Identification of Constituents of Human Neutrophil Azurophil Granules That Mediate Fungistasis againstHistoplasma capsulatum

Abstract

Previously we demonstrated that human neutrophils mediate potent and long-lasting fungistasis against Histoplasma capsulatum yeasts and that all of the fungistatic activity resides in the azurophil granules. In the present study, specific azurophil granule constituents with fungistatic activity were identified by incubation with H. capsulatum yeasts for 24 h and by quantifying the subsequent growth of yeasts via the incorporation of [ 3 H]leucine. Human neutrophil defensins HNP-1, HNP-2, and HNP-3 inhibited the growth of H. capsulatum yeasts in a concentration-dependent manner with maximum inhibition at 8 g/ml. At a concentration of 4 g/ml, all possible paired combinations of defensins exhibited additive fungistatic activity against H. capsulatum yeasts. Cathepsin G and bactericidal-permeability-increasing protein (BPI) also mediated fungistasis against H. capsulatum in a concentration-dependent manner. The fungistatic activities of combinations of cathepsin G and BPI were additive, as were those of combinations of cathepsin G or BPI with HNP-1, HNP-2, and HNP-3. Lysozyme and elastase exhibited modest antifungal activity, and azurocidin and proteinase 3 exhibited no significant fungistasis against H. capsulatum yeasts. Thus, defensins, cathepsin G, and BPI are the major anti-H. capsulatum effector molecules in the azurophil granules of human neutrophils.Histoplasma capsulatum is a dimorphic fungal pathogen of worldwide importance that causes a broad spectrum of disease activity. The course of H. capsulatum infection is mild in most immunocompetent individuals, but progressive disseminated infections occur in individuals immunocompromised by hematologic malignancies (6,25,30) or cytotoxic therapy (16,32,33) or in individuals infected with human immunodeficiency virus (15,20,31).Infection with H. capsulatum is acquired by inhalation of microconidia into the terminal bronchioles and alveoli of the lung. Inhaled microconidia subsequently convert into yeasts that are responsible for the pathogenesis of histoplasmosis (17). H. capsulatum yeasts are phagocytized by alveolar macrophages (M), within which they multiply (2, 7). Dividing yeasts destroy the alveolar M, and subsequently the yeasts are ingested by other resident alveolar M and by inflammatory phagocytes recruited to the locus of infection. Repetition of this cycle results in spread of infection to hilar lymph nodes and to other organs during the acute phase of primary histoplasmosis. Subsequently, the maturation of specific cell-mediated immunity against H. capsulatum activates M to halt yeast proliferation with gradual resolution of the disease process in immunocompetent hosts (7,22).The role of polymorphonuclear neutrophils (PMNs) in the cell-mediated immune response against H. capsulatum is unclear (7). However, even the earliest studies in a murine model of histoplasmosis described PMNs as being the predominant inflammatory cell type in the lungs during the first 36 h after intranasal inoculation with H. capsulatum macroconidia. In these studies the PMNs ...

Pulmonary immune responses induced in BALB/c mice by Paracoccidioides brasiliensis conidia

“…Furthermore, we observed an increase of lysozyme expression in the lungs of mice infected with P. brasiliensis conidia during the early stages of infection (1-4 days), with the expression concentrating mainly on PMNs and macrophages with a parallel decrease in the number of fungal propagules (Gonzalez et al, unpublished data). The microbicidal effect of lysozyme has been demonstrated against C. albicans and H. capsulatum [106,125,126]. These results suggest that in vivo, lysozyme could well be one of the molecules participating in the control of P. brasiliensis infection.…”

“…Thus, the phagocytic system constitutes an important effector mechanism of the natural and adaptative immune response leading to inhibition or destruction of several pathogens through the production of molecules that exert such effect, once they become activated by cytokines. Among these effector molecules, one can list degradative enzymes such as collagenases and elastases, lysozyme, defensins proteinases, reactive oxygen intermediates (ROI) and reactive nitrogen intermediates (RNI) [106][107][108].…”

Macrophage-independent Fungicidal Action of the Pulmonary Collectins

“…Rapid access to this intracellular niche also allows Hc to escape the immune defenses of the alveolar lining fluid, which include the antimicrobial proteins lactoferrin, lysozyme, and defensins (2).…”

Lung Diseases - Selected State of the Art Reviews

“…During any infectious process, phagocytic cells are the main effector cells, which upon their activation (e.g., by cytokines), produce microbicidal substances [i.e., degradative enzymes, defensins, lysozymes, reactive oxygen intermediates (ROS) and reactive nitrogen intermediates (RNI)] capable of inhibiting and destroying several pathogens (Nathan & Shiloh, 2000;Newman et al, 2000). Taking into account that both PMN and Ms are the principal components of the pulmonary inflammatory infiltrate of mice infected with P. brasiliensis conidia, the expression of microbicidal molecules produced by these phagocytic cells in the PCM model was determined.…”