Histochemical identification of sulfation position in chondroitin sulfate in various cartilages

Abstract

The ability of chondrocytes to synthesize chondroitin-4-sulfate (C4S) as opposed to chondroitin-6-sulfate (C6S) is a phylogenetically related phenomenon seen among adult higher vertebrates and developmentally during the embryogenesis of these vertebrates. While the embryonic cartilage may be initially a C6S matrix, C4S synthesis is seen to develop with time. We have histochemically localized these differences in sulfation with the cationic carbocyanine dye, Stains-all, in a spectrum of cartilages that vary in the sulfation position of their chondroitin sulfate. Cartilages from the rat and rabbit that are predominantly C4S stained magenta at pH 4.3, while the C6S-rich cartilage matrices from the regenerating rabbit ear and lamprey cranium stained blue. Embryonic chicken cartilages develop a gradient of magenta matrix with age, with increased concentration toward the articular surface. Both magenta and blue matrices were absent after pretreatment with chondroitinase ABC but were present after Streptomyces hyaluronidase digestion. The magenta staining was a property of the cartilage matrix as a whole, since isolated C4S and C6S stained blue. The differential staining was seen at pH 4.3, but not at pH 8.8, suggesting an interaction between the chondroitin sulfate and the adjacent tissue proteins.