Cell surface glycosaminoglycans: Identification and organization in cultured human embryo fibroblasts

A morphologically detectable cell coat, composed of glycoprotein, glycolipid, and glycosaminoglycan, is present on the external surface of most vertebrate cells. We have invetigated the composition and organization of glycosaminoglycans in the cell coat of cultured human embryo fibroblasts by labeling cells with ³H-glucosamine and Na₂³⁵SO₄ and subsequently treating cultures with specific enzymes. Components released were identified by chromatography and specific enzymatic digestion. In situ incubation with leech hyaluronidase (4 μg/ml) removed only hyaluronic acid from the cell surface whereas testicular hyaluronidase (0.5 mg/ml) removed both hyaluronic acid and chondroitin sulfate. Trypsin (0.1 mg/ml) released a large mass of glycopeptides in addition to hyaluronic acid, chondroitin sulfate, and heparan sulfate. The affinity of the cell coat for the cationic dye, ruthenium red, was reduced by leech hyaluronidase treatment. Sequential enzyme digestions of the cell surface showed that hyaluronic acid could be removed without the concomitant or subsequent release of sulfated glycosaminoglycans, suggesting that the hyaluronic acid is not a structural backbone for glycosaminoglycan complexes of the external cell surface.