Capsular specific IgM enhances complement-mediated phagocytosis and killing of Cryptococcus neoformans by methamphetamine-treated J774.16 macrophage-like cells

Abstract

Methamphetamine (METH) is a powerful and highly addictive stimulant that affects the central nervous system of users in the United States and worldwide, and its consumption is associated to the acquisition of HIV and AIDS-related infections. METH enhances cryptococcosis in mice, an opportunistic infection caused by the encapsulated fungus Cryptococcus neoformans. Due to its ability to survive within macrophages, C. neoformans is an ideal model to study pathogen-macrophage interactions. METH abrogates normal macrophage function, which might contribute to the higher rate and more rapid progression of infections in drug abusers. Hence, we investigated the role of complement and specific IgM to C. neoformans capsular polysaccharide on the function of J774.16 macrophage-like cells after exposure to METH. We found that complement and IgM significantly promotes complement-mediated phagocytosis of C. neoformans by J774.16 cells in comparison to co-incubation with complement alone. IgM enhances the expression of complement receptor 3 on the surface macrophages treated with the drug. Also, IgM-increased macrophage phagocytosis of C. neoformans may be associated with upregulation of GTPase-RhoA, a key regulator of the actin polymerization signaling cascade. Fungal cells incubated with complement and IgM in the presence of METH demonstrated higher number of cells per aggregate, a possible explanation for their enhanced ingestion by phagocytes. IgM increased killing of yeast cells by macrophages by inhibiting the alkalization of the phagosome and stimulating the intracellular production of nitric oxide. Together, our findings suggest that IgM stimulates the effector functions of macrophages against opportunistic pathogens in the setting of drug abuse.

Methamphetamine Impairs IgG1-Mediated Phagocytosis and Killing of Cryptococcus neoformans by J774.16 Macrophage- and NR-9640 Microglia-Like Cells

“…Recent evidence supporting this possibility showed that METH mediates structural changes to the capsular composition (11) and inhibits complement-mediated phagocytosis (13) of C. neoformans. Interestingly, specific IgM to GXM enhances complement-mediated phagocytosis of the fungus (13), suggesting that the pentameric structure of this isotype may prevent METH-induced capsular modifications and positively influence the interaction of C. neoformans with complement receptors in macrophages.…”

“…Phagocytosis assay. Macrophage-or microglia-like cells were incubated in 96-well microtiter plates (Corning) in the absence or presence of METH (25 or 50 M; Sigma) for 2 h. Cytochalasin D (25 M; Sigma) was used as a positive control because it potently inhibits actin polymerization and phagocytosis (13,15). Monolayers of J774.16 or NR-9640 cells were washed thrice with PBS, and feeding medium (13) supplemented with gamma interferon (IFN-␥; 100 U/ml) and bacterial lipopolysaccharide (LPS; 1 g/ml) was added, followed by the addition of preincubated cryptococci with MAb 18B7 (anti-cryptococcal GXM IgG1 generated and generously provided by Arturo Casadevall; 2 g/ml) for 1 h, in a macrophage/ microglia (10 5 cells):yeast/latex bead (Thermo Fisher) (10 6 cells) ratio of 1:10.…”

“…In this regard, METH enhances C. neoformans infection of the respiratory system and dissemination to the CNS of rodents by promoting fungal attachment, alteration of the polysaccharide capsule composition, release of immunosuppressive capsular material, and biofilm formation (11,12). Thus, C. neoformans is an excellent model organism to answer questions regarding hostpathogen interactions in the setting of METH due to the accessibility to specific antibodies (Abs), cell lines, and animal models (13).…”

“…Cryptococci easily replicate and release abundant amounts of polysaccharide-enclosed vesicles inside phagocytic cells that accumulate in their phagosome, resulting in the escape of yeast cells through lytic and nonlytic exocytosis (23)(24)(25). Even though METH compromises the ability of macrophages to maintain acidic phagolysosomes (13,16), the impact of this drug of abuse on the intracellular effects of specific Abs on the fate of a microbe within murine macrophages has not been extensively investigated. The intimate interaction of C. neoformans with macrophages is an ideal system to examine the role of METH in Ab function (13).…”

“…Even though METH compromises the ability of macrophages to maintain acidic phagolysosomes (13,16), the impact of this drug of abuse on the intracellular effects of specific Abs on the fate of a microbe within murine macrophages has not been extensively investigated. The intimate interaction of C. neoformans with macrophages is an ideal system to examine the role of METH in Ab function (13). Similarly and particularly important to cryptococcal infection, positron emission tomography has demonstrated that the highest accumulation and slowest clearance of METH in humans occur in the lungs and brain, respectively, with these organs being main disease-related targets of the fungus (26).…”