Synthesis of cellular and extracellular glycoproteins by cultured human keratinocytes and their response to retinoids

Abstract

The glycoproteins synthesized by human keratinocytes cultured on 3T3 feeder layers were studied by metabolic labelling. Keratinocytes freed of feeder cells synthesized a complex pattern of cellular and extracellular glycoproteins that was distinct from that of 3T3 cells, dermal fibroblasts and epidermal melanocytes. The effect of low concentrations of all- trans-retinoic acid and arotinoid ethyl ester on glycoprotein synthesis was examined in keratinocyte cultures depleted of vitamin A. Treatment with either retinoid resulted in a 2–3-fold increase in the amount of d-[ 3H]glucosamine-labelled material in the culture medium. Gel electrophoresis revealed increased incorporation of d-[ 3H]glucosamine into extracellular glycoproteins of M r 245 000, 170 000, 140 000, 130 000, 120 000 and 105 000 as well as into glycosaminoglycans in retinoid-treated cultures. The labelling of extracellular glycoproteins with l-[ 3H]leucine and l-[ 35S]methionine was also increased by retinoids suggesting increased synthesis of these components rather than an effect on their glycosylation. The M r 245 000 glycoprotein was identified as keratinocyte-derived fibronectin by immunoblotting, immunoprecipitation and specific binding to gelatin. The results show that retinoids increase the synthesis of glycoprotein as well as glycosaminoglycan components of the extracellular matrix in human keratinocyte cultures. It is suggested that retinoids select for a population of cells that synthesize relatively large amounts of glycosaminoglycan, fibronectin and other as yet unidentified extracellular glycoproteins.