Effect of in vitro aging on the biosynthesis of glycosaminoglycans by human skin fibroblasts. Modulation by the elastin-laminin receptor

The incorporation of a radioactive precursor H-3-glucosamine in glycoconjugates, essentially glycosaminoglycans (GAG) was evaluated in the culture medium and cell fraction of human skin fibroblasts. Using increasing passage numbers, we could estimate the effect of in vitro aging on these biosynthetic activities. The incorporation in different free (hyaluronan) and protein bound (proteoglycans) GAGs was evaluated after specific enzymatic digestion. Most newly synthesized GAGs were excreted in the extracellular medium. Incorporation of the tracer in hyaluronan, the major biosynthetic product, increased with passage number but its titratable concentration decreased with in vitro aging, suggesting a rapid post-synthetic degradation. The proportion of chondroitin sulfates 4 (A) and 6 (C) and heparan sulfate decreased and that of dermatan sulfate increased with increasing passage number. We explored the modulation of these biosynthetic activities by the elastin laminin receptor. Using agonists (elastin peptides) and an antagonist (melibiose) of the receptor, their action on GAG biosynthesis was evaluated. Both elastin peptides and melibiose increased incorporation of the tracer in GAGs, but only melibiose inhibited post-synthetic degradation of hyaluronan, therefore increasing its concentration. The effect of passage number on the receptor mediated modulations was also investigated. (C) 1999 Elsevier Science Ireland Ltd. All rights reserved.