Glycoproteins and glycosaminoglycans synthesized by human keratinocytes in culture. Their role in cell-substratum adhesion

Abstract

Glycoproteins and proteoglycans synthesized by human keratinocytes in medium containing D-[1-14C]glucosamine were extracted and analysed by polyacrylamide-gel electrophoresis in the presence of sodium dodecyl sulphate. Extraction of the labelled keratinocytes with 0.5% Triton X-100 removed most of the glycoconjugates and left the cytoskeleton and nuclear residue adherent to the substratum. In addition to the cytoskeletal proteins, there was a relatively simple profile of glycoproteins and glycosaminoglycans associated with this adherent cytoskeleton. These consisted of eight glycoproteins in the mol.wt. range 99000-232000, five proteins in the keratin region (mol.wt. 42000-61000), hyaluronic acid and a sulphated glycosaminoglycan. Surface labelling of the keratinocytes with galactose oxidase (with or without neuraminidase)/KB3H4 revealed that many of the glycoproteins were exposed on the cell surface. The importance of the glycoproteins and proteoglycans in attaching the keratinocytes to the substratum was examined by studying their expression after incubation in medium containing tunicamycin and their degradation after digestion with trypsin and hyaluronidase. These studies, together with an examination of the glycoconjugates released by sequential extraction with 0.5% Triton X-100 followed by 0.2% sodium dodecyl sulphate, revealed that the glycoprotein of mol.wt. 232000 has an important role in mediating the attachment of keratinocytes to the substratum.

Stimulating effects of Bacillus subtilis natto-fermented Radix astragali on hyaluronic acid production in human skin cells

“…HA plays important biological roles in the skin, such as holding moisture, maintaining the architecture and elasticity of tissues, and facilitating the exchange of nutrients and waste products (Laurent and Fraser, 1992;Manuskiatti and Maibach, 1996). HA is synthesized mainly by fibroblasts and keratinocytes, and distributed in the connective tissue of the dermis and the intercellular space of the epidermis (Roberts and Jenner, 1983;Tammi et al, 1988;Sakai et al, 1999). Age-dependent decrease of hyaluronic acid has been identified as one of the major factor responsible for the physical appearance of aged skin, including wrinkle formation and the loss in elasticity of the skin (Longas et al, 1987;Ghersetich et al, 1994).…”

Epithelial origin of cutaneous anchoring fibrils.

“…The basal lamina is a continuous layer of extracellular matrix molecules including type IV collagen, laminin, and entactin. Most basal lamina components have been shown to be epithelial cell products (Briggaman, 1981;Hay, 1981Hay, , 1982Kefalides et al, 1979;Kubo et al, 1984;Martin and Timpl, 1987;Roberts and Jenner, 1983) and are involved in the regulation of epithelial growth and differentiation (Foellmer et al, 1983). The basal keratinocytes adhere to their basal lamina via fine filaments known as anchoring filaments or microfilaments which extend from the hemidesmosomes through the lamina lucida into the lamina densa (see Fig.…”

Biosynthesis of proteoglycans by proliferating and differentiating normal human keratinocytes cultured in serum‐free medium

“…However, when epidermal cells were cultured in the absence of dermis, the synthesis of hyaluronic acid was considerably reduced, whereas the synthesis of sulfated GAGs was unaffected. Roberts and Jenner (1983) have shown that human keratinocytes when cultured in the presence of 3T3 feeder cells synthesize hyaluronic acid and sulfated GAGs. Similarly, Brown and Parkinson (1983) have demonstrated that human keratinocytes in culture synthesize hyaluronic acid, heparan sulfate, and chondroitin sulfate.…”