Hyaluronan distribution in the human and canine intervertebral disc and cartilage endplate

A biotinylated complex of aggrecan G1-domain and link protein was used to characterize the distribution of hyaluronan in paraffin-embedded sections of adult human and canine intervertebral disc and cartilage endplate. Limited chondroitinase ABC and trypsin digestions of the sections before staining was utilized to expose hyaluronan potentially masked by aggrecan. Hyaluronan concentration and hyaluronan to uronic acid ratio in different parts of the discs were measured as a background for the histological analysis. Hyaluronan staining was strong in the nucleus pulposus and inner parts of annulus fibrosus of both species, corroborated by biochemical assays of the same compartments. Particularly in human samples, hyaluronan in the interterritorial matrix of nucleus pulposus and annulus fibrosus was readily accessible to the probe without enzyme treatments. In contrast, the cell-associated hyaluronan signal was enhanced after trypsin or limited chondroitinase ABC-treatment of the sections, suggesting that pericellular hyaluronan was more masked by aggrecan than in the distant matrix. A puzzling feature of canine cartilage endplate cells was their intensive cell-associated hyaluronan signal, part of which appeared intracellular. Hyaluronan was abundant between the collagenous lamellae in annulus fibrosus, perhaps important in the plasticity of this tissue.