Cellular Immunity in a Cutaneous Model of Cryptococcosis

Abstract

The temporal development of cellular immune responses in mice inoculated cutaneously with viable Cryptococcus neoformans 145 was determined in vivo and in vitro by comparing several antigen preparations for their efficacy in the assays selected. Three antigens derived from C. neoformans 145, viz., a culture filtrate preparation (CneF-145), a membrane extract (B-HEX), and soluble cytoplasmic substances (SCS), were compared for their ability to detect delayed hypersensitivity (DH) in vivo in a footpad assay or to stimulate lymphocytes in vitro in a thymidine incorporation assay. DH to B-HEX could be demonstrated as early as 1 week after infection, whereas significant responses to SCS and CneF-145 were not regularly detected until 3 weeks after infection. Substantial reactions were observed to all three antigens up to 12 weeks, although they peaked at 2 to 3 weeks. Reactions to B-HEX and SCS were somewhat better than those to CneF. Differences in the efficacies of the three antigens were not obvious after the sixth week of infection, however. In vitro, lymph node cells from infected animals were stimulated significantly with all three antigens beginning at week 1. As with DH, however, responses to CneF-145 were usually less than those to SCS and B-HEX. In vitro lymphocyte responses waned after approximately 6 weeks, whereas DH responses were clearly positive through 12 weeks. In addition to the studies in infected animals, animals immunized with heat-killed cells of C. neoformans 145 or 184 were tested 6 to 8 days later for DH with CneF-145, CneF-184, or B-HEX derived from C. neoformans 145. The CneF-145 and CneF-184 were equally effective for detecting DH, regardless of the cryptococcal strain used for immunization. Likewise, the B-HEX detected equivalent responses in mice sensitized with each cryptococcal strain. Since all three antigens were soluble and easily extracted and since each elicited significant cellular immune responses in infected animals, further studies involving their specificity and the nature of their reactive components seems warranted as they may help evaluate immune responses in humans infected with this fungus.