The proteins synthesized and released by human astrocytoma cells cultured with radiolabeled amino acids were analyzed by polyacrylamide gel electrophoresis in sodium dodecyl sulfate and by trichloracetic acid precipitation. A select group of extracellular proteins was released from the astrocytomas. The most abundant extracellular proteins were at least 250,000 daltons. Five other major proteins (P 175, P 125, P 60, P 47, and P 40) were 175,000, 125,000, 60,000, 47,000, and 40,000 daltons, respectively. The complement of proteins retained by the cells was considerably more complex than those released. Clinical efforts to enhance the immunologic response directed against human astrocytomas must distinguish between extracellular and retained antigens. The pattern of proteins released by gliomas might be diagnostically useful if present in cerebrospinal fluid or serum.

With [3H]fucose as a marker, C6 glioma cells in culture released an 85,000 molecular weight molecule into the medium as the major extracellular glycoprotein. The quantity and extracellular/cytoplasmic ratio of this glycoprotein suggest that its cellular processing is different from that of five other released glycoproteins of molecular weights 55,000, 115,000, 130,000, 150,000, and 170,000. Nearly 40% of newly synthesized glycoproteins in the cells was released into the culture medium. Major glycoproteins retained by the cells migrated electrophoretically to molecular weight positions of 82,000, 110,000, 120,000, 140,000, and 160,000, and approximately one-third of these returned glycoproteins were labile to trypsinization. Both synthesis and release of these macromolecules were inhibited more than 95% with cycloheximide treatment, demonstrating that nearly all fucosylation was linked to protein synthesis. Since 40% of all glycoproteins was released under conditions of more than 99% cellular viability, it is likely that these extracellular glycoproteins are physiological products of membrane turnover and secretion, but not of cell lysis. The results provide a basis for the further study of glial differentiation and of shed glioma antigens.