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Comparative studies were performed on the uptake of 35SO4-labelled proteoglycans and of free 35SO4-glycosaminoglycan chains by cultured skin fibroblasts. Proteoglycans as well as protein-free polysaccharide chains are internalized by adsorptive pinocytosis. The maximum capacity for endocytosis of proteoglycans is about four times higher than for glycosaminoglycan chains when referred to disaccharide units. On a molar basis about 10 times more glycosaminoglycan chains than proteoglycans can be internalized within a given time. Since unlabelled carbohydrate chains do not compete for the uptake of labelled proteoglycans it seems likely that different receptors exist on the cell surface for endocytosis of both types of macromolecules. 14C-labelled hyaluronate was neither pinocytosed to a measurable extent by skin fibroblasts nor by intima cells cultured from the thoracic aorta. Rat liver hepatocytes efficiently internalized hyaluronate by adsorptive pinocytosis. It was concluded from competitive experiments with hyaluronate oligosaccharides that hyaluronate has to interact with at least two receptor molecules on the cell membrane to achieve endocytosis.

Cultured cells derived from bovine corneal stroma synthesize all types of sulfated glycosaminoglycans and distribute these macromolecules into topographically different compartments in a reproducible manner. Each compartment is characterized by a typical glycosaminoglycan distribution pattern. Corneal fibroblasts synthesize in vitro only small amounts of keratan sulfate in contrast to the in vivo conditions. We have, therefore, investigated the synthesis and topographical distribution of sulfated glycosaminoglycans as influenced by different culture conditions. The following results were obtained: 1) Cocultivation of epithelial and stromal fibroblasts from bovine cornea led to an increased incorporation of radiosulfate into sulfated glycosaminoglycans by about 50% as compared to the theoretical value. Glycosaminoglycan distribution of mixed cultures into different compartments showed no similarity compared with pure epithelial or stromal fibroblasts. 2) Addition of native or heat inactivated anterior chamber fluid to the culture medium was followed by a twofold increase of [35S]-sulfate incorporation and by an augmented intracellular and pericellular accumulation of labeled macromolecules. 3) Reduction of the incubation temperature led to a reduced synthesis of glycosaminoglycans without influencing their topographical distribution. Growth of stromal cells on type I collagen was accompanied by a reduced glycosaminoglycan synthesis of about 25%. Extracellular macromolecules reached only half of the normal value, while intracellularly their concentration was slightly increased. 4) None of the variations of the culture condition led to a significant change of the distribution pattern of sulfated glycosaminoglycans. Especially, no significant increase of keratan sulfate biosynthesis could be detected.

Lysosomal hydrolases participate substantially in the degradation of all classes of biological macromolecules. They act physiologically within the lysosome. The enzymes are either primarily included within primary lysosomes or are transported to these cell organelles after secretion and subsequent adsorptive pinocytosis. The involvement of these enzymes in a variety of pathological conditions can be understood on the basis of the known functions of lysosomal hydrolases. Inactivity of one or several of the enzymes causes lysosomal storage disorders. Similar metabolic consequences are found when the enzymes are unable to be concentrated within the lysosome. Lysosomal hydrolases participate, furthermore, in the pathogenesis of numerous diseases. A distinction can be made between lysosomal overload, pathologically-increased enzyme secretion into the extracellular space, and a release of lysosomal enzymes into the cytosol.