Antibodies to Cryptococcus neoformans glucuronoxylomannan enhance antifungal activity of murine macrophages

Abstract

Monoclonal antibodies (MAbs) to the capsular polysaccharide of the pathogenic fungus Cryptococcus neoformans can prolong survival and decrease organ fungal burden in experimental murine cryptococcosis. To investigate the mechanism of antibody-mediated protection, the interaction of C. neoformans and murine macrophage-like J774.16 cells was studied in the presence and absence of MAbs differing in isotype. Immunoglobulin G2a (IgG2a) and IgG2b isotype switch variants were isolated from an IgM hybridoma to complete the IgG subclass set. IgM, IgG1, IgG2a, IgG2b, IgG3, and IgA MAbs were studied for their ability to promote phagocytosis and reduce the number of CFU in C. neoformans and J774.16 cell cocultures. The MAbs in this set had similar if not identical fine specificities and were derived from a single B cell. All isotypes promoted phagocytosis; however, the IgG subclasses were more effective opsonins than IgM or IgA. All isotypes enhanced J774.16 anti-C. neoformans activity in vitro, as measured by a reduction in the number of CFU. The IgG1 MAbs were consistently more active in promoting opsonization and reducing the number of CFU. Addition of IgG1 MAb to C. neoformans and J774.16 cocultures resulted in rapid reduction in the number of CFU, which is consistent with fungal killing. Electron microscopy revealed that MAb-opsonized C. neoformans cells were internalized and appeared damaged. Administration of IgM, IgG1, IgG2a, and IgG2b isotype switch variant MAbs revealed that the IgG2a and IgG2b subclasses were the most and least effective isotypes, respectively, in prolonging survival in an intraperitoneal murine infection model. The results indicate that murine antibody subclasses differ in their ability to enhance macrophage anti-C. neoformans activity and suggest that antibody enhancement of macrophage function is a mechanism by which antibodies modify infection in vivo.