IN-SITU DETECTION OF SUPEROXIDE ANIONS WITHIN PORCINE ARTICULAR-CARTILAGE

Cartilage was isolated from pig articular joints, and the production o f reactive oxygen species (ROS) by chondrocytes embedded within the ca rtilage was assessed by two methods: the reduction of nitro blue tetra zolium and by the use of diaminobenzidine in the presence of manganese ions. Little constitutive generation of ROS was seen, but it could be detected after the addition of the calcium ionophore ionomycin. Furth er, the response seen was extremely heterogeneous; same cells showed a far greater release of ROS than others. Cells arranged in the columna r arrays of the deep zone were the most active, while those furthest f rom the cartilage/bone interface (i.e. nearer to the outer face of the cartilage) were unresponsive. Chondrocytes cultured in alginate beads also showed a similar heterogeneity in their response, suggesting tha t the isolation of these cells and the measurement of ROS production i n a population is not representative of the true situation.