Influence of staphylococcal metabolites on adhesive reactions in the system "Candida albicans-buccal epitheliocytes"

The influence of S. aureus and S. epidermitidis metabolites on the adhesive reactions in the system "C. albicans-buccal epitheliocytes" was studied. The study revealed that the treatment of C. albicans with S. aureus supernatants inhibited the adhesion of C. albicans to epitheliocytes, the degree of the inhibiting action of S. aureus supernatants in the system depending on their strain specificity. S. epidermitidis supernatants produced no adhesive effect. The irreversible decrease of the adhesive activity of C. albicans under the action of bacterial metabolites was, seemingly, the consequence of transformation of the receptor apparatus of C. albicans. At the same time S. aureus supernatants produced no essential influence on the adhesive potential and viability of buccal epitheliocytes.     

Adhesiveness of yeast-like fungi in the system "Candida albicans--vaginal epitheliocytes"

The influence of vaginal liquid on adhesive reactions in the system "Candida albicans - vaginal epitheliocytes". Preliminary treatment of C. albicans by mucin, natural or absorbed vaginal liquid resulted in decrease of adhesive activity (p<0.05). C. albicans did not change its adhesive properties in case of preliminary treatment of vaginal epitheliocytes by the same substances. In the system "Candida albicans - vaginal epitheliocytes" vaginal liquid has the expressed antiadhesive effect, which probably belongs to mucin.     

Antifungal activity of the amyrin derivatives and in vitro inhibition of Candida albicans adhesion to human epithelial cells

AIMS: The antifungal activity of amyrin pentacyclic triterpene and 15 synthetic derivatives was evaluated against Candida species. Additionally, inhibition of adhesion of Candida albicans to human epithelial cells in vitro was determined. METHODS AND RESULTS: Esterification of alpha- and beta-amyrin with a variety of acyl chlorides produced a series of analogue derivatives. These substances were synthesized to evaluate the antifungal properties against Candida species. Among the 15 derivatives, alpha- and beta-amyrin formiate (2) and alpha- and beta-amyrin acetate (3) were the most active, inhibiting all the Candida species tested in concentrations that ranged from 30 to 250 microg ml(-1). alpha- and beta-amyrin formiate inhibited the adhesion ability of C. albicans to buccal epithelial cells (BEC) in 65.3%. CONCLUSIONS: alpha- and beta-amyrin formiate and alpha- and beta-amyrin acetate derivatives exhibited potential antifungal activity against Candida spp. and amyrin formiate showed inhibition of the adhesion ability of C. albicans to buccal epithelial cells. SIGNIFICANCE AND IMPACT OF THE STUDY: This study demonstrated that two derivatives of amyrin pentacyclic triterpene exhibited significant antifungal activity against Candida species. Additionally, alpha- and beta-amyrin formiate was as effective as fluconazole in inhibiting the adhesion of C. albicans to buccal epithelial cells.     

Antiadhesive effect of the secretion of the oral cavity in the system "Candida albicans--buccal epitheliocytes"

The influence of the oral cavity secretion on adhesive reactions in the system "C. albicans--buccal epitheliocytes" was studied. The treatment of C. albicans with natural saliva led to decrease of adhesive activity. The treatment of C. albicans with antibody absorbed saliva at different temperature conditions led to different changes of adhesion. This effect was determined by the action of temperature-dependent and temperature-independent factors, supposedly of enzymatic nature.     

The chemiluminescence reactions of the blood neutrophils and of peritoneal exudate cells in Syrian hamsters to the intraperitoneal administration of cellular and microbial materials]

The luminol-dependent chemiluminescence (CL) activity of peritoneal exudate cells and blood neutrophils of Syrian hamsters inoculated intraperitoneally with heat-inactivated microbial particles of Candida albicans, (C. albicans), heated irradiated normal cells and native or heated irradiated malignant tumor cells was studied. The inoculation with particles of C. albicans and heated normal cells induced significant activation of CL of peritoneal exudate cells, but did not influence the CL reaction of blood neutrophils. The inoculation of animals with nonheated irradiated tumor cells led to increase of CL response of both peritoneal exudate cells and blood neutrophils. The inoculation with heated irradiated tumor cells did not activate CL of peritoneal exudate cells and led to slight, but long-lasting decrease of CL response of blood neutrophils.     

Induction of ERK-kinase signalling triggers morphotype-specific killing of Candida albicans filaments by human neutrophils

Candida albicans is among the most important fungal pathogens in humans. Morphological plasticity has been linked to its pathogenic potential as filamentous forms are associated with tissue invasion and infection. Here we show that human neutrophils discriminate between yeasts and filaments of C. albicans . Whereas filaments induced targeted motility, resulting in the establishment of close contact between neutrophils and fungal cells, yeast forms were largely ignored during coincubation. In transwell assays, C. albicans filaments induced significantly higher migratory activity in neutrophils than yeasts. Neutrophil motility based on actin rearrangement was essential for killing of C. albicans filaments but not involved in killing of yeast forms. Using inhibitors for MAP-kinase cascades, it was shown that recognition of C. albicans filaments by neutrophils is mediated via the MEK/ERK cascade and independent of JNK or p38 activation. Inhibition of the ERK signalling pathway abolished neutrophil migration induced by C. albicans filaments and selectively impaired the ability to kill this morphotype. These data show that invasive filamentous forms of C. albicans trigger a morphotype-specific activation of neutrophils, which is strongly dependent on neutrophil motility. Therefore, human neutrophils are capable of sensing C. albicans invasion and initiating an appropriate early immune response.     

A comparison of phospholipase activity, cellular adherence and pathogenicity of yeasts

Phospholipase A and lysophospholipase activities were measured in the culture fluid and in the blastospores of Candida albicans. When phospholipase activity was measured in six yeasts (four strains of C. albicans and a single strain each of Candida parapsilosis and Saccharomyces cerevisiae) a correlation was found between this activity and two potential parameters of pathogenicity. The C. albicans isolates which adhered most strongly to buccal epithelial cells and were most pathogenic in mice had the highest phospholipase activities. Non-pathogenic yeasts, including C. albicans isolates which did not adhere and did not kill mice, had lower phospholipase activities.     

Platelet-activating factor-induced early activation of NF-kappa B plays a crucial role for organ clearance of Candida albicans

In this study, we have investigated the mechanisms underlying organ susceptibility to candida infection. Infection of BALB/c mice with Candida albicans led to both an early (1-8 h) and late (24-48 h) activation of NF-kappaB in the organs resistant to C. albicans, including the lung and spleen. In susceptible organs such as the kidneys, early activation of NF-kappaB was not observed. The kinetics of TNF-alpha mRNA expression paralleled those of NF-kappaB activation in all organs examined. Blocking the effects of endogenous platelet-activating factor (PAF) by pretreatment with the PAF antagonist BN50739 or antioxidants significantly reduced the early activity of NF-kappaB and TNF-alpha mRNA expression, and increased the recovery of C. albicans in the lung and spleen. Importantly, administration of PAF 5 min prior to the infection resulted in the appearance of early activities of NF-kappaB and TNF-alpha mRNA expression, followed by a nearly complete clearance of the organisms in the kidneys. Pretreatment with anti-TNF-alpha Ab resulted in an enhanced susceptibility to C. albicans, and the PAF-mediated resistance was abrogated by anti-TNF-alpha in all organs examined. These data indicated that endogenously produced PAF in response to C. albicans is a key molecule involved in the early activation of NF-kappaB, which, in turn, renders the organ resistant to the fungus by promoting the production of anti-candidal proinflammatory cytokines such as TNF-alpha. Susceptible organs, including the kidneys, lack the capacity to generate a sufficient PAF-induced early NF-kappaB response.     

Acute endotoxemia prolongs the survival of rat lung neutrophils in response to 12-O-tetradecanoyl-phorbol 13-acetate

Acute endotoxemia is associated with prolonged survival of adherent neutrophils in the lung vasculature. In the present studies, the effects of inflammatory mediators on signaling pathways regulating neutrophil survival were examined. We found that the protein kinase C activator, 12-O-tetradecanoyl-phorbol 13-acetate (TPA), but not interferon-gamma (IFN-gamma), prolonged the survival of adherent vasculature lung neutrophils from endotoxemic rats, a response that was correlated with reduced apoptosis. Although endotoxin administration to rats induced the expression of the anti-apoptotic protein Mcl-1 in lung neutrophils, TPA had no effect on this response. Endotoxin administration also induced expression of total p38 and p44/42 mitogen activated protein kinases (MAPK) in neutrophils, as well as phosphatidyl inositol 3 kinase (PI3K) and its downstream target protein kinase B (PKB). Treatment of the cells with TPA increased p38 MAPK expression in cells from both control and endotoxin treated animals. Cells from endotoxin treated, but not control animals, were found to exhibit constitutive binding activity of nuclear factor kappa B (NF-kappaB) which was blocked by TPA. In contrast, constitutive CCAAT/enhancer binding protein (C/EBP) nuclear binding activity evident in neutrophils from control animals was reduced following endotoxin administration. Moreover, this response was independent of TPA. These data suggest that NF-kappaB plays a role in TPA-induced signaling leading to prolonged survival of adherent vascular neutrophils in the lung during acute endotoxemia.     

Reduced adherence and inhibition of hyphal development of Candida albicans by the antimicrobial agent polynoxylin

Polynoxylin (Anaflex R) was investigated for antimicrobial activities ancillary to its known cidal and static effects. Significant reductions in adherence of Candida albicans blastospores (oral strain) to human buccal epithelial cells were observed following polynoxylin treatment. The anti-adherence activity was concentration and time-dependent. Treatment of the epithelial cells did not result in significant reductions in adherence. Polynoxylin was also shown to inhibit the germination and hyphal development of C. albicans.     

Recognition of Candida albicans Als3 by the germ tube-specific monoclonal antibody 3D9.3

Monoclonal antibody 3D9.3 (MAb 3D9.3) reacts with the surface of Candida albicans germ tubes and recognizes a protein epitope. We used a two-step chromatography procedure to purify and identify the antigen (3D9) from C. albicans strain 66396 germ tubes. MAb 3D9.3 recognized two intense protein bands at 140 and 180 kDa. A comparative analysis between theoretical and experimental mass spectrum peaks showed that both bands corresponded to Als3. This conclusion was supported by lack of reactivity between MAb 3D9.3 and an als3 Δ /als3 Δ mutant strain, and the fact that an immunoglobulin preparation enriched for Als3 specificity recognized the purified 3D9 antigen. PCR demonstrated that C. albicans strain 66396 has two different-sized ALS3 alleles that correspond to the two purified protein bands. Strain- and species-specificity of the 3D9 epitope were studied with various C. albicans strains and Candida species, such as closely related Candida dubliniensis . The 3D9 epitope was detected only in C. albicans , demonstrating the utility of MAb 3D9.3 for differentiation between C. albicans and C. dubliniensis . Adhesion assays demonstrated that MAb 3D9.3 blocks adhesion of C. albicans germ tubes to human buccal epithelial cells and vascular endothelial cells.     

IL-33 priming regulates multiple steps of the neutrophil-mediated anti-Candida albicans response by modulating TLR and dectin-1 signals

IL-33 is known to play an important role in Th2 immunity. In this study, we investigated the effect of IL-33 pretreatment on anti-fungal response using an acute Candida albicans peritoneal infection model. IL-33 pretreatment induced a rapid fungal clearance and markedly reduced the C. albicans infection-associated mortality. The priming effect of IL-33 occurred during multiple steps of the neutrophil-mediated anti-fungal response. First, the anti-fungal effect occurred due to the rapid and massive recruitment of neutrophils to the site of infection as a result of the release of CXCR2 chemokines by peritoneal macrophages and by reversal of the TLR-induced reduction of CXCR2 expression in neutrophils during IL-33 priming. Second, conditioning of neutrophils by IL-33 activated the TLR and dectin-1 signaling pathways, leading to the upregulation of complement receptor 3 expression induced by C. albicans. Upregulated CR3 in turn increased the phagocytosis of opsonized C. albicans and resulted in the production of high levels of reactive oxygen species and the subsequent enhanced killing activity of neutrophils. Taken together, our results suggest that IL-33 can regulate the anti-fungal activity of neutrophils by collaborative modulation of the signaling pathways of different classes of innate immune receptors.     

Equilibrium analysis of binding of Candida albicans to human buccal epithelial cells

The effect of growth temperature on the binding of Candida albicans to human buccal epithelial cells (BECs) was examined using an equilibrium of binding analysis. Candida albicans was cultured in M9 medium either for 12 h at 25 degrees C or for 9 h at 25 degrees C and then shifted to 37 degrees C for 3 h. The temperature shift did not result in germ tube formation; however, the adherence of C. albicans to BECs was altered. Shifting temperature increased the yeast's ability to bind to BECs. A Langmuir adsorption isotherm was used to calculate the maximum number of available binding sites (N) and the apparent association constants of binding (Ka) for all resolvable adhesin-receptor interactions. Three classes of adhesin-receptor interactions were resolved when the yeast was cultured at 25 degrees C and included a low copy number site (N = 3.0 cfu/BEC; Ka = 2.11 X 10(-6) mL/cfu), a medium copy number site (N = 23.6 cfu/BEC, Ka = 8.21 X 10(-7) mL/cfu), and a high copy number site (N = 91.7 cfu/BEC, Ka = 3.35 X 10(-8) mL/cfu). Two classes of adhesin-receptor interactions were resolved when the incubation temperature was shifted to 37 degrees C: a low copy number site (N = 4.5 cfu/BEC, Ka = 3.98 X 10(-6) mL/cfu) and a high copy number site (N = 150.5 cfu/BEC, Ka = 8.47 X 10(-8) mL/cfu). Augmented C. albicans adherence to BECs due to the elevated growth temperatures appears to result from a temperature-regulated alteration in the C. albicans adhesin that recognizes a high copy number receptor site with relatively low affinity.     

Regulation of human polymorphonuclear neutrophil (PMN) activity against Candida albicans by large granular lymphocytes via release of a PMN-activating factor

Using a 24-hr radiolabel microassay developed in our laboratory that measures [3H]glucose uptake in residual Candida, we have identified the effector cells responsible for in vitro inhibition of Candida albicans growth as mainly polymorphonuclear neutrophils (PMN) and monocytes within the human peripheral blood cells. Highly purified T cells and large granular lymphocytes (LGL) that mediate natural killer activity which were obtained by Percoll density gradient centrifugation were found to have no innate activity against C. albicans. The LGL could not be activated by interferon-alpha, interferon-gamma or interleukin 2 to inhibit Candida growth although their K562 tumor cytotoxic activity was readily enhanced by these cytokines. Stimulation with heat-killed C. albicans also did not activate fungal growth inhibitory function in LGL and the supernatant of these activated LGL had no direct fungicidal activity. However, the activated LGL supernatant had the capability to enhance PMN function against C. albicans growth. Addition of recombinant human tumor necrosis factor, affinity-purified interferon-alpha, or interferon-gamma to PMN caused increased antifungal activity in PMN. However, antibodies to these cytokines had only a partial adverse effect on the ability of the activated LGL supernatant to stimulate PMN anti-Candida function. Therefore, the activated LGL supernatant appeared to contain a potent stimulator of PMN function which is as yet unidentified. These data indicate that LGL did not directly mediate anti-Candida activity but could indirectly influence C. albicans growth by activating PMN against the fungi through the release of a specific PMN-activating factor. Our findings therefore add another role to LGL which is the regulation of PMN function, the consequence of which is regulation of fungal immunity.     

Granulocyte colony-stimulating factor potentiates anti-Candida albicans growth inhibitory activity of polymorphonuclear cells

Abstract Granulocyte colony-stimulating factor (G-CSF) stimulates a subset of granulocyte colony forming cells and when administered to neutropenic individuals results in recovery of blood neutrophil numbers to normal levels. Therefore, G-CSF may be a useful therapeutic agent for infections in immunocompromised hosts. However, to date there has been only limited information that G-CSF activates the antimicrobial activity of neutrophils. In the present study, we found that recombinant G-CSF promotes the anti- Candida albicans activity of normal human blood polymorphonuclear (PMN) cells in vitro using both a ³H-glucose uptake procedure and a Candida colony counting assay. As little as 0.1 ng/ml G-CSF induced significant anti-Candida activity in the PMN cultures. G-CSF treatment also enhanced superoxide anion production by the PMNs in response to f-MLP as determined by the superoxide dismutase inhibitable cytochrome C reduction method. Such results show that G-CSF can promote the antimicrobial activity of peripheral blood PMNs against C. albicans .     

Differential adhesion of pathogenic Candida species to epithelial and inert surfaces

Abstract Growth in medium containing 500 mM galactose is known to promote the adhesion of Candida albicans to buccal epithelial cells or to acrylic in vitro. Of 5 other Candida species tested, only C. tropicalis (one strain) showed substantially increased adhesion to buccal cells (but not to acrylic) after growth under these conditions. A second strain of C. tropicalis as well as C. stellatoidea, C. parapsilosis, C. pseudotropicalis, C. guilliermondii and Saccharomyces cerevisiae showed little or no increased adhesion to either surface. However, after growth in medium containing 50 mM glucose, C. tropicalis and C. parapsilosis were significantly more adherent to acrylic than glucose-grown yeasts of the other species, including C. albicans . These results are discussed in relation to the colonization and infection potential of the pathogenic Candida species.     

In vitro effect of carbohydrates and enteric bacteria on adherence of Candida albicans

The adherence of Candida albicans to any cell is considered essential in the process that leads to colonization. Our objective in this study was to evaluate the effect of different carbohydrates and the presence of lactobacilli and Escherichia coli on the in vitro adherence of Candida albicans. The adherence to buccal epithelial cells was higher when growing at concentrations of galactose of 50, and 200 mM, as well as 50, 200, and 500 mM of sucrose, and 500 mM of mannose, compared with that obtained when growing in Sabouraud dextrose broth (p < 0.01). The presence of other microorganisms, such as Lactobacillus acidophilus and L. casei, caused a decrease in the in vitro adherence of C. albicans to buccal epithelial cells (p < 0.05), whereas E. coli did not modify this adherence at all.