Identification of constituents of human neutrophil azurophil granules thatmediate fungistasis against Histoplasma capsulatum

Previously we demonstrated that human neutrophils mediate potent and long-l asting fungistasis against Histoplasma capsulatum yeasts and that all of th e fungistatic activity resides in the azurophil granules. In the present st udy, specific azurophil granule constituents with fungistatic activity were identified by incubation with H. capsulatum yeasts for 24 h and by quantif ying the subsequent growth of yeasts via the incorporation of [H-3]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 inhi bition at 8 mu g/ml. At a concentration of 4 mu g/ml, all possible paired c ombinations of defensins exhibited additive fungistatic activity against H. capsulatum yeasts. Cathepsin G and bactericidal-permeability-increasing pr otein (BPI) also mediated fungistasis against H. capsulatum in a concentrat ion-dependent manner. The fungistatic activities of combinations of catheps in 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 a ntifungal activity, and azurocidin and proteinase 3 exhibited no significan t fungistasis against H. capsulatum yeasts. Thus, defensins, cathepsin G, a nd BPI are the major anti-H. capsulatum effector molecules in the azurophil granules of human neutrophils.