HOST-PATHOGEN INTERACTION DURING CRYPTOCOCCUS NEOFORMANS CNS INFECTION

Abstract

Cryptococcus neoformans (C. neoformans) is an opportunistic fungal pathogen widely distributed in the environment globally. C. neoformans infection initiates from the lung through inhaling the spores. While most healthy individuals can clear the fungus or contain the fungus in the granuloma, immunosuppressed patients and a small group of healthy populations fail in controlling the cryptococcal fungal pulmonary infection. In those cases, C. neoformans transmigrates from the lung to the central nervous system (CNS) and causes fatal meningoencephalitis, which accounts for 112,000 deaths each year worldwide. However, we have a very limited understanding of the transmigration of C. neoformans from the bloodstream to the brain in vivo, and the mechanism involved in the clearance of the organism in the brain remains poorly understood.

In this study, we first report a novel approach to quantitatively analyze the interactions between C. neoformans and brain endothelial cells in a mouse model using flow cytometry. Using this system, we show that C. neoformans was internalized by brain endothelial cells in vivo and that mice infected with acapsular or heat-killed C. neoformans yeast cells displayed a lower frequency of brain endothelial cells containing the yeast cell compared to mice infected with wild-type or viable yeast cells, respectively. We further demonstrate that brain endothelial cells were invaded by the serotype A strain (H99 strain) at a higher rate compared to the serotype D strain (52D strain).

Moreover, we found that clearance of C. neoformans in the brain correlates with accumulation and pro-inflammatory M1 polarization of Ly6Chi mononuclear phagocytes and that these phagocytes play a critical role in the clearance of C. neoformans in the brain. Notably, the accumulation of Ly6Chi mononuclear phagocytes coincides with enhanced secretions of TNF and IFN-γ in the brain. TNF receptor (TNFR) signaling, but not IFN-γ receptor (IFN-γR) signaling, mediates the recruitment of Ly6Chi mononuclear phagocytes to the brain in a cell-intrinsic manner. By contrast, IFN-γ induces M1 polarization of Ly6Chi mononuclear phagocytes. Disruption of TNFR or IFN-γR signaling enhances cryptococcal growth in the brain. Thus, Ly6Chi mononuclear phagocytes act as effector cells for cryptococcal clearance in the brain, involving TNFR as well as IFN-γR signaling.

Collectively, our study established that 1) internalization of C. neoformans by brain endothelial cells occurred in vivo and offered a powerful approach to quantitatively analyze fungal migration into the brain; 2) Ly6Chi mononuclear phagocytes accumulate in the brain during brain infection with C. neoformans and function as effector cells for clearance of C. neoformans in the brain involving TNFR signaling and IFN-γ signaling.