Protective effect of acidic mannan fraction of bakers' yeast on experimental candidiasis in mice

An acidic fraction of bakers' yeast mannan, WAM025, showed a significant protective effect against Candida albicans infection in mice, but a neutral fraction of the same bakers' yeast mannan, WNM, did not exhibit this effect. Moreover, pretreatment with WAM025 resulted in a marked reduction of proliferation of C. albicans cells in the organs of the infected mice. We investigated the stimulative effect of these mannan fractions on the function of mouse peritoneal phagocytes, and found that mice administered WAM025 showed a greater increase in the number of peritoneal exudate cells, macrophages and polymorphonuclear leucocytes (PMN), than the mice treated with WNM, especially in the proportion of PMN. Peritoneal phagocytes, PMN and macrophages obtained from WAM025-treated mice showed marked candidacidal activity. Of the phagocytes, PMN were responsible for the larger part of the candidacidal activity. The myeloperoxidase activities of PMN and macrophages in WAM025-treated PEC were greater than in untreated macrophages. The myeloperoxidase activity of WAM025-treated PMN was significantly greater than that of WAM025-treated macrophages. This activity paralleled the active oxygen-releasing activity of the phagocytes. On the other hand, the phagocytic activity of phagocytes from mice administered WNM or WAM025 for C. albicans cells was identical to that of untreated phagocytes. WAM025 seems to cause enhance elimination of the pathogen from mice, by increasing the number and candidacidal activity of phagocytic cells.     

Studies towards the potential of poliovirus as a vector for the expression of HPV 16 virus-like-particles

In this study we tested the hypothesis that after administration of a single intraperitoneal dose of concanavalin A (Con-A) to mice, the proportion of neutrophils and macrophages in the peritoneal exudate and their phagocytic and candidacidal activities should change with time. The number of neutrophils in the peritoneal exudate was greatly increased 6 h after administration of Con-A, and those cells were able to kill both intracellular and extracellular yeast and germ tube forms of Candida albicans. Addition of catalase to the culture medium reduced the killing of C. albicans, suggesting that the candidacidal activity depended on the myeloperoxidase system. The survival of mice pretreated with Con-A and submitted to an inoculum of C. albicans 6 h afterwards was twice higher than that of controls, which suggests that neutrophils were able to clear the experimental infection. One day after the treatment, the population of neutrophils in the exudate was about 45%, but after 2 days it was reduced to only 5% and the candidacidal activity was also reduced. After 4 days the exudate contained over 95% of macrophages, the candidacidal activity reached a maximum, and the phagocytosis mediated by both complement receptors and mannose receptors was increased. Uptake of FITC-mannose-BSA by macrophages was maximal on about the 4th day and was inhibited by mannan, suggesting that treatment with Con-A increased the activity of mannose receptors. These results support the hypothesis that activation of cellular immunity by Con-A occurred in two phases, one dominated by neutrophils, and the other by macrophages expressing increased activity of mannose receptors.     

Protecting effect of chitin and chitosan on experimentally induced murine candidiasis

Chitin and chitosan were found to exhibit a protective effect on mice administered these polysaccharides intraperitoneally against infection of the viable cells of Candida albicans NIH A-207 strain. A significant difference was observed between the protective effects of chitin and chitosan, i.e., chitin was much more effective than chitosan when the C. albicans cells were challenged via the intravenous route. In intraperitoneal inoculations of C. albicans cells, however, chitosan provided stronger resistance for mice than chitin. It has also been revealed that the number of polymorphonuclear leukocytes from circulating blood of chitin-administered mice increased remarkably compared with that of untreated and chitosan-treated mice, and that the increase of active oxygen-generating phagocytic cells was significant. On the other hand, the number of peritoneal exudate cells (PEC) and the amounts of active oxygen generated from these cells in chitosan-treated mice were larger than those of chitin-treated mice. However, candidacidal activities of PEC per fixed cell number in mice treated with chitin or with chitosan were almost the same and greater than those of untreated mice.     

Polysaccharide-rich fraction of Agaricus brasiliensis enhances the candidacidal activity of murine macrophages

A polysaccharide-rich fraction (ATF) of medicinal mushroom Agaricus brasiliensis was evaluated on the candidacidal activity, H2O2 and nitric oxide (NO) production, and expression of mannose receptors by murine peritoneal macrophages. Mice received three intraperitoneal (i.p.) injections of ATF and after 48 h their peritoneal resident macrophages were assayed against Candida albicans yeast forms. The treatment increased fungicidal activity and it was associated with higher levels of H2O2, whereas NO production was not affected. We also found that the treatment enhances mannose receptor expression by peritoneal macrophages, which are involved in the attachment and phagocytosis of non-opsonized microorganisms. Treatment of animals with ATF was able to enhance the clearance of C. albicans during the first 6 h after the experimental i.p. infection. Our results suggest that this extract can increase host resistance against some infectious agents through the stimulation of microbicidal activity of macrophages.     

Lactoferrin feeding augments peritoneal macrophage activities in mice intraperitoneally injected with inactivated Candida albicans

Oral administration of lactoferrin (LF), an innate-defense protein present in exocrine secretions such as milk and in neutrophils, is reported to improve host-protection against infections with microorganisms including pathogenic fungi, possibly due to an immunomodulatory effect. This study aimed to evaluate the effect of bovine LF feeding on peritoneal macrophage activities in mice intraperitoneally injected with inactivated Candida albicans. Time course analysis during the 14 days following Candida-priming revealed that LF administration slightly increased the number of peritoneal exudate cells, and significantly enhanced the production of superoxide anion (O2(-)) and nitric oxide (NO) by peritoneal macrophages at day 7. LF administration facilitated NO production and Candida hyphal-growth inhibition by macrophages derived from Candida-primed mice but not non-primed mice, suggesting that the action of LF is dependent on the immune status of the host. LF administration altered the kinetics of cytokines in the peritoneal lavage fluid of Candida-primed mice. Enhancement of cytokine levels by LF was observed for IL-12 at day 5 and IFN-gamma at day 9, but not for TNF-alpha or IL-10. In conclusion, LF feeding augmented the activities of macrophages in a manner dependent on Candida-priming and these effects may be related to enhanced cytokine levels.     

Effect of N-acetylchitohexaose against Candida albicans infection of tumor-bearing mice

A water-soluble oligosaccharide, N-acetylchitohexaose (NACOS-6), was able to enhance the protective effect against Candida albicans infection in mice during the early phase of tumor-bearing. A significant decrease in the number of C. albicans cells in the kidneys of NACOS-6-treated tumor-bearing mice was observed 8 days after the fungal infection, or 15 days after the tumor transplantation. The candidacidal activity of polymorphonuclear leukocytes from NACOS-6-treated tumor-bearing mice did not differ from that of NACOS-6-untreated tumor-bearing mice. On the other hand, the candidacidal activities of both macrophages and T lymphocytes increased following administration of NACOS-6 in the early phase of tumor-bearing. The culture supernatant of T lymphocytes from NACOS-6-treated tumor-bearing mice also potentiated the candidacidal activity of casein-induced macrophages. An enhancement of natural killer cell activity of splenic lymphocytes obtained from NACOS-6-treated tumor-bearing mice was also observed.     

Toll-like receptor 2 suppresses immunity against Candida albicans through induction of IL-10 and regulatory T cells

Toll-like receptor (TLR) 2 and TLR4 play a pivotal role in recognition of Candida albicans. We demonstrate that TLR2(-/-) mice are more resistant to disseminated Candida infection, and this is associated with increased chemotaxis and enhanced candidacidal capacity of TLR2(-/-) macrophages. Although production of the proinflammatory cytokines TNF, IL-1alpha, and IL-1beta is normal, IL-10 release is severely impaired in the TLR2(-/-) mice. This is accompanied by a 50% decrease in the CD4+CD25+ regulatory T (Treg) cell population in TLR2(-/-) mice. In vitro studies confirmed that enhanced survival of Treg cells was induced by TLR2 agonists. The deleterious role of Treg cells on the innate immune response during disseminated candidiasis was underscored by the improved resistance to this infection after depletion of Treg cells. In conclusion, C. albicans induces immunosuppression through TLR2-derived signals that mediate increased IL-10 production and survival of Treg cells. This represents a novel mechanism in the pathogenesis of fungal infections.     

Low dose of Concanavalin-A enhances innate immune response and prevents liver injury in mice infected with Candida albicans

The mechanisms through which Candida albicans is recognized by immune cells and how it triggers host defence are not completely understood. In this study, we evaluated the effect of Concanavalin-A on the clearance of C. albicans by infected mice and their production of proinflammatory cytokine tumor necrosis factor-alpha (TNF-alpha). Subgroups of 5 animals were pretreated with Con-A (250 mug mL(-1) PBS) and after 96 h were infected intraperitoneally with 10(7) cells of C. albicans CR15 (an isolate from a HIV+ person); 30 min, 2, 6, 24 or 72 h after infection the mice were sacrificed. Phagocytosis of C. albicans by peritoneal macrophages increased 30 min after infection in mice pretreated with Con-A. The liver presented the greatest number of CFUs, and this number was reduced by pretreatment with Con-A. Control animals infected with C. albicans presented a significant increase in plasmatic alanine aminotransferase, which was not observed in mice treated with Con-A. Two hours after infection the production of TNF-alpha in the liver of mice pretreated with Con-A was significantly increased. These results suggest that a single dose of Con-A caused a beneficial modulating action of the inflammatory response during infection with C. albicans.     

Type I IFNs stimulate nitric oxide production and resistance to Trypanosoma cruzi infection

The participation of type I IFNs (IFN-I) in NO production and resistance to Trypanosoma cruzi infection was investigated. Adherent cells obtained from the peritoneal cavity of mice infected by the i.p. route produced NO and IFN-I. Synthesis of NO by these cells was partially inhibited by treatment with anti-IFN-alphabeta or anti-TNF-alpha Abs. Compared with susceptible BALB/c mice, peritoneal cells from parasite-infected resistant C57BL/6 mice produced more NO (2-fold), IFN-I (10-fold), and TNF-alpha (3.5-fold). Later in the infection, IFN-I levels measured in spleen cell (SC) cultures from 8-day infected mice were greater in C57BL/6 than in infected BALB/c mice, and treatment of the cultures with anti-IFN-alphabeta Ab reduced NO production. IFN-gamma or IL-10 production by SCs was not different between the two mouse strains; IL-4 was not detectable. Treatment of C57BL/6 mice with IFN-I reduced parasitemia levels in the acute phase of infection. Mice deprived of the IFN-alphabetaR gene developed 3-fold higher parasitemia levels in the acute phase in comparison with control 129Sv mice. Production of NO by peritoneal macrophages and SCs was reduced in mice that lacked signaling by IFN-alphabeta, whereas parasitism of macrophages was heavier than in control wild-type mice. We conclude that IFN-I costimulate NO synthesis early in T. cruzi infection, which contributes to a better control of the parasitemia in resistant mice.     

Antifungal type 1 responses are upregulated in IL-10-deficient mice

C57BL/6 mice are highly resistant to infections caused by Candida albicans and Aspergillus fumigatus. To elucidate the role of IL-10 produced by C57BL/6 mice during these infections, parameters of infection and immunity to it were evaluated in IL-10-deficient and wild-type mice with disseminated or gastrointestinal candidiasis or invasive pulmonary aspergillosis. Unlike parasitic protozoan infection, C. albicans or A. fumigatus infection did not induce significant acute toxicity in IL-10-deficient mice, who, instead, showed reduced fungal burden and fungal-associated inflammatory responses. The increased resistance to infections as compared to wild-type mice was associated with upregulation of innate and acquired antifungal Th1 responses, such as a dramatically higher production of IL-12, nitric oxide (NO) and TNF-alpha as well as IFN-gamma by CD4+ T cells. Pharmacological inhibition of NO production greatly reduced resistance to gastrointestinal candidiasis, thus pointing to the importance of IL-10-dependent NO regulation at mucosal sites in fungal infections. These results are reminiscent of those obtained in genetically susceptible mice, in which IL-10 administration increased, and IL-10 neutralization decreased, susceptibility to C. albicans and A. fumigatus infections. Collectively, these observations indicate that the absence of IL-10 augments innate and acquired antifungal immunity by upregulating type 1 cytokine responses. The resulting protective Th1 responses lead to a prompt reduction of fungal growth, thus preventing tissue destruction and lethal levels of proinflammatory cytokines.     

A novel B-2 suppressor cell regulating susceptibility/resistance of mice to Toxoplasma gondii infection

Irradiation treatment enhanced resistance of C57BL/6, but not BALB/c against Toxoplasma gondii infection. Six Gy-irradiated (IR) C57BL/6 recipients of B-2 cells from T. gondii-infected C57BL/6 died after infection. B-2 suppressor cells from infected C57BL/6 enhanced production of IL-4 and IL-10 in peritoneal exudate cells (PECs), and down-regulated NO release in peritoneal macrophages after infection. On the other hand, B-2 suppressor cells were not detected in a strain, BALB/c, resistant against infection. These data indicated that irradiation-sensitive B-2 cells regulated susceptibility/resistance in mice against T. gondii infection.     

Enhanced proinflammatory response to the Candida albicans gpi7 null mutant by murine cells

The Candida albicans gpi7/gpi7 null mutant strain (Deltagpi7), which is affected in glycosylphosphatidylinositol (GPI) anchor biosynthesis, showed a reduced virulence following systemic infection of C57BL/6 mice. In vitro production of TNF-alpha, IL-6 and IL-1beta by macrophages in response to Deltagpi7 cells was significantly increased as compared to control (wild type GPI7/GPI7 and revertant gpi7/GPI7) cells; this probably contributes to the enhanced recruitment of neutrophils to the peritoneal cavity in response to Deltagpi7 cells. Survival of knockout mice for Toll-like receptor (TLR) 2 and TLR4 following intravenous injection of Deltagpi7 cells showed no significant differences as compared to C57BL/6 mice. In vitro production of TNF-alpha by macrophages and neutrophil recruitment were significantly inhibited in TLR2-/- mice in response to control yeast strains. Interestingly both TNF-alpha production and neutrophil recruitment in response to Deltagpi7 were significantly increased in all three types of mice, with no differences among them, and laminarin failed to inhibit this increased production of TNF-alpha. These results indicate that the enhanced proinflammatory response to Deltagpi7 does not involve recognition through TLR2, TLR4 nor dectin-1. Therefore, complete GPI anchors confer surface properties that are involved in modulation of cytokine production by macrophages in response to C. albicans.     

Enhanced killing of Candida albicans by murine macrophages treated with macrophage colony-stimulating factor: evidence for augmented expression of mannose receptors

The effect of macrophage colony-stimulating factor (CSF-1) on killing of Candida albicans by murine peritoneal macrophages was determined. The killing capacity of resident peritoneal macrophages was unaffected by CSF-1. However, proteose-peptone-elicited peritoneal exudate macrophages that had been pretreated with CSF-1 (greater than or equal to 1000 U/ml) for 24 or 48 hr exhibited a significantly enhanced capacity to kill C. albicans. CSF-enhanced killing appeared to be independent of endogenously produced interferon-alpha/beta (IFN) in that enhancement by these two agents differed with regard to onset of the effect, target cell responsiveness, and duration of augmented killing. In addition, a highly specific anti-IFN antiserum that totally neutralized IFN augmentation of candidacidal activity had no effect on CSF-induced enhancement. Evidence was obtained indicating that CSF, unlike IFN, augmented mannose-inhibitable binding and ingestion of C. albicans, suggesting that augmented expression of mannose-receptors by CSF-treated macrophages was at least partially responsible for the enhanced killing.     

Antifungal and antitumor activities of a monoclonal antibody directed against a stress mannoprotein of Candida albicans

Immunization of mice with a stress mannoprotein of >200 kDa from the cell wall of Candida albicans led to the production of monoclonal antibody (Mab) C7. The immunogen is a major target of secretory IgA and its expression is regulated by different environmental conditions including temperature, pH, glucose concentration and ammonium sulphate in the culture medium. Mab C7 reacted with a peptide epitope present in the >200 kDa antigen as well as in a number of antigens from the blastoconidium and germ tube cell wall, including enolase. In addition to its reactivity with C. albicans, Mab C7 also reacted with antigens present in C. krusei, C, tropicalis, C. glabrata, C. dubliniensis and C. lusitaniae, as well as in Cryptococcus neoformans, Scedosporium prolificans and Aspergillus fumigatus. Mab C7 exhibited four important biological activities, namely inhibition of adhesion of C. albicans to a variety of surfaces, inhibition of germination of C. albicans, direct candidacidal activity and direct tumoricidal activity. In tumor cells, Mab C7 reacted with nucleoporin Nup88, a reactivity that can be utilized for diagnostic and prognostic purposes.     

Induction of candidicidal activity in mice by immunization with Candida albicans ribosomes

Abstract Mice immunized with ribosomes from Candida albicans are protected against experimental systemic candidiasis. In this study we investigated the candidacidal activity of spleen cells from immunized animals as measured by ⁵¹Cr release from pre-labelled yeast cells. It was found that the anti-candidal cytotoxic activity of splenocytes from immunized mice was significantly higher than that of spleen cells from non-immunized controls with various effector to target (E:T) ratios, but optimal results were obtained with an E:T ratio of 10:1. The cytotoxic activity of splenocytes as measured by the ⁵¹Cr release assay correlated well with the capacity of the cells to inhibit candidal growth as determined by quantitative plating. This candidacidal activity was not antibody dependent but increased killing was obtained by adding fresh (but not heat inactivated) mouse serum. The enhanced candidicidal activity was inhibited by removal of plastic- or nylon-adherent cells from the cell suspension but not by treatment with anti-Thy 1.2 serum and complement. The data indicate that a candidacidal cell population is induced in the spleens of animals immunized with C. albicans ribosomes.     

Interaction between LPS-induced NO production and IDO activity in mouse peritoneal cells in the presence of activated Vα14 NKT cells

In this study, we demonstrated that lipopolysaccharide (LPS) markedly increased nitric oxide (NO) production and indoleamine 2,3-dioxygenase (IDO) activity in mouse peritoneal cells in the presence of activated Vα14 natural killer T cells. Moreover, LPS-induced NO production in peritoneal cells from IDO-knockout (KO) mice was more increased than that from wild-type mice. However, there was no significant difference in the expression of inducible nitric oxide synthase (iNOS) mRNA and protein between the wild-type and IDO-KO mice. No significant difference was also observed in the ratio of CD3- and DX5-positive cells and F4/80- and TLR4-positive cells in peritoneal cells between the wild-type and IDO-KO mice. Since the IDO activity was enhanced by an NO inhibitor, NO may be post-translationally consumed by inhibiting the IDO activity. IDO is well known to play an important role in immunosuppression during inflammatory disease. Therefore, the inhibition of IDO by NO may exacerbate inflammation in the peritoneal cavity.     

LAAE-14, a new anti-inflammatory drug, increases the survival of Candida albicans-inoculated mice

LAAE-14, a lipidic acid-amido ether derivative, has been recently described as a new anti-inflammatory drug. We have studied the effect of treatment with this compound on the susceptibility of mice to in vivo experimental Candida albicans infection. ICR mice orally treated with LAAE-14 (25 mg kg(-1)) and experimentally intravenously infected showed a significantly increased survival as compared to control mice. In vitro, the compound did not inhibit the growth of C. albicans yeast cells or the yeast-to-hyphal transition. The in vitro production of prostaglandin E2 by peritoneal macrophages in response to the yeasts and hyphae of C. albicans was significantly decreased upon treatment with LAAE-14, in a dose-dependent manner. Thus, reduced prostaglandin production during fungal infection could be an important factor in controlling fungal colonisation and infection.     

Candida albicans double-stranded DNA can participate in the host defense against disseminated candidiasis

In the present work, we studied the in vitro immunomodulatory properties of double-stranded Candida albicans DNA and its protective effect in murine disseminated candidiasis. DNA induced the production of TNF-alpha by peritoneal macrophages and splenocytes in vitro through a chloroquine-dependent mechanism. Yeast DNA acted synergistically with IFN-gamma in triggering the secretion of nitric oxide by macrophages and enabled them to stimulate the proliferation of T cells in response to soluble anti-CD3. The effect of DNA on splenocytes is associated with an enhanced synthesis of IFN-gamma, IL-2 and IL-10. In vivo, DNA decreased the mortality and lowered the kidney contamination in mice intraperitoneally inoculated with C. albicans simultaneously with an increase in the specific proliferative response and cytokine production. The present results indicate that C. albicans DNA can provide protection against disseminated infection.     

Production and function of cytokines in natural and acquired immunity to Candida albicans infection.

Host resistance against infections caused by the yeast Candida albicans is mediated predominantly by polymorphonuclear leukocytes and macrophages. Antigens of Candida stimulate lymphocyte proliferation and cytokine synthesis, and in both humans and mice, these cytokines enhance the candidacidal functions of the phagocytic cells. In systemic candidiasis in mice, cytokine production has been found to be a function of the CD4+ T helper (Th) cells. The Th1 subset of these cells, characterized by the production of gamma interferon and interleukin-2, is associated with macrophage activation and enhanced resistance against reinfection, whereas the Th2 subset, which produces interleukins-4, -6, and -10, is linked to the development of chronic disease. However, other models have generated divergent data. Mucosal infection generally elicits Th1-type cytokine responses and protection from systemic challenge, and identification of cytokine mRNA present in infected tissues of mice that develop mild or severe lesions does not show pure Th1- or Th2-type responses. Furthermore, antigens of C. albicans, mannan in particular, can induce suppressor cells that modulate both specific and nonspecific cellular and humoral immune responses, and there is an emerging body of evidence that molecular mimicry may affect the efficiency of anti-Candida responses within defined genetic contexts.     

Murine Defense Mechanism against Candida albicans Infection: II. Opsonization,Phagocytosis, and Intracellular Killing of C. albicans

The phagocytic and intracellular killing activities of normal mouse phagocytes against Candida albicans were studied to elucidate the role of these activities in nonspecific and specific defense mechanisms. In the presence of fresh normal mouse serum, viable C. albicans cells were ingested by mouse peripheral blood leukocytes (PBLs) and peritoneal macrophages (PMPs) at the same rate. Serum-chelation experiments indicated that the factors involved in the alternative complement pathway are opsonins for C. albicans. PBLs killed intracellular C. albicans more effectively than PMPs. Lymphokine-activated PMPs manifested marked intracellular killing activity and the occurrence of increased superoxide anion- and singlet oxygen production, in the absence of increased myeloperoxidase (MPO) production, suggests that the enhanced, MPO-independent, oxidative mechanism may play an important role in the candidacidal activity. Specific rabbit antibodies played no role in the phagocytosis and intracellular killing of C. albicans. These results suggest that PMNs and factors involved in the alternative complement pathway, and lymphokine-activated macrophages play major roles in the protection of mice against C. albicans infection.