PROTEINS IN THE CELL-WALL AND MEMBRANE OF CRYPTOCOCCUS-NEOFORMANS STIMULATE LYMPHOCYTES FROM BOTH ADULTS AND FETAL CORD-BLOOD TO PROLIFERATE

Cryptococcus neoformans is an encapsulated yeast that infects patients who have defective cell-mediated immunity, including AIDS, but rarely infects individuals who have intact cell-mediated immunity. Studies o f the immune response to C. neoformans have been hampered by a paucity of defined T-lymphocyte antigens, and hence, the understanding of the T-cell response is incomplete. The goal of this study was to separate C. neoformans into its component parts, determine whether those compo nents stimulate lymphocyte proliferation, perform preliminary characte rization of the proteins, and establish the potential mechanism of lym phocyte proliferation, The lymphocyte response to fungal culture mediu m, whole organisms, disrupted organisms, and the yeast intracellular f raction or cell wall and membrane was studied by determining thymidine incorporation and by determining the number of lymphocytes at various times after stimulation, The cell wall and membrane of C. neoformans stimulated lymphocyte proliferation, while the intracellular fraction and culture filtrate did not, The optimal response occurred on day 7 o f incubation, with 4 x 10(5) peripheral blood mononuclear cells per we ll and with 13 mu g of cryptococcal protein per ml. The number of lymp hocytes increased with time in culture, indicating that thymidine inco rporation was accompanied by proliferation, Proteinase K treatment of the cell wall and membrane abrogated lymphocyte proliferation, indicat ing that the molecule was a protein. [S-35]methionine labeling, sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and fluorography were performed to analyze the proteins contained in the cell wall and membrane, intracellular fraction, and culture filtrate, At least 18 di screte bands were resolved from the cell wall and membrane. Since a la rge percentage of healthy adults responded to the cryptococcal cell wa ll and membrane, a mitogenic effect was investigated by testing prolif eration of fetal cord blood lymphocytes, The percentage of fetal sampl es that proliferated in response to the cell wall and membrane was sim ilar to the percentage of fetal samples that proliferated in response to Staphylococcus enterotoxin B, a microbial mitogen. Thus, a protein in the cell wall and membrane of C, neoformans is a potent stimulant o f lymphocyte proliferation and has mitogenic properties, which may hav e important implications for cell-mediated immunity to C. neoformans.