755 GLYCOPROTEINS IN CULTURE MEDIUM: A COMPARISON FROM CYSTIC FIBROSIS (CF) AND CONTROL SKIN FIBROBLASTS

Abstract

Glycoproteins of high molecular weight are found associated with the cell surface or in growth media of fibroblasts and other cell types in culture. Several abnormalities of glycoproteins in CF have been reported. Therefore the glycoproteins from the media of CF and matched control skin fibroblasts were isolated and partially characterized. Cells were labeled with L-[3H]fucose and grown in medium containing 10% serum. For some experiments the cells were subsequently grown in serum free medium for 48 hours prior to harvest. Media were examined from cells in both log and confluent phases of the growth cycle. The four methods used to isolate the glycoproteins were cryoprecipitation, ammonium sulfate precipitation, heparin precipitation and a heparin-sepharose column. The precipitated proteins were characterized by polyacrylamide gel electrophoresis, isoelectric focusing and protein and carbohydrate content. Different classes of glycoproteins were obtained with each of the four methods. Only heparin precipitation yielded a single radioactive glycoprotein (Mr>200,000) demonstrated by gel electrophoresis. Several qualitative and quantitative differences were noted. In all cases less protein per cell was precipitated by heparin from the CF medium when compared to control. USPHS grant AM16859 and CF Foundation Student Traineeships.