Organ-Specific Innate Immune Responses in a Mouse Model of Invasive Candidiasis

Molecular Signaling Section, Laboratory of Molecular Immunology, National Institute of Allergy and Infectious Diseases, Bethesda, MD 20892, USA.
Journal of Innate Immunity (Impact Factor: 4.35). 11/2010; 3(2):180-99. DOI: 10.1159/000321157
Source: PubMed

ABSTRACT In a fatal mouse model of invasive candidiasis (IC), fungal burden changes with variable dynamics in the kidney, brain, spleen, and liver and declines in all organs except for the kidney, which inexorably loses function. Since leukocytes are required to control Candida, we hypothesized that differential leukocyte infiltration determines organ-specific outcome of the infection. We defined leukocyte accumulation in the blood, kidney, brain, spleen, and liver after infection using fluorescent-activated cell sorting (FACS) and immunohistochemistry. Accumulation of Ly6c(int)CD11b(+) neutrophils predominated in all organs except the brain, where CD45(int)CD11b(+)CD11c(-) microglia were the major leukocytes detected, surrounding foci of invading Candida. Significantly more neutrophils accumulated in the spleen and liver than in the kidney during the first 24 h after infection, when neutrophil presence is critical for Candida control. Conversely, at later time points only the kidney continued to accumulate neutrophils, associated with immunopathology and organ failure. The distribution of neutrophils was completely different in each organ, with large abscesses exclusively forming in the kidney. Candida filamentation, an essential virulence factor, was seen in the kidney but not in the spleen or liver. IC induced Ly6c(hi)CD11b(+) inflammatory monocyte and NK1.1(+) cell expansion in the blood and all organs tested, and MHCII(+)F4/80(+)CD11c(-) macrophage accumulation, mainly in the spleen and liver. This study is the first detailed analysis of leukocyte subsets accumulating in different target organs during IC. The results delineate immune responses to the same pathogen that are highly idiosyncratic for each organ tested. The work provides novel insights into the balance between effective host defense and immunopathology in IC.

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Available from: Chyi-Chia Richard Lee, Sep 26, 2015
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    • "21. Take a small aliquot before the last wash to determine neutrophil count, viability by trypan blue exclusion and FACS analysis using a fluorescent viability dye (Lionakis et al., 2011), and purity by flow cytometry using staining with APC-conjugated anti-CD45, PE-conjugated anti-Ly6G and APC-Cy7–conjugated anti-CD11b. "
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    ABSTRACT: Neutrophils represent the first line of defense against bacterial and fungal pathogens. Indeed, patients with inherited and acquired qualitative and quantitative neutrophil defects are at high risk for developing bacterial and fungal infections and suffering adverse outcomes from these infections. Therefore, research aiming at defining the molecular factors that modulate neutrophil effector function under homeostatic conditions and during infection is essential for devising strategies to augment neutrophil function and improve the outcome of infected individuals. This unit describes a reproducible density gradient centrifugation-based protocol that can be applied in any laboratory to harvest large numbers of highly enriched and highly viable neutrophils from the bone marrow of mice both at the steady state and following infection with Candida albicans as described in UNIT. In another protocol, we also present a method that combines gentle enzymatic tissue digestion with a positive immunomagnetic selection technique or Fluorescence-activated cell sorting (FACS) to harvest highly pure and highly viable preparations of neutrophils directly from mouse tissues such as the kidney, the liver or the spleen. Finally, methods for isolating neutrophils from mouse peritoneal fluid and peripheral blood are included. Mouse neutrophils isolated by these protocols can be used for examining several aspects of cellular function ex vivo including pathogen binding, phagocytosis and killing, neutrophil chemotaxis, oxidative burst, degranulation and cytokine production, and for performing neutrophil adoptive transfer experiments. © 2015 by John Wiley & Sons, Inc.
    Current protocols in immunology / edited by John E. Coligan ... [et al.] 08/2015; 110:3.20.1-3.20.15. DOI:10.1002/0471142735.im0320s110
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    • "Additionally, as yeast cells are less immuno-stimulatory, these cells may disseminate within the bloodstream without causing an extensive host immune activation, compared to hyphae. However, in the final stages of systemic infection in mouse models, development of hyphae is favored (Lionakis et al., 2011). "
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    ABSTRACT: A range of attributes determines the virulence of human pathogens. During interactions with their hosts, pathogenic microbes often undergo transitions between distinct stages, and the ability to switch between these can be directly related to the disease process. Understanding the mechanisms and dynamics of these transitions is a key factor in understanding and combating infectious diseases. The human fungal pathogen Candida albicans exhibits different morphotypes at different stages during the course of infection (candidiasis). For example, hyphae are considered to be the invasive form, which causes tissue damage, while yeast cells are predominant in the commensal stage. Here, we described interactions of C. albicans with its human host in a game theoretic model. In the game, players are fungal cells. Each fungal cell can adopt one of the two strategies: to exist as a yeast or hyphal cell. We characterized the ranges of model parameters in which the coexistence of both yeast and hyphal forms is plausible. Stability analysis of the system showed that, in theory, a reduced ability of the host to specifically recognize yeast and hyphal cells can result in bi-stability of the microbial populations' profile. Inspired by the model analysis we reasoned that the types of microbial interactions can change during invasive candidiasis. We found that positive cooperation among fungal cells occurs in mild infections and an enhanced tendency to invade the host is associated with negative cooperation. The model can easily be extended to multi-player systems with direct application to identifying individuals that enhance either positive or negative cooperation. Results of the modeling approach have potential application in developing treatment strategies.
    Frontiers in Microbiology 02/2014; 5:41. DOI:10.3389/fmicb.2014.00041 · 3.99 Impact Factor
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    • "The mice were weighed every day. All of the mice were sacrificed after 5 days, and the kidneys were harvested (as most of the Candida cells in systemic candidiasis murine models are found in the kidneys [39]). The kidneys were divided into equal portions for fungal burden determination and histopathological evaluation. "
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    ABSTRACT: Candida is the most common fungal pathogen of humans worldwide and has become a major clinical problem because of the growing number of immunocompromised patients, who are susceptible to infection. Moreover, the number of available antifungals is limited, and antifungal-resistant Candida strains are emerging. New and effective antifungals are therefore urgently needed. Here, we discovered a small molecule with activity against Candida spp. both in vitro and in vivo. We screened a library of 50,240 small molecules for inhibitors of yeast-to-hypha transition, a major virulence attribute of Candida albicans. This screening identified 20 active compounds. Further examination of the in vitro antifungal and anti-biofilm properties of these compounds, using a range of Candida spp., led to the discovery of SM21, a highly potent antifungal molecule (minimum inhibitory concentration (MIC) 0.2 - 1.6 µg/ml). In vitro, SM21 was toxic to fungi but not to various human cell lines or bacterial species and was active against Candida isolates that are resistant to existing antifungal agents. Moreover, SM21 was relatively more effective against biofilms of Candida spp. than the current antifungal agents. In vivo, SM21 prevented the death of mice in a systemic candidiasis model and was also more effective than the common antifungal nystatin at reducing the extent of tongue lesions in a mouse model of oral candidiasis. Propidium iodide uptake assay showed that SM21 affected the integrity of the cell membrane. Taken together, our results indicate that SM21 has the potential to be developed as a novel antifungal agent for clinical use.
    PLoS ONE 01/2014; 9(1):e85836. DOI:10.1371/journal.pone.0085836 · 3.23 Impact Factor
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