cartilage-sparing properties of these compounds.