Neutrophil Candidacidal Activity Research Articles

Perinuclear Anti-Neutrophil Cytoplasmic Antibodies (pANCA) Impair Neutrophil Candidacidal Activity and Are Increased in the Cellular Fraction of Vaginal Samples from Women with Vulvovaginal Candidiasis.

Vulvovaginal candidiasis (VVC) is primarily caused by Candida albicans and affects 75% of childbearing age women. Although C. albicans can colonize asymptomatically, disease is associated with an increased Candida burden, a loss of epithelial tolerance and a breakdown in vaginal microbiota homeostasis. VVC symptoms have been ascribed to a powerful inflammatory response associated with the infiltration of non-protective neutrophils (PMN). Here, we compared the immunological characteristics of vaginal fluids and cellular protein extracts obtained from 28 VVC women and from 23 healthy women colonized by Candida spp. We measured the levels of antibodies against fungal antigens and human autoantigens (anti-Saccharomyces cerevisiae antibodies (ASCA), C. albicans germ tube antibodies (CAGTAs) and perinuclear anti-neutrophil cytoplasmic antibodies (pANCA)), in addition to other immunological markers. Our results show that the pANCA levels detected in the cellular protein extracts from the vaginal fluids of symptomatic women were significantly higher than those obtained from healthy colonized women. Consistent with a potential physiologically relevant role for this pANCA, we found that specific anti-myeloperoxidase antibodies could completely neutralize the ex vivo killing capacity of polymorphonuclear cells. Collectively, this preliminary study suggests for the first time that pANCA are found in the pathogenic vaginal environment and can promptly impair neutrophil function against Candida, potentially preventing a protective response.

EphA2 Is a Neutrophil Receptor for Candida albicans that Stimulates Antifungal Activity during Oropharyngeal Infection

SUMMARYDuring oropharyngeal candidiasis (OPC), Candida albicans proliferates and invades the superficial oral epithelium. Ephrin type-A receptor 2 (EphA2) functions as an oral epithelial cell β-glucan receptor that triggers the production of proinflammatory mediators in response to fungal infection. Because EphA2 is also expressed by neutrophils, we investigated its role in neutrophil candidacidal activity during OPC. We found that EphA2 on stromal cells is required for the accumulation of phagocytes in the oral mucosa of mice with OPC. EphA2 on neutrophils is also central to host defense against OPC. The interaction of neutrophil EphA2 with serum- opsonized C. albicans yeast activates the MEK-ERK signaling pathway, leading to NADPH subunit p47phox site-specific phospho-priming. This priming increases intracellular reactive oxygen species production and enhances fungal killing. Thus, in neutrophils, EphA2 serves as a receptor for β-glucans that augments Fcγ receptor-mediated antifungal activity and controls early fungal proliferation during OPC.

β-glucan priming enhances neutrophil Fcγ receptor-mediated antifungal activity during oral mucosal infection

Abstract Neutrophils are key to control fungal proliferation in the oral cavity. Recently we identified that the ephrin type-A receptor 2 (EphA2) functions as an oral epithelial cell β-glucan receptor that triggers epithelial cell production of proinflammatory mediators in response to Candida albicans infection. Because EphA2 is also expressed by neutrophils, we investigated the role of EphA2 in neutrophil candidacidal activity during OPC. We found that when OPC was induced in EphA2−/− mice, there was delayed accumulation of phagocytes into the oral tissues and more severe disease relative to wild-type mice. Depletion of neutrophils in EphA2−/− mice did not further increase oral fungal burden, nor did adoptive transfer of EphA2−/− neutrophils into CD18−/− mice decrease oral fungal burden, indicating that EphA2 on neutrophils plays a central role in the host defense against OPC. Mice lacking the Fcγ receptor I (CD64) were also highly susceptible to OPC. In vitro studies showed that EphA2 signaling augments neutrophil Fcγ receptor-mediated antifungal activity. The interaction of serum-opsonized C. albicansyeast with neutrophil EphA2 activated the MEK-ERK signaling pathway, leading to NADPH subunit p47phox site specific phospho-priming. EphA2-induced priming of neutrophil p47phox increased intracellular ROS production and enhanced killing of opsonized C. albicans yeast. Thus, in neutrophils, EphA2 serves as a receptor for β-glucans that enhances Fcγ receptor-mediated antifungal activity and is crucial to control early fungal proliferation during OPC.

EphA2 is a Neutrophil Receptor for Candida Albicans that Stimulates Antifungal Activity During Oropharyngeal Infection

During oropharyngeal candidiasis (OPC), Candida albicans proliferates and invades the superficial oral epithelium. The ephrin type-A receptor 2 (EphA2) functions as an oral epithelial cell β-glucan receptor that triggers the production of proinflammatory mediators in response to fungal infection. Because EphA2 is also expressed by neutrophils, we investigated the role of EphA2 in neutrophil candidacidal activity during OPC. We found that when OPC was induced in EphA2–/– mice, there was delayed accumulation of phagocytes in the oral tissues and more severe disease relative to wild-type mice. Depletion of neutrophils in EphA2–/– mice did not further increase oral fungal burden, nor did adoptive transfer of EphA2–/– neutrophils into CD18−/− mice decrease oral fungal burden, indicating that EphA2 on neutrophils plays a central role in the host defense against OPC. Mice lacking the Fcγ receptor I were also highly susceptible to OPC. In vitro studies showed that EphA2 signaling augments neutrophil Fcγ receptor-mediated antifungal activity. The interaction of serum-opsonized C. albicans yeast with neutrophil EphA2 activated the MEK-ERK signaling pathway, leading to NADPH subunit p47phox site specific phospho-priming. EphA2-induced priming of neutrophil p47phox increased intracellular ROS production and enhanced killing of opsonized C. albicans yeast. Thus, in neutrophils, EphA2 serves as a receptor for β-glucans that enhances Fcγ receptor-mediated antifungal activity and is crucial to control early fungal proliferation during OPC.

Leucocyte Phagocytosis In Children With Urinary Schistosomiasis And Asymptomatic Malaria Parasitemia

In the participants considered for this study, leucocyte migration, neutrophil candidacidal activity and ability to generate reactive oxygen were determined as percentage migration index (%M. I), candidacidal phagocytic index (%C.I) and bacterial stimulated nitroblue tetrazolium (NBT) dye reduction index (%NBT) respectively. Also, malaria density was counted from thick blood film of glass slide stained with Giemsa stain. The participants were 54 school children having urinary schistosomiasis without malaria parasites (USS-M), 18 children with both urinary schistosomiasis and malaria parasites (USS+M), 46 children with malaria parasites without urinary schistosomiasis (M-USS) and 29 controls. The mean % M.I was least while %NBT index was highest in USS+M subjects but M-USS subjects had least %C.I. Malaria density was higher in M-USS subjects than USS+M subjects. The results of this study showed that low prevalence and reduced severity of malaria parasites in children with urinary schistosomiasis may be due to adequate production of leucocyte migration inhibitory factor (LMIF) and reactive oxygen species. Key words: Leucocyte phagocytosis, malaria, schistosomiasis, Nigeria Afr. J. Clin. Exper. Microbiol. 2005; 6(2): 81-86

A monoclonal antibody to Candida albicans enhances mouse neutrophil candidacidal activity.

A monoclonal antibody (MAb) to Candida albicans (MAb B6.1) that protects against candidiasis and the nonprotective MAb B6 were compared for ability to support neutrophil (polymorphonuclear leukocyte [PMN]) candidacidal activity. Both MAbs are immunoglobulin M, and each recognizes distinct C. albicans mannan cell wall determinants. PMN candidacidal activity was assessed by transmission electron microscopy and by an in vitro killing assay. The results indicated that MAb B6.1, but not MAb B6, enhances ingestion and killing of yeast cells by PMN in the presence of serum complement.

Effect of fluconazole on the interactions between human neutrophils and Candida albicans

The effects of fluconazole on the candidacidal activity of human neutrophils were investigated. Three clinical isolates of Candida albicans were studied in a colony count assay and the presence of fluconazole during incubation of yeasts with neutrophils had no effect on neutrophil candidacidal activity. These results, using a different assay and three additional isolates, confirm previous studies showing that fluconazole does not interfere with the ability of neutrophils to kill C. albicans.

Effect of Friend leukemia virus infection on susceptibility to Candida albicans

Previous studies have demonstrated that Friend leukemia virus (FLV) induces a profound immunosuppression in susceptible mice. The studies described in this report indicate that mice infected with FLV have an increased susceptibility to subsequent infection with the opportunistic pathogen Candida albicans, as measured by increased numbers of C. albicans CFU in the kidneys of FLV-infected mice relative to uninfected controls. Experiments in which the NB-tropic and N-tropic strains of FLV were used suggest that virus replication or the resulting virus burden may be important in the observed increased susceptibility to C. albicans. Since neutrophils are believed to be important in the response of mice to systemic Candida infections, the effect of FLV infection on neutrophil candidacidal activity was investigated. The percentage of neutrophils present in unfractionated Proteose Peptone-elicited peritoneal exudates of mice infected with FLV for 14 days was significantly lower than in uninfected control mice or mice infected with FLV for 6 or 10 days. When neutrophils from FLV-infected and control mice were purified, adjusted to equal concentrations, and tested for in vitro candidacidal activity, neutrophils from mice infected with FLV for 14 days were deficient in their ability to kill C. albicans relative to normal controls and mice infected with FLV for 6 or 10 days. Addition of normal mouse serum increased killing in all groups but did not restore candidacidal activity of neutrophils from mice infected with FLV for 14 days to levels of control neutrophils or neutrophils from mice infected for 6 or 10 days with the virus. These results suggest a defect in neutrophil function, at the later stages of FLV infection, involving in vitro candidacidal activity. In addition, neutrophils from FLV-infected mice may be deficient in in vivo chemotactic activity. These defects in neutrophil function could account, at least in part, for the observed increased susceptibility of FLV-infected mice to C. albicans.

Effect of cilofungin (LY121019), a fungal cell wall synthesis inhibitor, on interactions of Candida albicans with human neutrophils.

The effects of cilofungin, a lipopeptide antifungal agent active against several species of Candida, on human neutrophil candidacidal activity were investigated. While no increase or decrease in neutrophil killing of blastospores was observed when cilofungin was simultaneously incubated with neutrophils and yeasts, preincubation of yeasts with serum containing cilofungin resulted in approximately 90% decrease in killing of five of six strains. Attachment/phagocytosis of these strains was unchanged from controls. We conclude that cilofungin interferes with opsonization of most strains of C. albicans, resulting in a marked decrease in neutrophil candidacidal activity, a phenomenon which may be of significance in vivo in certain tissues where access to antibodies and/or complement is limited.

A Micromethod for Measuring Neutrophil Candidacidal Activity in Neonates

We developed a micromethod, requiring but a few drops of blood, for measuring the ability of neutrophils to kill ingested Candida albicans. The neutrophils of 10 normal adults killed 26.2 +/- 9.5% of ingested C. albicans in 2.5 hr in a standard fungicidal assay, and 21.4 +/- 6.5% in our new micromodification. Neutrophils from 14 full term, healthy 30- to 40-hr-old infants, studied with the micromethod, killed 23.2 +/- 6.0% of ingested C. albicans. We conclude that the neutrophils of normal neonates are competent in this sphere of antimicrobial activity.

Cyclic 3',5'-adenosine monophosphate in the human lukocyte: synthesis, degradation, andeffects n neutrophil candidacidal activity.

Prostaglandins E(1) and E(2) (PGE(1) and PGE(2)) stimulate adenyl cyclase activity in broken cell preparations of normal human leukocytes, whereas prostaglandin F(1a) produces no effect. PGE(1) and PGE(2) also cause increased accumulation of cyclic 3',5'-adenosine monophosphate-(3)H ((3)H-labeled AMP) in intact leukocytes which have been preincubated with adenine-(3)H in vitro. Theophylline inhibits leukocyte phosphodiesterase activity and potentiates the stimulatory effect of the prostaglandins on intracellular accumulation of cyclic 3',5'-AMP-(3)H. The ability of human granulocytes in vitro to kill Candida albicans was consistently inhibited by PGE(1) and theophylline. This effect was reproduced by dibutyryl cyclic 3',5'-AMP, a lipid-soluble analogue of the endogenous nucleotide. The inhibition of candidacidal activity could not be accounted for by drug effects on phagocytosis, oxygen consumption, or hexose monophosphate shunt activity. These results are consistent with the hypothesis that increased intracellular concentrations of cyclic 3',5'-AMP impair the granulocyte's ability to kill C. albicans, but the precise mechanism of inhibition has not yet been defined.

Interaction of Candida albicans with human leukocytes and serum.

A quantitative assay of candidacidal activity based on differential staining of non-viable Candida albicans by methylene blue was developed and applied to studies of leukocytes from normal individuals and patients with fungal and other infections. Serum factors were necessary for optimal phagocytosis of C. albicans but lacked direct candidacidal activity. Normal human neutrophils (38 studies) killed 29.0 +/- 7.4% of ingested C. albicans in 1 hr. Eosinophils and monocytes killed a smaller percentage. Neutrophil candidacidal activity did not require protein or ribonucleic acid synthesis by the leukocyte but was inhibited by anaerobic conditions, potassium cyanide, and colchicine. Leukocytes of a patient with hereditary myeloperoxidase deficiency and of three children with chronic granulomatous disease phagocytized C. albicans normally, yet failed to kill them. Our data suggest that the neutrophil can play an important role in resistance to Candida infection and that the lysosomal enzyme myeloperoxidase and its oxidant substrate hydrogen peroxide are the major participants in neutrophil candidacidal activity.