Candidicidal activity of the human recombinant myeloperoxidase system

Abstract

The effect of the human recombinant myeloperoxidase (recMPO) system on the viability of Candida albicans blastospores was determined. Cells were incubated in the presence of the enzyme, H202, and various substrates. In optimal conditions, 100% mortality was reached with only 41 mU of recMPO per ml, confirming its powerful candidicidal activity.To fight invading microorganisms, polymorphonuclear neutrophils use essentially two mechanisms. The first one, which is nonoxidative, involves such molecules as defensins, permeability-inducing proteins, proteases, or lysozyme (12). The second process occurs via the respiratory burst; this oxygen-dependent system leads to the production of reactive oxygen metabolites, such as H202 (reviewed in reference 5). The toxicity of the generated hydrogen peroxide is increased by reaction with a granular enzyme called myeloperoxidase (MPO) and a halide to form potent halide oxidation products such as hypochlorous acid (HOCl) (9).The candidicidal activity of the natural MPO system has been documented (6, 11); it has been shown also that MPO-deficient polymorphonuclear neutrophils lose their candidicidal property (4) and that in monocytes maturing into macrophages, the decrease in MPO content is accompanied by a decline in candidicidal activity (13). Moreover, a few MPO-deficient individuals have developed serious hematogenous Candida infections (15).In view of the potential interest of MPO as an antifungal agent, we studied the effects of a recombinant form of the enzyme (recMPO) (14) on the viability of Candida albicans blastospores, using as substrates H202 generated by glucose and glucose oxidase and physiological (Cl-) and (I-) halides. In addition, the candidicidal activity of the recMPO system was evaluated at different pHs and in the presence of ammonium ions, which by reaction with HOCl yields strongly oxidizing monochloramines (15).Candidicidal system. Experiments were conducted with a C. albicans strain, 10231, obtained from the American Type Culture Collection. Cells were grown on YM (Difco) agar slopes for 30 h at 30°C and then collected by rinsing with 1 ml of 0.9% NaCl. The suspension was centrifuged and resuspended in 20 mM NaKPO4 buffer, pH 7.5, to a density of 106 cells per ml. For each assay, 1 ml of the suspension was incubated with gentle stirring for 60 min at 37°C, with the various components of the reaction system (i.e., recMPO, glucose, and glucose oxidase) and a halide, in a final volume of 5 ml of NaKPO4 buffer, pH 7.5. Aliquots of the reaction mix were collected, diluted with 0.9% NaCl and spread in triplicate onto Sabouraud dextrose agar plates containing 50 ,ug of chloramphenicol per ml. Colonies were counted after incubation of the plates for 48 h at 30°C.An initial control experiment determined the maximal * Corresponding author.H202 concentration tolerated by C. albicans blastospores; to this end, the concentration of glucose was maintained at 1%while the concentration of glucose oxidase was varied. As shown in Fig. 1, catalase-positive C. albicans sup...

Vaginal myeloperoxidase and flora in the pig‐tailed macaque

“…However, exogenous MPO appeared to transiently increase vaginal MPO levels, suggesting that, at least for a brief period of time, this method of augmentation of endogenous vaginal defenses might be effective. If MPO works in synergy with vaginal H 2 O 2 -producing organisms, then a similar protective effect against STD, as has been seen in 6itro, may occur in 6i6o [2,5,7,13,15]. This effect remains to be adequately tested in this or another suitable model.…”

“…Following sample collection at time 0, the five animals randomly selected for MPO treatment received a vaginal infusion of 1.5 ml of sterile lubricant (KY gel, nonbacteriostatic; Johnson & Johnson Corp., new Brunswick, New Jersey) mixed with 250 mg of recombinant MPO. Recombinant MPO, which has been shown to be microbicidal [2,13], was generously provided by N. Moguilevsky and A. Bollen of the Service de Génétique Appliquée, Université Libre de Bruxelles, Belgium. Sample collection was repeated at 4 and 24 hours.…”