Observations on the morphology of the proteinpolysaccharide complex of bovine nasal cartilage and its relationship to collagen.

Abstract

The investigation has been carried out by electron microscope examination of intact cartilage and of various fractions of an aqueous extract of that tissue. The materials were stained, either with an 0·5 % solution of bismuth nitrate in 0·1 m nitric acid in water at a pH of 1·2, or with a similar solution of bismuth nitrate in 0·1 m nitric acid in 90 % acetone. It is considered that in both circumstances the bismuth was bound exclusively to the sulphate group of chondroitin sulphate and keratosulphate. The protein and chondroitin sulphate moieties of the proteinpolysaccharide complex were isolated after alkaline treatment. The former contains 15 % keratosulphate and it is only this part which stains. Both the protein and the chondroitin sulphate were visualized as discrete groups of particles in which the average particle diameter was 23 Å. It is suggested that each group represents a network of keratosulphate or chondroitin sulphate A ( CSA ) chains respectively, cross-linked by bismuth ions. And that each individual particle represents a segment of a carbohydrate chain which has assumed a coiled configuration owing to the neutralization of its net charge by bismuth. The light fraction of the aqueous extract of cartilage contains the proteinpolysaccharide complex alone. After precipitation with aqueous bismuth nitrate its appearance was essentially similar to that of its CSA and protein fractions, except that the average particle size was 47 Å. After precipitation by bismuth nitrate in acetone, on the other hand, it was seen as single rows of particles, the length of the rows varying from 1100 Å to 1500 Å, and the particles having an average diameter of 30 Å. Each row is interpreted as a proteinpolysaccharide macro-molecule, and it is considered that each particle represents an individual carbohydrate chain, whereas the intervals between the particles represent the unstained protein moiety. The indi­viduality of the macromolecules in these circumstances is ascribed to the low dielectric constant of acetone. The relationship of the proteinpolysaccharide complex to collagen was studied in the heavy fraction of the aqueous extract of cartilage and in intact cartilage. It was observed that the heavy fraction consists of proteinpolysaccharide and collagen and that a pro­portion of the complex is bound to collagen. In each collagen period, proteinpolysaccharide macromolecules were attached transversely round the circumference of the fibre over the a and b 1 bands. A similar relationship was noted in intact cartilage.