- Immunological phenomena associated with chronic mucocutaneous candidiasis have recently been intensively studied by many workers (reviewed by Kirkpatrick, Rich & Bennett, 1971). The results have shown that there is no common immunological denominator in this disease. The most common finding, however, is defective cellular immunity, which may or may not be accompanied by failure of in vitro lymphocyte transformation.
Immunological Feautures in a Case of Chronic Granulomatous Candidiasis and its Treatment with Transfer Factor
VALDIMARSSON, C. B. S. WOOD, J. R. HOBBS AND P. J. L. HOLT
- The ability of Candida albicans to establish an infection involves multiple components of this fungal pathogen, but its ability to persist in host tissue may involve primarily the immunosuppressive property of a major cell wall glycoprotein, mannan. Mannan and oligosaccharide fragments of mannan are potent inhibitors of cell-mediated immunity and appear to reproduce the immune deficit of patients with the mucocutaneous form of candidiasis. However, neither the exact structures of these inhibitory species nor their mechanisms of action have yet been clearly defined. Different investigators have proposed that mannan or mannan catabolites act upon monocytes or suppressor T lymphocytes, but research from unrelated areas has provided still other possibilities for consideration. These include interference with cytokine activities, lymphocyte-monocyte interactions, and leukocyte homing. To stimulate further research of the immunosuppressive property of C. albicans mannan, we have reviewed (i) the relationship of mannan to other antigens and virulence factors of the fungus; (ii) the chemistry of mannan, together with methods for preparation of mannan and mannan fragments; and (iii) the historical evidence for immunosuppression by Candida mannan and the mechanisms currently proposed for this property; and (iv) we have speculated upon still other mechanisms by which mannan might influence host defense functions. It is possible that understanding the immunosuppressive effects of mannan will provide clues to novel therapies for candidiasis that will enhance the efficacy of both available and future anti-Candida agents. Immunosuppressive properties observed for isolated Candida mannan and its catabolites in vivo and in vitro provide additional evidence that fungal mannan is responsible for patient immune dysfunction.
Candida mannan: chemistry, suppression of cell-mediated immunity, and possible mechanisms of action.
R D Nelson, N Shibata, R P Podzorski, and M J Herron
- Phagocytic cells of the innate immune system, such as macrophages and neutrophils, are a primary line of defense against microbial infections. Patients with defects in innate immunity, such as those with chronic granulomatous disease or neutropenia, are extremely sensitive to a variety of infections. When a phagocyte recognizes the presence of an invading cell, it engulfs the microbe with its membrane to form the phagosome, an intracellular compartment containing the microbe. This compartment matures by fusion with lysosomes to create the phagolysosome, an organelle replete with antimicrobial compounds and an acidic pH. Internalization creates a hostile environment for the microorganism, which, of course, is the intent. The phagolysosome is a precarious neighborhood even before the onslaught of antimicrobial compounds. Engulfment by the macrophage thrusts the microorganism into an alien milieu, one devoid of key nutrients necessary for metabolism and division. Surviving the antimicrobial assault in the phagolysosome depends on the microbe’s ability to synthesize the proteins and other cellular components necessary to counteract these stresses. Thus, a pathogen must find the requisite nutrients to provide the building blocks for these complex macromolecules and the energy with which to synthesize them.
In this article we consider the initial responses of several microbes to nutrient deprivation inside the macrophage. The first of these, Mycobacterium tuberculosis, the bacterium that causes tuberculosis, resides for prolonged periods within the macrophage, in which it can proliferate and subsequently spread throughout the body. The second, the yeast Saccharomyces cerevisiae, is killed efficiently by the macrophage. The third, the opportunistic fungal pathogen Candida albicans, survives ingestion by changing rapidly from a yeast to a filamentous morphology, lysing the macrophage from the inside out. Once free, C. albicans cells are able to disseminate through the body. The interaction of C. albicans with the macrophage is transient, as opposed to the long-term persistence of M. tuberculosis. Although the outcomes of this macrophage capture are quite different among the three microbes, the initial responses of all three to the internal environment are remarkably similar: induction of the glyoxylate cycle, a pathway that permits the utilization of compounds with two carbons (C2 compounds), such as acetate, to satisfy cellular carbon requirements.
Systemic fungal infections have increased dramatically in prevalence and severity over the last few decades, in concert with the number of patients living for extended periods with significant immune dysfunction. AIDS, cancer chemotherapy, and organ transplantation have all contributed to this rise, as has the widespread use of antibiotics. The most common systemic fungal infection is candidiasis, which accounts for well over half of these invasive mycoses (3). A single species, C. albicans, causes the majority of these infections. C. albicans, which also causes oropharyngeal thrush and vaginitis, is normally a commensal of the mammalian gastrointestinal tract, in which it lives without adverse effects on the host. Both C. albicans and S. cerevisiae are readily phagocytosed by cultured macrophages in the presence of serum. While the macrophages efficiently kill S. cerevisiae, engulfment induces C. albicans cells to grow in a filamentous morphology. These hyphal filaments can penetrate through the membrane of the phagocytic cell, releasing the fungal cell back into the extracellular medium while killing the macrophage in the process. The different outcomes are not surprising; C. albicans is a common pathogen while S. cerevisiae is rarely found in human hosts.
The primary function of the glyoxylate cycle is to permit growth when C2 compounds, such as ethanol and acetate, are the only sources of carbon. The glyoxylate pathway (also dubbed the glyoxylate shunt, for clear reasons) bypasses these decarboxylations, allowing C2 compounds to serve as carbon sources in gluconeogenesis and to be incorporated into glucose and, from there, into amino acids, DNA, and RNA. Glucose, as the preferred carbon source in most organisms, can be both converted into five-carbon sugars (such as ribose and deoxyribose) via the pentose phosphate pathway and catabolized to acetyl-CoA via glycolysis. In microorganisms, however, glucose is frequently not available, and simple carbon compounds provide the only accessible carbon.
With the population of immunocompromised people on the rise, the frequency of invasive fungal infections continues to increase, making the need for effective treatments more imperative.
Life and Death in a Macrophage: Role of the Glyoxylate Cycle in Virulence
Michael C. Lorenz and Gerald R. Fink
- Fungal pathogens can be recognized by the immune system via their beta-glucan, a potent proinflammatory molecule that is present at high levels but is predominantly buried beneath a mannoprotein coat and invisible to the host. To investigate the nature and significance of “masking” this molecule, we characterized the mechanism of masking and consequences of unmasking for immune recognition. We found that the underlying beta-glucan in the cell wall of Candida albicans is unmasked by subinhibitory doses of the antifungal drug caspofungin, causing the exposed fungi to elicit a stronger immune response. Using a library of bakers’ yeast (Saccharomyces cerevisiae) mutants, we uncovered a conserved genetic network that is required for concealing beta-glucan from the immune system and limiting the host response. Perturbation of parts of this network in the pathogen C. albicans caused unmasking of its beta-glucan, leading to increased beta-glucan receptor-dependent elicitation of key proinflammatory cytokines from primary mouse macrophages. By creating an anti-inflammatory barrier to mask beta-glucan, opportunistic fungi may promote commensal colonization and have an increased propensity for causing disease.
A drug-sensitive genetic network masks fungi from the immune system.
- The interaction of Candida albicans with macrophages is considered a crucial step in the development of an adequate immune response in systemic candidiasis. An in vitro model of phagocytosis that includes a differential staining procedure to discriminate between internalized and non-internalized yeast was developed. Upon optimization of a protocol to obtain an enriched population of ingested yeasts, a thorough genomics and proteomics analysis was carried out on these cells. Both proteins and mRNA were obtained from the same sample and analyzed in parallel. The combination of two-dimensional PAGE with MS revealed a total of 132 differentially expressed yeast protein species upon macrophage interaction. Among these species, 67 unique proteins were identified. This is the first time that a proteomics approach has been used to study C. albicans-macrophage interaction. We provide evidence of a rapid protein response of the fungus to adapt to the new environment inside the phagosome by changing the expression of proteins belonging to different pathways. The clear down-regulation of the carbon-compound metabolism, plus the up-regulation of lipid, fatty acid, glyoxylate, and tricarboxylic acid cycles, indicates that yeast shifts to a starvation mode. There is an important activation of the degradation and detoxification protein machinery. The complementary genomics approach led to the detection of specific pathways related to the virulence of Candida. Network analyses allowed us to generate a hypothetical model of Candida cell death after macrophage interaction, highlighting the interconnection between actin cytoskeleton, mitochondria, and autophagy in the regulation of apoptosis. In conclusion, the combination of genomics, proteomics, and network analyses is a powerful strategy to better understand the complex host-pathogen interactions.
Integrated proteomics and genomics strategies bring new insight into Candida albicans response upon macrophage interaction.
- Intestinal homeostasis is critical for efficient energy extraction from food and protection from pathogens. Its disruption can lead to an array of severe illnesses with major impacts on public health, such as inflammatory bowel disease characterized by self-destructive intestinal immunity. However, the mechanisms regulating the equilibrium between the large bacterial flora and the immune system remain unclear. Intestinal lymphoid tissues generate flora-reactive IgA-producing B cells, and include Peyer’s patches and mesenteric lymph nodes, as well as numerous isolated lymphoid follicles (ILFs). Here we show that peptidoglycan from Gram-negative bacteria is necessary and sufficient to induce the genesis of ILFs in mice through recognition by the NOD1 (nucleotide-binding oligomerization domain containing 1) innate receptor in epithelial cells, and -defensin 3- and CCL20-mediated signalling through the chemokine receptor CCR6. Maturation of ILFs into large B-cell clusters requires subsequent detection of bacteria by toll-like receptors. In the absence of ILFs, the composition of the intestinal bacterial community is profoundly altered. Our results demonstrate that intestinal bacterial commensals and the immune system communicate through an innate detection system to generate adaptive lymphoid tissues and maintain intestinal homeostasis.
Lymphoid tissue genesis induced by commensals through NOD1 regulates intestinal homeostasis
Djahida Bouskra, Christophe Brézillon, Marion Bérard, Catherine Werts, Rosa Varona, Ivo Gomperts Boneca & Gérard Eberl
http://www.nature.com/nature/journal/v456/n7221/full/nature07450.html – a1#a1
- Candida albicans is a component of the normal flora of the alimentary tract and also is found on the mucocutaneous membranes of the healthy host. However, when immune defenses are compromised or the normal microflora balance is disrupted, Candida transforms itself into an opportunistic pathogenic killer. Candida is the leading cause of invasive fungal disease in premature infants, diabetics, and surgical patients, and of oropharyngeal disease in AIDS patients. As the induction of cell-mediated immunity to Candida is of critical importance in host defense, we sought to determine whether human dendritic cells (DC) could phagocytose and degrade Candida and subsequently present Candida antigens to T cells. Immature DC obtained by culture of human monocytes in the presence of granulocyte-macrophage colony-stimulating factor and interleukin-4 phagocytosed unopsonized Candida in a time-dependent manner, and phagocytosis was not enhanced by opsonization of Candida in serum. Like macrophages (M), DC recognized Candida by the mannose-fucose receptor. Upon ingestion, DC killed Candida as efficiently as human M, and fungicidal activity was not enhanced by the presence of fresh serum. Although phagocytosis of Candida by DC stimulated the production of superoxide anion, inhibitors of the respiratory burst (or NO production) did not inhibit killing of Candida, even when phagocytosis was blocked by preincubation of DC with cytochalasin D. Further, although apparently only modest phagolysosomal fusion occurred upon DC phagocytosis of Candida, killing of Candida under anaerobic conditions was almost equivalent to killing under aerobic conditions. Finally, DC stimulated Candida-specific lymphocyte proliferation in a concentration-dependent manner after phagocytosis of both viable and heat-killed Candida cells. These data suggest that, in vivo, such interactions between DC and C. albicans may facilitate the induction of cell-mediated immunity. Despite appropriate therapy, mortality from systemic Candida infections in immunocompromised individuals is nearly 30%. In human immunodeficiency virus-infected individuals who have not yet developed advanced immunodeficiency, the prevalence of oropharyngeal Candida is from 7 to 48% of patients. As the immunodeficiency in AIDS patients progresses, the prevalence of oral candidiasis increases to 43 to 93%. Furthermore, the development of oral candidiasis in the early stages of human immunodeficiency virus infection is highly predictive of worsening immunodeficiency.
Candida albicans Is Phagocytosed, Killed, and Processed for Antigen Presentation by Human Dendritic Cells
Simon L. Newman and Angela Holly
- Systemic candidiasis remains a major cause of disease and death, particularly among patients suffering from hematological malignancies. In an attempt to contribute to the discovery of useful biomarkers for its diagnosis and therapeutic monitoring, we embarked on a mapping of Candida albicans immunogenic proteins specifically recognized by antibodies produced during the natural course of systemic Candida infection in this high-risk population. About 85 immunoreactive protein species were detected with systemic candidiasis patients’ serum specimens by using immunoproteomics (i.e., two-dimensional electrophoresis followed by Western blotting), and identified through a combination of peptide mass fingerprinting by matrix-assisted laser desorption/ionization-time of flight-mass spectrometry (MALDI-TOF-MS), de novo peptide sequencing using nano-electrospray ionization-ion trap (ESI-IT) MS, and genomic database searches. This proteomic approach has led to the characterization of 42 different housekeeping enzymes as C. albicans antigens. Their biological significance is also discussed. Furthermore, this study is the first to report that 26 of them exhibit antigenic properties in C. albicans, and 35 of them become targets of the human antibody response to systemic candidiasis. Our findings suggest that the production of antibodies to C. albicans phosphoglycerate kinase and alcohol dehydrogenase during systemic candidiasis could be associated with a differentiation of the human immune response. We also highlight the relationship between changes in maintenance of circulating levels of specific anti-Candida antibodies and patients’ outcome. Some of these variations, especially the rise of high anti-enolase antibody concentrations, appear to be related to recovery from systemic candidiasis in these patients, which might serve as markers for predicting their outcome.
Proteomics-based identification of novel Candida albicans antigens for diagnosis of systemic candidiasis in patients with underlying hematological malignancies.
- Humans rely on their native microbiota for nutrition and resistance to colonization by pathogens; furthermore, recent discoveries have shown that symbiotic microbes make essential contributions to the development, metabolism, and immune response of the host. Co-evolved, beneficial, human–microbe interactions can be altered by many aspects of a modern lifestyle, including urbanization, global travel, and dietary changes, but in particular by antibiotics. The acute effects of antibiotic treatment on the native gut microbiota range from self-limiting “functional” diarrhea to life-threatening pseudomembranous colitis. The long-term consequences of such perturbations for the human–microbial symbiosis are more difficult to discern, but chronic conditions such as asthma and atopic disease have been associated with childhood antibiotic use and an altered intestinal microbiota. Because many chemical transformations in the gut are mediated by specific microbial populations, with implications for cancer and obesity, among other conditions, changes in the composition of the gut microbiota could have important but undiscovered health effects.
The Pervasive Effects of an Antibiotic on the Human Gut Microbiota, as Revealed by Deep 16S rRNA Sequencing
Les Dethlefsen , Sue Huse, Mitchell L. Sogin, David A. Relman
- A new study has uncovered the genetic wiring diagram underlying the infectiousness of Candida albicans, a fungus that causes thrush in babies, vaginal infections in women, and life-threatening infections in chemotherapy and AIDS patients. The study, led by Dr. Gerald R. Fink, Director of the Whitehead Institute for Biomedical Research, reveals that one key to Candida’s infectiousness lies in its ability to switch from a rounded form to filamentous forms. Fungal infections in hospitalized patients have almost doubled throughout the 1980s, often with life-threatening results in individuals with weakened immune systems. Candida, in particular, poses a serious threat and is associated with high mortality rates in patients undergoing chemotherapy. Candida is also a major cause of infection in hospitalized patients, especially those in Intensive Care Units, patients after major injuries or surgery, patients with burns, and premature babies.
In this study, Dr. Fink and his colleagues used molecular biology techniques to identify the components of the filamentation circuit in yeast. With the recently completed yeast genome to guide them, the scientists began to knock out suspicious genes and, by a process of elimination, discovered the culprits that are responsible for filamentation. Once scientists identified the key yeast filamentation genes, they simply plucked out the analogous genes in Candida. “Candida albicans is three hundred million years apart evolutionarily from yeast-as far away in evolution as humans are from turtles-and yet, the basic logic circuit was conserved,” says Dr. Fink.
Discovery Of Genetic Pathways May Provide New Ways To Combat Candida Infections
Gerald R. Fink, et al.
- The mechanisms by which corticosteroids (CCs) improve the outcome of AIDS patients with severe Pneumocystis carinii pneumonia (PCP) are unclear. We studied IL-1b and TNFα release from alveolar macrophages (AMs) of patients receiving CCs for the treatment of PCP and also the effect of in vitro hydrocortisone on this release. Cytokine release from AMs of AIDS patients with pulmonary complications not receiving CCs (group 1) was compared with that from AM of those receiving CCs for PCP (group 2). The AMs of HIV-negative normal subjects (group 3) served as controls. AU participants were nonsmokers or exsmokers. We found that lipopolysaccharide-stimulated AM from group 2 released significantly less interleukin-1 beta (IL-1b) and tumor necrosis factor alpha (TNFα) than AM from group 1 and was similar to that from group 3
Effect of corticosteroids on IL 1β and TNFα releae by alveolar macrophages from patients with AIDS and Pneumocystis carinii pneumonia
ZHENG BO HUANG ; EDEN E.
- A transporter in the colon called SLC5A8 plays an important role in enabling the colon to get the last bit of good out of food before the unusable is flushed away, according to research currently published online as an accelerated communication in the Journal of Biological Chemistry. The finding that SLC5A8 is the transporter helps clarify why fruits and vegetables are good for you and why antibiotics, which wipe out good bacteria along with bad, should only be taken when absolutely necessary, upset the model and colonic cells get sick and may even become cancerous. “We do not make the enzyme to digest cellulose; bacteria make the enzyme in the colon,” says Dr. Ganapathy. “Therefore, you need to eat dietary fiber to provide the food for bacteria. Otherwise, they are not going to su'”/>rvive there. Antibiotics can wipe out good bacteria as well, leaving a void where disease-causing bacteria can grow.” “The gut is a huge immune organ; there are more immune cells in our gut than there are in the rest of the body put together,” says Dr. Robert G. Martindale, MCG gastrointestinal surgeon and nutritionist with a special interest in probiotics giving patients good bacteria to restore a healthy flora. “The work that Dr. Ganapathy is doing is showing very nicely that if, in fact, we keep this short-chain fatty acid transporter healthy, we then can keep the whole immune system healthy.” Immune cells also have a specific receptor for short-chain fatty acids on the cell surface, and Drs. Ganapathy and Martindale are pursuing the idea that the SLC5A8 transporter is delivering these fatty acids to immune cells to interact with the receptors and keep the immune cells vigilant as well.
Transporter’s function provides support for eating vegetables, limiting antibiotics
Dr. Vadivel Ganapathy
- Although Candida albicans is present in many mammals including humans, normal bacterial flora and various immune factors usually restrict the growth of C. albicans in the alimentary tracts of immune competent hosts. Infection of the alimentary tract mucosae, including the mucosae of the oropharynx, esophagus, and gastrointestinal tract, with C. albicans is occurring with greater frequency, presumably because of the increased population of immune compromised individuals. Recent evidence suggests that cell-mediated immunity, and more specifically, CD41 T lymphocytes, play an important role in resistance to mucosal candidiasis. Patient populations with AIDS or other defects in cellular immune function show an increased incidence of mucocutaneous, but not necessarily disseminated, candidiasis, whereas patients with phagocytic cell defects, such as those that occur in patients with neutropenic or chronic granulomatous disease states, show a higher incidence of disseminated candidiasis. A combination of defective cell-mediated immunity and phagocytic cell defects in athymic beige (bg/bg nu/nu) mice was found to predispose them to severe mucosal candidiasis with subsequent Candida dissemination. Existing mouse models of mucosal candidiasis use combinations of chemically induced immune suppression, elimination or alteration of the host microflora by administration of antibiotics, high inocula, trauma, infant animals, or animals with congenital, functional, physiological, immunological, or metabolic defects to facilitate colonization of the gastrointestinal tract by C. albicans.
New Model Of Oropharyngeal and Gastrointestinal Colonization by Candida albicans in CD41 T-Cell-Deficient Mice for Evaluation of Antifungal Agents
AMY M. FLATTERY, GEORGE K. ABRUZZO, CHARLES J. GILL, JEFFREY G. SMITH, AND KEN BARTIZAL
- Underlying acquired immunity to the fungus Candida albicans is usually present in adult immunocompetent individuals and is presumed to prevent mucosal colonization progressing to symptomatic infection. Exploration of immunological events leading to Candida resistance or susceptibility has indicated the central role of the innate and adaptive immune systems, the relative contribution of which may vary depending on the site of the primary infection. Nevertheless, acquired resistance to infection results from the development of Th1 responses. Cytokines produced by Th1 cells activate phagocytic cells to a Candidacidal state. In contrast, cytokines produced by Th2 cells inhibit Th1 development and deactivate phagocytic effector cells. Because reciprocal influences have been recognized between innate and adaptive Th immunity, it appears that an integrated immune response determines the life-long commensalism of the fungus at the mucosal level, as well as the transition from mucosal saprophyte to pathogen. However, if the ability of C. albicans to establish a disseminated infection involves neutropenia as a major predisposing factor, its ability to persist in infected tissues or to behave as a commensal may involve primarily downregulation of host cell-mediated adaptive immunity. As a commensal, C. albicans may be endowed with the ability to elude the host’s immunological surveillance, thus allowing its persistence on mucosal surfaces. Th1 and Th2 CD41 T-cells develop from a common, naïve CD41 T-cell precursor, and several parameters have been shown to influence the pathway of differentiation of CD41 T-cell precursors. Among these, cytokines appear to play a major role, acting not only as modulators of antifungal effector functions but also as key regulators in the development of the different Th subsets from precursor Th cells. Studies in mice have shown that development of protective AntiCandidal Th1 responses requires the concerted actions of several cytokines, such as interferon (IFN)-g, transforming growth factor (TGF)-b, interleukin (IL)-6 , tumor necrosis factor (TNF)-a, and IL-12, in the relative absence of inhibitory Th2 cytokines, such as IL-4 and IL-10, which inhibit development of Th1 responses. Early in infection, neutralization of Th1 cytokines (IFN-g and IL-12) leads to the onset of Th2 rather than Th1 responses, while neutralization of Th2 cytokines (IL-4 and IL-10) allows development of Th1- rather than Th2-cell responses. TNF/lymphotoxin (LT)-a and IL-6 deficiencies render mice highly susceptible to C. albicans infections. Studies in humans have reinforced this concept, by showing that acquired immunity to C. albicans correlates with the expression of local or peripheral Th1 reactivity, whereas susceptibility to the infection seen in thermally injured patients, in patients with human immunodeficiency virus (HIV) infection, or in patients with chronic mucocutaneous or hepatosplenic candidiasis correlates with a biased Th2 response to the fungus. Altogether these data demonstrate that susceptibility to primary and secondary C. albicans infections in cytokine-deficient mice correlates with the failure to develop antiCandidal, protective Th1 responses and with the occurrence of unprotective IL-4- and IL-10-producing Th2 cells. However, an important immunoregulatory role has been attributed to neutrophils recently. Neutrophils, more than macrophages, were endowed with the ability to produce directive cytokines such as IL-10 and IL-12. Most importantly, IL-12 appeared to be released in response to a low-virulence Candida strain that initiates Th1 development in vivo, but IL-10 was released in response to a virulent strain. Human neutrophils also produced bioactive IL-12 in response to a mannoprotein fraction of C. albicans, capable of inducing Th1 cytokine expression in peripheral blood mononuclear cells. By producing directive cytokines such as IL-10 and IL-12, neutrophils influenced antifungal Th-cell development, as evidenced by the inability of neutropenic mice to mount protective antiCandidal Th1 responses. Production of IL-12 by neutrophils occurred independently of TNF-a and IFN-g. It was impaired upon iron overload but increased upon in vitro priming with IL-4 through upregulation of IL-4 receptor expression. Human studies confirm the multiple and complex role neutrophils have in candidiasis. First, risk factors for invasive fungal infections are not the same in all neutropenic patients. Secondly, chronic systemic candidiasis initiated by neutropenia may persist in spite of normal neutrophil counts and adequate antifungal therapy. Third, some patients, particularly transplant recipients who have adequate or even normal neutrophil counts, may be at high risk for invasive mycoses.
Innate and adaptive immunity in Candida albicans infections and saprophytism
- Candida albicans is a common commensal organism that occasionally causes opportunistic infections. As shown by the increased number of fungal infections in AIDS, the frequency of candidiasis has rapidly increased during the last 2 decades. In addition to AIDS, immunosuppression is induced by treatments of solid malignant tumors, lymphoproliferative disorders, and organ transplantation. In immunocompromised patients, Candida cells easily invade the host’s organs and multiply, causing lethal damage to the lungs, kidneys, liver, and intestines.
The prevention and treatment of Candidal infection have therefore become important for immunocompromised patients. Although the host’s defense system against Candida cells has not yet been completely clarified, it has been reported that both humoral and cellular immunities contribute to protection against Candida cells. In the former, antibodies to Candida cell antigens enhance phagocytosis of neutrophils and macrophages. Salivary proteins, such as secretory immunoglobulin A, secretory components, histatins, lysozyme, lactoferrin, transferrin, lactoperoxidase, mucins, and defensins have also been nominated as the humoral agents that prevent Candida cell adhesion and growth in the oropharyngeal cavity, whereas cellular agents, such as neutrophils, macrophages, and T and NK cells, play important roles in the front line against Candida cells, exhibiting phagocytosis and killing. For sufficient phagocytosis, opsonization of Candida cells is required. However, macrophages can trap nonopsonized blastoconidia by using their mannose receptors. To kill the trapped blastoconidia sufficiently, neutrophils and macrophages generate reactive oxygen intermediates (ROI) and nitric oxide (NO). The generation of ROI and NO is regulated by multiple cytokines. Among them, granulocyte-macrophage colony-stimulating factor (GM-CSF), interferons, and prostaglandins strongly induce NO synthase (NOS) and activate other enzymes associated with ROI generation. However, the virulence of blastoconidia is correlated with their resistance to phagocytes. It has been reported that Candida cells with high levels of hyphal wall protein 1 (HWP1) and C. albicans drug resistance proteins 1 and 2 (CDR1 and -2) were resistant not only to antifungal drugs, but also to phagocytes.
Clinically, there are two types of candidiasis: body surface candidiasis, including mucocutaneous candidiasis, and deep (organ) candidiasis. Surface Candidal infection is relatively easily cured, but deep candidiasis is highly resistant to antifungal drug therapy. Along with these approaches, we synthesized a short lactoferrin peptide, FKCRRWQWRM, and examined its influences on blastoconidia and phagocytes. We found that the peptide possessed superior activities in both kinds of cells, suggesting its usefulness for the treatment of candidiasis.
Lactoferrin Peptide Increases the Survival of Candida albicans- Inoculated Mice by Upregulating Neutrophil and Macrophage Functions, Especially in Combination with Amphotericin B and Granulocyte-Macrophage Colony-Stimulating Factor
Toyohiro Tanida, Fu Rao, Toshihiro Hamada, Eisaku Ueta, and Tokio Osaki
- Candida albicans infections often occur during or shortly after antibacterial treatment. Phagocytosis by polymorphonuclear neutrophil granulocytes (PMN) is the most important primarily defence mechanism against C. albicans. Certain antibiotics such as some fluoroquinolones (FQ) are known to influence phagocyte functions. Thus, we investigated the influence of older and newer FQ on the phagocytosis and killing of C. albicans by human PMN paying special attention to CD11b expression of these cells as an indicator of the degree of their activation. In order to obtain comprehensive and comparable results we tested 13 FQ over a wide range of concentrations and in a time dependent manner in a standardized approach. When used at therapeutic concentrations, the FQ tested did not influence to a clinically significant degree the phagocytosis or the killing of C. albicans by human PMN and also not their activation. However, at high concentrations those FQ with cyclopropyl-moiety at position N1 showed increase in CD11b expression and diminished phagocytosis and oxidative burst.
Influence of fluoroquinolones on phagocytosis and killing of Candida albicans by human polymorphonuclear neutrophils
Thomas Grúger; Caroline Mörler; Norbert Schnitzler; Kerstin Brandenburg; Sabine Nidermajer; Regine Horré; Josef Zúndorf
- We studied the effects of eight antibiotics, cyclosporin and corticosteroids on the in vitro secretion of GM-CSF and G-CSF by monocytes, T lymphocytes and endothelial cells. The aim was to evaluate a possible mechanism for these drugs in the delay of haemopoietic recovery after high-dose chemotherapy or bone marrow transplantation. Corticosteroids were prominent inhibitors of GM-CSF secretion by monocytes and T lymphocytes, but not by endothelial cells. In contrast, G-CSF secretion by monocytes was unchanged whereas that of endothelial cells was enhanced in the presence of corticosteroids. Cyclosporin efficiently down-regulated GM-CSF secretion by T lymphocytes and had also a minor effect on CSF secretion by endothelial cells, whereas monocyte secretion was unaffected. Stimulated T lymphocytes derived from patients under treatment with cyclosporin had impaired capacity to secrete GM-CSF compared to controls. Among the antibiotics, cephalosporins inhibited GM-CSF secretion by T lymphocytes, and GM- and G-CSF secretion by endothelial cells. Ciprofloxacin and sulphmethoxazole had minor effects on GM-CSF secretion by T lymphocytes and endothelial cells. No antibiotic significantly influenced GM-CSF secretion by monocytes.
Effects of immunosuppressive drugs and antibiotics on GM-CSF and G-CSF secretion in vitro by monocytes, T lymphocytes and endothelial cells
Lenhoff S.; Olofsson T.
- Some antimicrobial agents have been reported to modify the host immune and inflammatory responses both in vivo and in vitro. Fosfomycin (FOM) and clarithromycin (CAM) have immunomodulatory activity on human lymphocyte function. In the present study, we examined the effects of FOM and CAM on cytokine synthesis by lipopolysaccharide (LPS)- stimulated human monocytes in comparison with that of dexamethasone in vitro. The three drugs demonstrated positive or negative effects on the synthesis of various cytokines by LPS-primed monocytes. They suppressed the synthesis of tumor necrosis factor alpha, interleukin 1 alpha (IL-1 alpha), IL-1 beta, the IL-1 receptor antagonist, and granulocyte- macrophage colony-stimulating factor in a concentration-dependent manner at concentrations between 1.6 and 40 micrograms/ml. On the contrary, the drugs showed different actions on the synthesis of IL-6 and IL-10. Namely, FOM enhanced both IL-6 and IL-10 synthesis, CAM enhanced only IL-10 synthesis, but dexamethasone deeply suppressed the synthesis of both cytokines. These data indicate that antibacterial agents may modify acute-phase inflammatory responses through their effects on cytokine synthesis by monocytes.
Modulatory effect of antibiotics on cytokine production by human monocytes in vitro
K Morikawa, H Watabe, M Araake and S Morikawa
- Experimental subcutaneous Candida albicans infections in mice were used to examine the manner in which this pathogen is cleared in animals recovering from cyclophosphamide-induced leucopenia. In this system, infections at the inoculation sites progressed rapidly during a 6 day period of leucopenia to form arrays of parallel filamentous organisms that effectively isolated those in the interior from contact by neutrophils, even when the leucopenia had resolved. Dense collections of organisms also developed at sites of metastatic infection in the kidneys. A majority of the organisms were found to be viable when they were retrieved from the infected subcutaneous sites of animals that had recovered from leucopenia and whose abscesses had begun to drain spontaneously. Removal of the protective arrays of fungal cells appeared to be accomplished by drainage of abscess contents through the surface of the skin or into the collecting system of the kidney. Drainage of the subcutaneous abscesses did not occur in the cyclophosphamide-treated animals until after the neutrophilic infiltrates had developed, suggesting that this drainage process was mediated by neutrophils rather than by the organisms themselves. In summary, the above findings demonstrate that C. albicans infections in leucopenic hosts may progress to the extent that they would be very difficult to clear solely through the microbicidal processes of returning neutrophils. However, neutrophils also appear to promote the removal of masses of viable fungal cells to the exterior of the body.
Arrays of Candida albicans pseudohyphae that protect the organisms from neutrophil fungicidal mechanisms in experimental infections of mice
P.G. Sohnle; B.L. Hahn; D.K. Wagner
- Phagocytes are an essential defence against infection. Since drugs which affect their function may alter the outcome of infections, we have studied the effect of nine antibiotics on phagocyte function in vitro. The effects of antibiotics on the respiratory burst function of phagocytes from healthy adult donors were investigated using lucigenin-enhanced chemiluminescence in response to serum-opsonised zymosan. Aminoglycosides showed dose-dependent suppression of polymorphonuclear leucocyte chemiluminescence, except streptomycin which caused enhancement. Erythromycin caused profound suppression of chemiluminescence from both polymorphonuclear leucocytes and monocytes. Benzylpenicillin and the cephalosporins caused variable suppression of phagocyte chemiluminescence: cefotaxime increased monocyte chemiluminescence in some experiments. None of the drugs produced suppression at clinically relevant plasma concentrations, but erythromycin and some other drugs are preferentially concentrated in phagocytes to levels which suppress their oxidative metabolism in vitro. It is therefore possible that some antibiotics alter phagocyte function: ex vivo studies of phagocyte function in patients taking antibiotics would be valuable.
Antibiotic effects on phagocyte chemiluminescence in vitro
- A. Pierce, W. O. Tarnow-Mordi and I. A. Cree
- Drug-induced neutropenia is one of the most common causes of neutropenia. Drugs can decrease neutrophil production through toxic, idiosyncratic, or hypersensitivity mechanisms or increase peripheral neutrophil destruction through immune mechanisms. It may result from aminopyrine, propylthiouracil , penicillin, or other antibiotics. Severe dose-related neutropenia occurs predictably after cytotoxic cancer drugs or radiation therapy suppresses bone marrow production. Neutropenia due to ineffective marrow production can occur in megaloblastic anemias caused by vitamin B12 or folate deficiency. Usually, macrocytic anemia and sometimes mild thrombocytopenia develop simultaneously. Infections can cause neutropenia by impairing neutrophil production or by inducing immune destruction or rapid use of neutrophils.
The Merck Manuals Online Medical Library
- With the increasing number of immune compromised patients, fungi have emerged as major causes of human disease. Risk factors for systemic candidiasis include presence of intravascular catheters, receipt of broad-spectrum antibiotics, injury to the gastrointestinal mucosa and neutropenia. Within a species, the fungal morphotype (e.g. yeast, pseudohyphae and hyphae of Candida albicans) may be an important determinant of the host response. Whereas yeasts and spores are often effectively phagocytosed, the larger size of hyphae precludes effective ingestion.
Differentiation of CD4+ T cells along a T-helper (Th) cell type 1 (Th1) or type 2 (Th2) pathway and development of specific Th responses, is an essential determinant of the host’s susceptibility or resistance to invasive fungal infections. Development of Th1 responses is influenced by the concerted action of cytokines, such as interferon (INF)-c, interleukin (IL)-6, tumour necrosis factor (TNF)-a, and IL-12, in the relative absence of Th2 cytokines, such as IL-4 and IL-10 (Romani, 2002). Oropharyngeal candidiasis (OPC) is among the most common mycotic infections of immunocompromised patients. Development of infection depends upon both systemic and local determinants. Risk factors for oral candidiasis include extremes in age, diabetes mellitus, particularly when glycemic control is poor, nutritional deficiencies, use of broad spectrum antibiotics and immunosuppression (especially of cell-mediated immunity) (Klein et al., 1984; Guggenheimer et al., 2000). Local factors that promote infection include dentures, salivary abnormalities, treatment with inhaled steroids, and destruction of mucosal barriers with radiotherapy for head and neck cancers or cytotoxic chemotherapy. Human immunodeficiency virus (HIV) is one of the most important predisposing conditions worldwide. AIDS patients have a particularly high incidence of mucosal candidiasis, which is often recurrent and, when it involves the esophagus, can be disabling (Sangeorzan et al., 1994). Local defence mechanisms against mucosal infection include salivary proteins, such as lactoferrin, beta-defensins, histatins, lysozyme, transferrin, lactoperoxidase, mucins, and secretory immunoglobulin A. These impair adhesion and growth of Candida in the oropharyngeal cavity. Development of OPC has been associated with a salivary Th2-type cytokine profile (Leigh et al., 1998). Cell-mediated immunity plays the dominant role in prevention of candidiasis at the gastrointestinal surfaces. In AIDS, development of oropharyngeal and oesophageal candidiasis correlates with declining CD4+ lymphocyte counts. OPC is also associated with T cell immunosuppression from corticosteroid therapy, organ transplantation, cancer chemotherapy and chronic mucocutaneous candidiasis (CMC). Candida species have emerged as an important cause of bloodstream and deep tissue infections. Risk factors for Candidaemia include breakdown of mucosal barriers due to cytotoxic chemotherapy and surgical procedures, neutropenia, changes in the gut flora due to antibiotics, and invasive interventions that breach the skin, such as intravenous lines and drains (Wey et al., 1989). Common sites of dissemination include the bloodstream, kidney, liver, spleen, and endovascular structures. Quantitative and qualitative abnormalities of neutrophils and monocytes are associated with systemic candidiasis. Patients with lymphoma, leukaemia, chronic granulomatous disease, and recipients of intensive cancer chemotherapy with resultant neutropenia are at increased risk for disseminated infection. Similar to the situation with Aspergillus hyphae, the large size of Candida hyphae and pseudohyphae may preclude phagocytosis. Achieving a balance between Th1 and Th2 cytokines may be important for optimal antifungal protection while minimizing immune-mediated damage. In vivo models indicate that T regulatory cells attenuate Th1 antifungal responses, induce tolerance to the fungus and participate in the development of long lasting protective immunity after yeast priming (Montagnoli et al., 2002; Romani, 2004).
Dendritic cells play an important role in linking innate with adaptive immunity. Dendritic cells that ingest the yeast form induce differentiation of CD4+ T cells toward a Th1 pathway. In contrast, hyphae induce Th2 responses (d’Ostiani et al., 2000). Neutrophils, macrophages and natural killer (NK) cells also modulate adaptive responses to the fungus. Neutrophils differentially induce Th1 and Th2 responses depending on whether the exposure is to yeast or hyphae. The syndrome of chronic disseminated candidiasis (CDC, also known as hepatosplenic candidiasis) predominantly affects patients with haematological malignancies upon recovery from neutropenia. CDC is characterized by increased serum levels of IL-10 and local production of Th2-inducing cytokines by hepatocytes and by infected mononuclear cells (Roilides et al., 1998b; Letterio et al., 2001). Thus, although neutropenia is a major predisposing factor, the propensity for persistence of the fungus in infected tissues may be a consequence of cell-mediated immune dysregulation with suppression of Th1 and overexpression of Th2 responses.
The immune response to fungal infections
Shmuel Shoham1 and Stuart M. Levitz
Section of Infectious Diseases, Washington Hospital Center, Washington, DC, and 2Department of Medicine, Boston Medical Center and Boston University School of Medicine, Boston, MA, USA
British Journal of Haematology, 129, 569–582
- Neutrophil-mediated inflammation is terminated through the programmed cell death or apoptosis of the neutrophil, a process that can be inhibited by soluble mediators released during an inflammatory response. It has been reported, however, that the phagocytosis of intact bacteria can accelerate apoptosis. We evaluated the effects of the phagocytosis of a common nosocomial pathogen, Candida albicans, on the expression of apoptosis. Phagocytosis of killed Candida induced a dose-dependent increase in the apoptosis of normal neutrophils after 18 h of in vitro culture, from 40.7 +/- 9.1% to 81.7 +/- 4.5%, while supernatants from neutrophil: Candida co-cultures actually inhibited apoptosis. Induction of apoptosis was not dependent on phagocytosis, since opsonization of yeast with serum failed to increase apoptosis, while inhibition of phagocytosis with latrunculin B resulted in a slightly increased apoptotic rate. Increased apoptosis induced by Candida was associated with increased activity of the membrane-associated apoptotic enzyme, caspase 8, and with increased expression of the active form of the key executioner caspase, caspase 3. Increased apoptosis was associated with depletion of intracellular glutathione (GSH), and could be inhibited by the addition of exogenous GSH. These data demonstrate an important physiologic role for host-pathogen interactions in the resolution of inflammation and suggest that the response to an invading pathogen is an important stimulus to the restoration of normal immunologic homeostasis.
Phagocytosis of Candida Albicans Induces Apoptosis of Human Neutrophils
Rotstein, Dalia; Parodo, Jean; Taneja, Ravi; Marshall, John C.
- The surface of the pathogenic yeast Candida albicans is coated with phospholipomannan (PLM), a phylogenetically unique glycolipid composed of _-1,2-oligomannosides and phytoceramide. This study compared the specific contribution of PLM to the modulation of signaling pathways linked to the survival of C. albicans in macrophages in contrast to Saccharomyces cerevisiae. The addition of PLM to macrophages incubated with S. cerevisiae mimicked each of the disregulation steps observed with C. albicans and promoted the survival of S. cerevisiae. Externalization of membranous phosphatidylserine, loss of mitochondrial integrity, and DNA fragmentation induced by PLM showed that this molecule promoted yeast survival by inducing host cell death. These findings suggest strongly that PLM is a virulence attribute of C. albicans and that elucidation of the relationship between structure and apoptotic activity is an innovative field of research.
The yeast C. albicans has been reported to inhibit tumor necrosis factor-_-induced DNA fragmentation in macrophages (33) and to induce apoptosis of macrophages (24) and neutrophils. Macrophages undergo apoptotic cell death after infection with C. albicans strains capable of hyphal formation (24), and activation of caspase 3 has been observed after endocytosis of C. albicans by neutrophils. Extensive literature exists on the effects of surface glycolipids from pathogens on the control of host cell apoptosis.
Candida albicans Phospholipomannan Promotes Survival of Phagocytosed Yeasts through Modulation of Bad Phosphorylation and Macrophage Apoptosis
Stella Ibata-Ombetta, Thierry Idziorek¶, Pierre-Andre´ Trinel, Daniel Poulain, and Thierry Jouault
- Interactions of microorganisms with integrins are central to the host defense mechanisms. The leukocyte integrin CD11b/CD18 is the principal adhesion receptor on leukocytes for Candida albicans, a major opportunistic pathogen. In this study we have investigated the roles of three regions within the receptor, the inserted (I) and lectin-like domains within the CD11b subunit, and the CD18 subunit, in CD11b/CD18-C. albicans interactions. We report four major findings. 1) A mutation in CD18 exerts a dominant negative effect on the function of the CD11b/CD18 complex. This interpretation is based on the observation that in the absence of CD18, the CD11b subunit alone binds C. albicans well, but a single point mutation at Ser138 of CD18 abolishes CD11b/CD18 binding of the fungus. 2) The lectin-like domain is not sufficient for CD11b/CD18-C. albicans interactions. Rather, the lectin-like domain appears to influence CD11b/CD18 binding activity by modulating the function of the I domain. 3) The I domain is the primary binding site for C. albicans in the receptor and is sufficient to support an efficient interaction. 4) We have identified specific amino acid sequences within the I domain that engage the microorganism. Compared with other ligands of CD11b/CD18, C. albicans has some unique as well as common contact sites within the I domain of the receptor. Such unique contact sites may underlie the ability of C. albicans to modulate CD11b/CD18 function and raise the possibility for selective interference of the microorganism-host leukocyte interactions.
Interaction of the Fungal Pathogen Candida albicans with Integrin CD11b/CD18: Recognition by the I Domain Is Modulated by the Lectin-Like Domain and the CD18 Subunit
Christopher B. Forsyth, Edward F. Plow and Li Zhang
- Studies of host-parasite relationships at the cellular level, using Candida albicans and rabbit alveolar macrophages or guinea pig neutrophils are presented. Guinea pig neutrophils killed the intracellular Candida cells presumed by myeloperoxidase-halide-hydrogen peroxide system. In contrast, rabbit alveolar macrophages did not kill the intracellular Candida cells although their phagocytic rate was almost comparable to that of neutrophils. Phagocytizing macrophages were eventually destroyed by the intracellular proliferation of Candida cells and formation of germ tubes and pseudomycelia. No significant improvement of Candidacidal activity was observed with macrophages from normal and immunized rabbits in immune serum. The mode of phagocytosis by macrophages and neutrophils were also studied under the scanning electron microscope.
Phagocytosis of Candida albicans by rabbit alveolar macrophages and guinea pig neutrophils.
- We examined the in vitro effect of Candida albicans on NO production by macrophages. Candida albicans suppressed not only NO production but also expression of inducible NO synthase (iNOS) mRNA by murine IFN-γ and bacterial LPS-stimulated peritoneal macrophages. The suppression was not associated with inhibition but rather stimulation of IL-1β production. This effect was observed when more than 1 × 103/ml of Candida albicans were added to macrophage cultures (1 × 106 cells/ml) and reached a maximal level at 1 × 106/ml. The NO inhibitory effect of Candida albicans was mediated predominantly by as yet unidentified soluble factor(s) and to a lesser extent by direct contact. In addition, heat- or paraformaldehyde-killed Candida albicans did not show this inhibitory activity. Culture supernatant of Candida albicans also inhibited NO production by activated macrophages in a dose-dependent manner, and increased IL-1β production. Finally, the inhibitory effect was not mediated by IL-10 and transforming growth factor- Candida albicans beta (TGF-β), since neutralizing antibodies to these cytokines did not influence -induced reduction in macrophage NO production. Our results suggest that Candida albicans may evade host defence mechanism(s) through a soluble factor-mediated suppression of NO production by stimulated macrophages, and that the effect is independent of production of immunosuppressive cytokines such as IL-10 and TGF-β. Candida albicans, an ubiquitous fungal microorganism, forms part of the normal microflora in the gastrointestinal tract and vagina even in individuals who do not have an apparent immunological dysfunction, suggesting the presence of certain mechanisms that evade the host defence system against this pathogen. It has been well documented that the host defence mechanism against mucosal infection with Candida albicans is mediated mainly by cellular immunity. In experimental models, protection against candidiasis is closely associated with the synthesis of IL-12 and induction of Th1 cells. Neutralization of endogenously synthesized IFN-γ and IL-12 by specific antibodies prevents the development of protective Th1 responses and exacerbates infections with Candida albicans. A number of macrophage-derived cytokines, including IL-10 and TGF-β, exert anti-inflammatory effects by inhibiting the production of proinflammatory cytokines, such as IL-1, IL-6, and tumour necrosis factor-alpha (TNF-α). In addition, these cytokines also suppress the production of NO by IFN-γ-stimulated macrophages.
Several investigators have demonstrated alterations by Candida albicans of the fungicidal activity of phagocytic cells. For example, Hilger & Danley and Danley et al. indicated that live Candida albicans suppressed the release of H2O2 by neutrophils, while dead organisms did not. On the other hand, Smail et al. demonstrated that Candida albicans produced a crude hyphal inhibitory product which inhibited superoxide anion production and release of azurophilic and specific granule components by activated neutrophils. Furthermore, Diamond et al. demonstrated that Candida albicans released small peptides which inhibited adhesion of the fungus and neutrophils. Other investigations also described the suppressive effects of Candida albicans on both cellular and humoral immunity. Considered together, these observations demonstrate the presence of several mechanisms by which Candida albicans evades the host defence systems and multiplies in host tissue. The present results may describe a new mechanism that allows Candida albicans to resist macrophage fungicidal activity.
Candida albicans suppresses nitric oxide (NO) production by interferon-gamma (IFN-γ) and lipopolysaccharide (LPS)-stimulated murine peritoneal macrophages
T Chinen, M H Qureshi, Y Koguchi, and K Kawakami
- 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.
Production and function of cytokines in natural and acquired immunity to Candida albicans infection.
- This study was designed to test a) whether carbohydrates other than glucose decreased the phagocytic capacity of neutrophils in normal human subjects, b) the duration of this effect, and c) the effect of fasting on neutrophilic phagocytosis. Venous blood was drawn from the arm after an overnight fast and at 0.5, 1, 2, 3, or 5 hr postprandial and this was incubated with a suspension of Staphylococcus epidermidis. The phagocytic index (mean number of bacteria viewed within each neutrophil) was determined by microscopic examination of slides prepared with Wright’s stain. Oral 100-g portions of carbohydrate from glucose, fructose, sucrose, honey, or orange juice all significantly decreased the capacity of neutrophils to engulf bacteria as measured by the slide technique. Starch ingestion did not have this effect. The decrease in phagocytic index was rapid following the ingestion of simple carbohydrates. The greatest effects occurred between 1 and 2 hr postprandial, but the values were still significantly below the fasting control values 5 hr after feeding (P < 0.001). The decreased phagocytic index was not significantly associated with the number of neutrophils. These data suggest that the function and not the number of phagocytes was altered by ingestion of sugars. This implicates glucose and other simple carbohydrates in the control of phagocytosis and shows that the effects last for at least 5 hr. On the other hand, a fast of 36 or 60 hr significantly increased (P < 0.001) the phagocytic index.
Role of sugars in human neutrophilic phagocytosis
Albert Sanchez, J. L. Reeser, H. S. Lau, P. Y. Yahiku, R. E. Willard, P. J. McMillan, S. Y. Cho, A. R. Magie, and U. D. Register
- The innate immune response was once considered to be a limited set of responses that aimed to contain an infection by primitive ‘ingest and kill’ mechanisms, giving the host time to mount a specific humoral and cellular immune response. In the mid-1990s, however, the discovery of Toll-like receptors heralded a revolution in our understanding of how microorganisms are recognized by the innate immune system, and how this system is activated. Several major classes of pathogen-recognition receptors have now been described, each with specific abilities to recognize conserved bacterial structures.
The first receptor on the surface of macrophages to be described as a mannan receptor was the C‑type-lectin mannose receptor (MR). By contrast, recognition of the shorter linear structures of O‑bound mannan is performed by TLR4, and results in cytokine production. Interestingly, TLR4 stimulation is lost during the germination of yeast into hyphae, which leads to a loss of interferon-γ (IFNγ) production capacity. The lectin domain mediates recognition of both the yeast and hyphal forms of C. albicans, as well as several other fungi. Recognition by CR3 does not trigger protective host responses, such as the respiratory burst, and can repress pro-inflammatory signals. Dectin 1 is a myeloid-expressed transmembrane receptor and possesses a single extracellular, nonclassical C–type-lectin-like domain that specifically recognizes β-(1,3)-glucans. Dectin 1 can recognize several fungi, including C. albicans yeast, although it does not appear to recognize C. albicans hyphae. In addition to mannoproteins, mannans and β-glucans, other structures of C. albicans can also be recognized as fungal PAMPs. Recently, it has been demonstrated that chitin induces recruitment of immune cells that mainly release IL–4 and IL-13.
More is known about the receptors that are involved in the induction of cytokine production by C. albicans. At least four TLRs (TLR2, TLR4, TLR6 and TLR9) are involved in triggering these responses. Moreover, TLR2 ligands fail to induce
the release of IL-12 and TH1-type IFNγ, thus promoting conditions that are favourable for TH2- or regulatory T cell (TReg)-type responses. C. albicans induces immunosuppression through TLR2-mediated IL–10 release, and this leads to the generation of CD4+CD25+ TReg cells with immunosuppressive potential. In addition to inducing direct anti-inflammatory effects, C. albicans has developed strategies to either block or avoid recognition by stimulatory PRRs.
The challenge ahead is to understand the level of complexity that underlies the response that is triggered by pathogen recognition. In this Review, we use the fungal pathogen Candida albicans as a model for the complex interaction that exists between the host pattern-recognition systems and invading microbial pathogens.
An integrated model of the recognition of Candida albicans by the innate immune system
Mihai G. Netea, Gordon D. Brown, Bart Jan Kullberg & Neil A. R. Gow
- In a paper published June 30 in the online version of Hypertension, Frank DeLano, a research scientist at UC San Diego, and Geert Schmid-Schönbein, a professor of bioengineering, describe how they successfully reversed the SHR animals’ symptoms of high blood pressure, a pre-diabetes condition called insulin resistance, and immune suppression. H. Glenn Bohlen, a professor in the Department of Cellular and Integrative Physiology at Indiana University Medical School, wrote in an accompanying editorial in Hypertension that the new study will likely be important to people suffering from obesity as well as hypertension. “With the national and international emphasis on obesity and its attendant cardiovascular problems, there is a tendency to forget that essential hypertension affects about the same percentage of humans as does serious obesity and an even higher percentage of the population than does type 2 diabetes mellitus,” wrote Bohlen. “The elegant study by Delano and Schmid-Schönbein points to a potentially very important overlap of an insulin resistance mechanism with hypertension in the spontaneously hypertensive rat (SHR).”The researchers showed that the SHR animals have protease activity in their circulation that cleaves more than just insulin receptors. In these animals, proteases also cleave significant numbers of CD18, an important binding receptor on the surface of infection-fighting leukocytes. CD18 gives these cells the ability to adhere to the walls of blood vessels as a way to home in on infections. With the loss of CD18 receptors, leukocytes of the SHR animals are unable to bind to the wall of blood vessels, resulting in a compromised immune system. “These results point to a single mechanism that explains multiple and diverse cell dysfunctions encountered in hypertensive rats, and they also suggest that a similar mechanism may be operating in humans suffering simultaneously from hypertension, diabetes, and other metabolic conditions,” said Schmid-Schönbein The team went on to test whether administration of a protease-blocking drug could reverse the multiple metabolic complications in the rat strain. They administered doxycycline, a seemingly unlikely Candidate to have such a beneficial effect. Infectious disease specialists often prescribe doxycycline, an antibiotic, to counter bacterial infections. However, in laboratory tests doxycycline also blocks the activity of certain proteases in the SHR strain of rat. The researchers found that protein receptors on the surface of SHR cells become clipped off as the animals develop hypertension. They used a novel visualization technique to show that after several weeks of ingesting doxycycline in their drinking water, the SHR rats developed cells that again bristled with normal CD18 and insulin receptors. The animals’ metabolic conditions simultaneously improved; blood pressure normalized and symptoms of immune suppression disappeared. Hypertension, insulin resistance and immune suppression links
Frank DeLano, a research scientist at UC San Diego, and Geert Schmid-Schönbein
- Chronic inflammation rather than invasion is characteristic of some forms of superficial candidiasis such as denture stomatitis. We hypothesized that Candida albicans may play a critical role in the pathogenesis of inflammatory lesions observed in chronic candidiasis by activating the proinflammatory cytokine interleukin-1β (IL-1β) from epithelial stores of the precursor. The aim of this study was therefore to demonstrate the proteolytic cleavage and activation of the inactive precursor of IL-1β (pro-IL-1β) by C. albicans. After incubation of either blastospores or hyphae with the inactive precursor, proteolytic cleavage was monitored by sodium dodecyl sulfate-polyacrylamide gel electrophoresis Western immunoblotting analysis, and the biological activity of the cleavage products was tested in a bioassay. We report here that late-stationary-growth-phase blastospores as well as hyphae of C. albicans, but not exponentially growing cells, can efficiently cleave pro-IL-1β to yield fragments of molecular masses compatible with mature biologically active IL-1β (17 to 19 kDa). Assays conducted in the presence of selected proteinase inhibitors suggest that the cleavage of pro-IL-1β involves the participation of one or more aspartyl proteinases. Cleavage products showed a dose-dependent IL-1β-like activity in a thymocyte proliferation bioassay, which was inhibited by anti-IL-1β neutralizing antibodies. The present data thus suggest a role for C. albicans proteinases in the activation and maintenance of the inflammatory response at epithelial surfaces.
Proteolytic Activation of the Interleukin-1β Precursor by Candida albicans
Beauséjour, Annie, Grenier, Daniel, Goulet, Jean-Paul
- This study examined CD11/CD18-mediated adhesion in neutrophil emigration during acute and recurrent Pseudomonas aeruginosa-induced pneumonia. Neutrophil emigration during acute pneumonia was studied in anti-CD18 antibody or murine-IgG-pretreated rabbits 4 hours after intrabronchial instillation of P. aeruginosa. To examine emigration in recurrent pneumonias, rabbits given P. aeruginosa on day 0 received anti-CD18 antibody or IgG on day 7. A second instillate was placed either at the initial site or in a separate lobe, and emigration into alveolar spaces was quantitated morphometrically after 4 hours. The results show that CD11/CD18 was required for neutrophil emigration in acute pneumonias and in recurrent pneumonias that occurred at a site distant from the initial infection. However, when the recurrent pneumonia occurred in the previously inflamed site, CD11/CD18 was not required. When the same number of organisms were instilled on days 0 and 7, emigration was reduced to 15 to 20 percent of the number that migrated initially and only CD18-independent adhesion pathways were used. Increasing the concentration of organisms threefold increased emigration through both CD18-dependent and CD18-independent pathways. These data indicate that P. aeruginosa induces CD11/CD18-dependent emigration during acute pneumonia and recurrent pneumonia at previously uninflamed sites. However, adhesion pathways are altered in regions of chronic inflammation, and a greater proportion of neutrophil emigration occurs through CD11/CD18-independent pathways.
Role of CD 11/CD 18 in neutrophil emigration during acute and recurrent Pseudomonas aeruginosa-induced pneumonia in rabbits
T Kumasaka, NA Doyle, WM Quinlan, L Graham and CM Doerschuk
- Some secreted proteins do not remain cell associated but forage into the extracellular environment. If such proteins are hydrolytic enzymes, they have capacity to hydrolyze large or complex substrates into small units that can be transported into the cell as a source of nutrition. If the degradation of host targets facilitates colonization or invasion, then such enzymes also function as virulence factors. In the last few years, several studies have examined clinical isolates for one or more phospholipase, proteinase, or hemolytic activities. These studies demonstrate the production of these activities by some but not all isolates. There are differences based on the site of isolation or the presence of type 2 diabetes mellitus. One of the characteristics of most of the hydrolytic activities is that there are multiple enzymes that may be expressed under different conditions.
Candida albicans Cell Wall Proteins
- LaJean Chaffin
Microbiology and Molecular Biology Reviews, September 2008, p. 495-544, Vol. 72, No. 3,1092-2172
- The study indicates that host age is a determining factor in yeast carriage. From the neonatal period, humans go through several dentition periods, and the emergence and substitution of teeth and changes in living habits greatly change the environment of the oral cavity, and therefore influence colonization by oral commensal organisms, certainly
including Candida spp. Russell and Lay showed that the frequency of oral yeast carriage at birth was low, doubled by the time that infants were discharged from the hospital at about seven-day old, and increased sharply after one month old. Kleinegger et al. demonstrated that the frequency of oral yeast carriage was 44% of the examined individuals in a group aged from 0.5 to 1.5, 24% in a 5-7-year-old group, 40% in a 15- to 18-year-old group.
In a review by Odd in 1988, the highest reported frequencies were 71% of school children in the United Kingdom, and 56% of children in Israel. Different sampling and identification methods for Candida spp. would also certainly influence the results. Because of the uneven distribution of C. albicans throughout the oral cavity, swab samples can yield false-negative culture more often than oral rinse samples or imprint culture. The number of false-negative cultures was reduced by the swabbing of two sitesrather than one. In conclusion, our observations indicate that there is an increased risk of dental caries with C. albicans carriage rates in preschool and school children.
Oral Candida albicans carriage in healthy preschool and school children
Rokiewicz D, Daniluk T, Zaremba ML2, Cylwik-Rokicka D , Stokowska W,
Pawińska M, Dąbrowska E, Marczuk-Kolada G, Waszkiel D
- The prevalence of opportunistic fungal infections has increased dramatically among the aged population in recent years. This work investigated the effect of ageing on murine defences against Candida albicans. Aged C57BL/6 mice that were experimentally infected intravenously had a significantly impaired survival and a higher tissue fungal burden compared with young mice. In vitro production of tumour necrosis factor (TNF)- by macrophages from aged mice in response to yeast cells and hyphae of C. albicans was significantly lower than production by macrophages from young mice. In vitro production of cytokines, such as TNF- and gamma interferon (IFN-), by antigen-stimulated splenocytes from mice intravenously infected with C. albicans cells was also diminished in old mice. This decrease in production of T helper 1 cytokines in old mice correlated with a diminished frequency of IFN–producing CD4+ T lymphocytes, although the ability to develop an acquired resistance upon vaccination (primary sublethal infection) of mice with the low-virulence PCA2 strain was not affected in aged mice. The diversity of antigens recognized by C. albicans-specific antibodies in sera from infected aged mice was clearly diminished when compared with that from infected young mice. Taken together, these data show that aged mice develop an altered innate and adaptive immune response to C. albicans and are more susceptible to systemic primary candidiasis.
Impaired immune response to Candida albicans in aged mice
Celia Murciano, Eva Villamón, Alberto Yáñez, José-Enrique O’Connor, Daniel Gozalbo and M. Luisa Gil
- Previous work by our group showed that aged C57BL/6 mice develop an altered innate and adaptive immune response to Candida albicans and are more susceptible to systemic primary candidiasis. In this work, we used young (2-3 months old) and aged (18-20 months old) C57BL/6 mice to study in vitro the influence of aging on (1) the fungicidal activity of neutrophils and macrophages, (2) the production of cytokines by resident peritoneal macrophages in response to C. albicans, and (3) cell surface Toll-like receptor (TLR) 2 expression on resident peritoneal macrophages. Our results indicate that murine phagocytes have a fungicidal activity well preserved with aging. In vitro production of proinflammatory cytokines (IL-6, IL-1beta, and tumor necrosis factor-alpha and chemokines (MIP-2) by purified (CD11b(+)) peritoneal macrophages in response to yeasts and hyphae of C. albicans was significantly lower in aged mice as compared with young mice. However, the production of IL-10 by macrophages, in response to C. albicans, was similar in both young and aged animals. Moreover, baseline TLR2 surface expression level was lower on aged macrophages than on control macrophages. Taken together, these data indicate that the increased susceptibility to C. albicans disseminated infections in aged mice is correlated with defects in TLR2 expression and in cytokine production, but not with an impaired fungicidal activity.
Influence of aging on murine neutrophil and macrophage function against Candida albicans.
Murciano Celia; Yáñez Alberto; O’Connor José E; Gozalbo Daniel; Gil María Luisa
- The pathogenic yeast Candida albicans and its derived molecules stimulate a wide range of macrophage secretory functions and may adapt to escape being killed by this phagocyte. In this study, phagocytosis of C. albicans and of the nonpathogenic yeast Saccharomyces cerevisiae was shown to be associated with phosphorylation of the mitogen-activated protein kinase (MAPK)/extracellularly regulated kinase (ERK) pathway in the absence of significant activation of either p38MAPK or stress-activated protein kinase/c-Jun N-terminal kinase. However, although 80% of endocytosed C. albicans survived after 1 h, 80% of S. cerevisiae cells were killed. Considerable quantitative differences were observed between the two species in the sequential phosphorylation of MAPK/ERK kinase (MEK), extracellularly regulated kinase-1, and 90-kDa-ribosomal S6 kinases. A lower level of activation of the pathway by C. albicans was associated with a species-specific overexpression of the MEK phosphatase MAPK phosphatase (MKP)-1. Killing of both C. albicans and S. cerevisiae could be reduced using PD98059, which mimics MKP-1 and inhibits MEK phosphorylation, suggesting that specific MKP-1 activation by C. albicans could contribute to its ability to escape the yeast lytic potential of macrophages.
Role of extracellular signal-regulated protein kinase cascade in macrophage killing of Candida albicans
Stella Ibata-Ombetta, Thierry Jouault, Pierre-André Trinel and Daniel Poulain
- Candida albicans is the most common opportunistic fungal pathogen of humans, causing systemic disease in immunocompromised patients. Host resistance to C. albicans infections is mediated predominantly by neutrophils and monocytes/macrophages. We have previously shown that exposure of a human epithelial cell line (HEp2) to C. albicans or to a culture filtrate of C. albicans caused actin rearrangement in the HEp2 cells. Since shifting of actin from the filamentous to the globular form may be crucial to the activity of phagocytes, we assessed in the present study the effect of the C. albicans metabolite (lyophilized culture filtrate) on the cytoskeleton of murine peritoneal macrophages and on their phagocytic activity. Our results showed a significant decrease in phagocytosis of C. albicans, ranging from 53-63% and a 25% reduction for C. glabrata cells. Using confocal laser scanning microscopy an actin rearrangement in the macrophages could be demonstrated that may be associated with the decrease of phagocytosis. We also tested the effect of mannan and of the secreted aspartic proteinase (Sap) inhibitor–pepstatin, on the activity of the metabolite in order to define the putative component and found no influence. In conclusion, our data indicate that a C. albicans metabolite affects phagocytic activity of macrophages, probably by alterations in their cytoskeleton.
Candida albicans metabolite affects the cytoskeleton and phagocytic activity of murine macrophages.
- Macrophages are widely distributed immune system cells that play an indispensable role in homeostasis and defense. They can be phenotypically polarized by the microenvironment to mount specific functional programs. Polarized macrophages can be broadly classified in two main groups: classically activated macrophages (or M1), whose prototypical activating stimuli are IFNgamma and LPS, and alternatively activated macrophages (or M2), further subdivided in M2a (after exposure to IL-4 or IL-13), M2b (immune complexes in combination with IL-1beta or LPS) and M2c (IL-10, TGFbeta or glucocorticoids). M1 exhibit potent microbicidal properties and promote strong IL-12-mediated Th1 responses, whilst M2 support Th2-associated effector functions. Beyond infection M2 polarized macrophages play a role in resolution of inflammation through high endocytic clearance capacities and trophic factor synthesis, accompanied by reduced pro-inflammatory cytokine secretion. Similar functions are also exerted by tumor-associated macrophages (TAM), which also display an alternative-like activation phenotype and play a detrimental pro-tumoral role. Here we review the main functions of polarized macrophages and discuss the perspectives of this field.
Macrophage activation and polarization.
- Macrophages are cells that function as a first line of defence against invading microorganisms. One of the hallmarks of macrophages is their ability to become activated in response to exogenous ‘danger signals’. Most microbes have molecular patterns (PAMPS) that are recognized by macrophages and trigger this activation response. There are many aspects of the activation response to PAMPS that are recapitulated when macrophages encounter endogenous danger signals. In response to damaged or stressed self, macrophages undergo physiological changes that include the initiation of signal transduction cascades from germline-encoded receptors, resulting in the elaboration of chemokines, cytokines and toxic mediators. This response to endogenous mediators can enhance inflammation, and thereby contribute to autoimmune pathologies. Often the overall inflammatory response is the result of cooperative activation signals from both exogenous and endogenous signals. Macrophage activation plays a critical role, not only in the initiation of the inflammatory response but also in the resolution of this response. The clearance of granulocytes and the elaboration of anti-inflammatory mediators by macrophages contribute to the dissolution of the inflammatory response. Thus, macrophages are a key player in the initiation, propagation and resolution of inflammation. This review summarizes our understanding of the role of macrophages in inflammation. We pay particular attention to the endogenous danger signals that macrophages may encounter and the responses that these signals induce. The molecular mechanisms responsible for these responses and the diseases that result from inappropriately controlled macrophage activation are also examined.
Macrophage activation by endogenous danger signals.
- Phagocytosis is the first step of defense against infections from the innate immune system, as it is the process of internalization of pathogens by cells with phagocytic activity, such as macrophages, which is followed by pathogen killing and destruction. Thus, phagocytosis assays are used as assays for one function of the innate immune system. As fungal infections are of increasing relevance and phagocytic mechanisms are dependent on the pathogenic organism and its viability, we established a microtiter plate phagocytosis assay based on viable, fluorescence-labeled Candida albicans. The distinction between internalized yeast cells and cells attached to macrophages was done via quenching of FITC—fluorescence by trypan blue, and the remaining fluorescence was quantified and used as indicator of the phagocytosis efficiency. As a proof of principle we showed that compounds acting on the dynamics of the actin cytoskeleton of the macrophages reduced the phagocytosis efficiency in a concentration dependent manner.
Phagocytosis Assay Based on Living Candida albicans for the Detection of Effects of Chemicals on Macrophage Function
Nina Klippel; Ursula Bilitewski
- Production of reactive oxygen species (ROS) is an important aspect of phagocyte-mediated host responses. Since phagocytes play a crucial role in the host response to Candida albicans, we examined the ability of Candida to modulate phagocyte ROS production. ROS production was measured in the murine macrophage cell line J774 and in primary phagocytes using luminol-enhanced chemiluminescence. J774 cells, murine polymorphonuclear leukocytes (PMN), human monocytes, and human PMN treated with live C. albicans produced significantly less ROS than phagocytes treated with heat-killed C. albicans. Live C. albicans also suppressed ROS production in murine bone marrow-derived macrophages from C57BL/6 mice, but not from BALB/c mice. Live C. albicans also suppressed ROS in response to external stimuli. C. albicans and Candida glabrata suppressed ROS production by phagocytes, whereas Saccharomyces cerevisiae stimulated ROS production. The cell wall is the initial point of contact between Candida and phagocytes, but isolated cell walls from both heat-killed and live C. albicans stimulated ROS production. Heat-killed C. albicans has increased surface exposure of 1,3-beta-glucan, a cell wall component that can stimulate phagocytes. To determine whether surface 1,3-beta-glucan exposure accounted for the difference in ROS production, live C. albicans cells were treated with a sublethal dose of caspofungin to increase surface 1,3-beta-glucan exposure. Caspofungin-treated C. albicans was fully able to suppress ROS production, indicating that suppression of ROS overrides stimulatory signals from 1,3-beta-glucan. These studies indicate that live C. albicans actively suppresses ROS production in phagocytes in vitro, which may represent an important immune evasion mechanism.
Live Candida albicans suppresses production of reactive oxygen species in phagocytes.
- The two-stage neuroinflammatory process, containment and progression, proposed to underlie neurodegeneration may predicate on systemic inflammation arising from the gastrointestinal tract. Helicobacter infection has been described as one switch in the pathogenic-circuitry of idiopathic parkinsonism (IP): eradication modifies disease progression and marked deterioration accompanies eradication-failure. Moreover, serum Helicobacter-antibody-profile predicts presence, severity and progression of IP. Slow gastrointestinal-transit precedes IP-diagnosis and becomes increasingly-apparent after, predisposing to small-intestinal bacterial-overgrowth (SIBO). Although IP is well-described as a systemic illness with a long prodrome, there has been no comprehensive overview of the blood profile. Here, it is examined in relation to Helicobacter status and lactulose-hydrogen-breath-testing for SIBO. A robust finding of reduced lymphocyte count in 126 IP-probands and 79 spouses (without clinically-definite IP), compared with that in 381 controls (p<0.001 in each case), was not explained by Helicobacter-status or breath-hydrogen. This complements a previous report that spouses were ‘down-the-pathway’ to ‘clinically-definite’ disease. In 205 other controls without clinically-definite IP, there were strong associations between sporadic cardinal features and immunoglobulin class concentration, not explained by Helicobacter-status. Premonitory states for idiopathic parkinsonism associated with relative lymphopenia, higher serum immunoglobulin concentrations and evidence of enteric-nervous-system damage may prove viral in origin. Although only 8% of the above 79 spouses were urea-breath-test-positive for Helicobacter, all 8 spouses with clinically-definite IP were (P<0.0001). Transmission of a ‘primer’ to a Helicobacter-colonised recipient might result in progression to the diagnostic threshold. Twenty-five percent of the 126 probands were seropositive for anti-nuclear autoantibody. In 20 probands, monitored before and serially after anti-Helicobacter therapy, seropositivity marked a severe hypokinetic response (p=0.03). It may alert to continuing infection, even at low-density. Hyperhomocysteinemia is a risk factor for dementia and depression. Serum homocysteine exceeded the target in 43% of the 126 IP-probands. It was partially explained by serum B12 (12% variance, p<0.001), but not by Helicobacter-status (gastric-atrophy uncommon in IP) or levodopa treatment. Immune-inflammatory activation increases homocysteine production. Since an estimated 60% of probands are hydrogen-breath-test positive, SIBO, with its increased bacterial utilisation of B12, is a likely cause. Thus, two prognostic indicators in established IP fit with involvement of Helicobacter and SIBO.
Blood profile holds clues to role of infection in a premonitory state for idiopathic parkinsonism and of gastrointestinal infection in established disease.
Andre Charlett, R John Dobbs, Sylvia M Dobbs, Clive Weller, Mohammad A A Ibrahim, Tracy Dew, Roy Sherwood, Norman L Oxlade, J Malcolm Plant, James Bowthorpe, Andrew J Lawson, Alan Curry, Dale W Peterson and Ingvar T Bjarnason
- Chronic inflammatory diseases like rheumatoid arthritis, inflammatory bowel disease, psoriasis, and liver disease cause “sickness behaviors,” including fatigue, malaise, and loss of social interest. However, it has been unclear how inflammation in other organs in the body can impact the brain and behavior. The researchers found that in mice with inflamed livers, white blood cells called monocytes infiltrated the brain. These findings support previous research demonstrating the presence of immune cells in the brain following organ inflammation, challenging the long-held belief that the blood-brain barrier prevents immune cells from accessing the brain.
Liver inflammation also stimulated cells in the blood to make an immune chemical (TNFα). When the researchers blocked the signaling of this immune chemical, microglia produced less CCL2, and monocytes stayed out of the brain. In the mice with inflamed livers, preventing the entry of monocytes into the brain reduced sickness behaviors; mice showed more mobility and social interaction. These findings suggest that people with chronic inflammatory diseases may benefit from treatments that limit monocyte access to the brain. “Sickness behavior significantly impacts quality of life. Our findings further our understanding and may generate potential new avenues for treatment of these often crippling symptoms,” said Swain. “The brain is the master coordinator of many of our bodies’ defense responses, so it must be able to sense injury and inflammation in distant body organs. This study starts to explain the peripheral communication signals that activate the brain,” said Nancy Rothwell, PhD, DSc, at the University of Manchester, an expert on brain inflammation who is unaffiliated with the study.
How Inflammatory Disease Causes Fatigue.
Mark Swain, MD
- Matrix metalloproteinases (MMPs) are zinc-dependent endopeptidases that are a subfamily of metzincins. Matrix metalloproteinases are responsible for much of the turnover of extracellular matrix components and are key to a wide range of processes including tissue remodeling and release of biological factors. Imbalance between the MMPs and endogenous tissue inhibitors of metalloproteinases (TIMPs) can result in dysregulation of many biologic processes and lead to the development of malignancy, cardiovascular disease, and autoimmune and inflammatory disorders. MMP production by monocyte/macrophages is dependent on the cell type, state of differentiation, and/or level of activation and whether they are infected, e.g., by HIV-1. MMP expression by
HIV-1 infected monocytes and macrophages may alter cellular trafficking and contribute to HIVassociated pathology such as HIV-associated dementia (HAD). This review will provide a classification of the MMP super-family with particular reference to those produced by monocyte/macrophages, describe their regulation and function within the immune system, and indicate their possible roles in the pathogenesis of disease, including
Dysregulation in the levels and control of MMPs can lead to pathological processes (including tumor growth and migration, arthritis, cirrhosis, aortic aneurysms, and fibrosis) and diseases (such as glaucoma, lupus, scleroderma, multiple sclerosis, and HIV-1 associated dementia). MMPs play an important role in immunological functions including ECM for leukocyte migration, modulating chemokine, and cytokine activity through both their activation and inactivation and defensins activation.
MMPs regulate numerous biologic processes through their proteolytic function, in both normal and pathological states. They alter the cellular milieu and cell behavior through proteolytic turnover of matrix components, by releasing molecules expressed on the surface of cells and by cleaving cell surface receptors or cell-cell adhesion proteins.
Cells of the monocyte/macrophage lineage, including blood monocytes, dendritic cells, and tissue macrophages such as microglia secrete diverse MMPs in large quantities.
Induction of MMPs can modulate DC functions including DC migration through endothelial barriers. Cell migration is critical for dendritic cells (DCs) in the initiation of the immune response.
HIV-infected patients frequently report gingival inflammation and may develop progressive periodontal tissue breakdown or require dental extraction due to weakened attachment. HIV-related periodontal disease, including gingivitis; periodontitis; and bacterial, viral and fungal infections has been associated with increased levels of MMP-1, -3, and -8 in saliva from HIV-infected individuals, and these MMPs may play a role in the development of HIV-associated periodontitis.
Matrix metalloproteinases, their production by monocytes and macrophages and their potential role in HIV-related diseases
Nicole L. Webster and Suzanne M. Crowe
- A variety of morphological changes in the basement membrane (BM) are known to occur in inflammatory diseases. Modifications of the BM can be associated with significant changes in protein content. Candida albicans (C. albicans) is normally a commensal organism and is a member of the natural flora of a large number of healthy individuals. However, under certain conditions, C. albicans can invade host tissues, causing inflammation and tissue damage. The aim of this study was to investigate the effect of C. albicans on the expression and production of structural (laminin-5 and type IV collagen) and inflammatory [matrix metalloproteinases (MMPs) and their inhibitors] proteins by human oral epithelial cells. Using engineered normal human oral mucosa infected with 10(5) C. albicans/cm2 for different periods of time, we were able to demonstrate that this yeast promotes significant laminin-5 and type IV collagen gene activation and protein secretion. These effects were accompanied by MMP-2 and MMP-9 gene activation. Interestingly, only the levels of active MMP-9 rose. The increase in MMP levels was paralleled by a decrease in the secretion of type 2 matrix metalloproteinase tissue inhibitors (TIMP-2). Our results demonstrated that C. albicans has a significant effect on tissue structure through BM protein and MMP modulation. This might help C. albicans overcome the mechanical and biological defenses of the tissue and allow it to disseminate, causing severe infections. If C. albicans uses MMPs (mainly MMP-9) to disseminate, inhibition of this protease could be of interest in treating a variety of inflammatory disorders, including oral candidiasis.
Basement membrane protein and matrix metalloproteinase deregulation in engineered human oral mucosa following infection with Candida albicans.
- Dendritic cells (DC) function as professional phagocytes to kill Candida albicans and subsequently present it to the adaptive immune system. Monocytes, macrophages and DC were generated from five individual donors and their Candida-killing capacity and cytokine release were assessed. Compared to monocytes and macrophages, DC from healthy volunteers were significantly less effective in C. albicans-stimulated cytokine release, killing of C. albicans blastoconidia and damaging of C. albicans hyphae. In conclusion, while important as antigen-presenting cells and initiators of the adaptive immune system, DC are poor in both intracellular killing and damaging of C. albicans hyphae. Effective handling of large numbers of C. albicans is the prime task of the innate immune system consisting of large numbers of neutrophils and monocytes.
Human dendritic cells are less potent at killing Candida albicans than both monocytes and macrophages
NETEA Mihai G.; GIJZEN Karlijn; COOLEN Neeltje; VERSCHUEREN Ineke; FIGDOR Carl; VAN DER MEER Jos W. M.; TORENSMA Ruurd; KULLBERG Bart Jan
- Morphological plasticity of Candida albicans is a major virulence factor. Using pH-dependent dimorphism we show, that human dendritic cells (DC) recognize filamentous forms and blastoconidia of a virulent C. albicans isolate (strain SC5314). Heat inactivated and viable blastoconidia are rapidly phagocytosed by human DC. However, viable yeast cells start to filament inside the DC at later stages of infection, leading to penetration and loss of cellular integrity. The cytokine burst of human DC induced upon contact with Candida is dominated by the granulocyte-activating, chemotactic factor IL-8 and the proinflammatory mediator TNF-alpha. Blastoconidia induce markedly lower cytokine levels than filamentous forms. Whereas IL-8 secretion is mainly cell mass dependent, release of TNF-alpha, a major proinflammatory cytokine, is clearly dependent on the morphology of Candida.
Polymorphism of Candida albicans is a major factor in the interaction with human dendritic cells.
- Dendritic cells (DC) are antigen presenting cells that act as sentinels, acquiring antigen and transporting it to lymphoid tissue where they have the unique ability to activate naïve T cells. From this pivotal position at the intersection of innate and adaptive immunity, DC shape many aspects of the developing immune response. They can determine whether non-responsiveness (tolerance) or an active immune response occurs, whether a type 1 or type 2 response predominates, and they may control tissue specific homing of antigen specific effector cells. Microbial products play a central role in modulating DC function and influencing these different immune outcomes. Using molecules including toll-like receptors, DC recognise and respond to microbe specific molecular structures. DC can distinguish between and initiate different responses to even closely related organisms. Emerging evidence suggests that intestinal DC are critical for regulation of immunity in the gut. They are likely to be pivotal in the balance between tolerance and active immunity to commensal microorganisms that is fundamental to inflammatory conditions, including Crohn’s disease and ulcerative colitis. Here, we describe how interactions between DC and microbial products contribute to regulation of immune responses and explore the concept that interactions between DC and commensal organisms may be responsible for maintaining intestinal immune homeostasis.
The dendritic cell: its role in intestinal inflammation and relationship with gut bacteria
A J Stagg, A L Hart, S C Knight, and M A Kamm
- Candida albicans is a component of the normal flora of the alimentary tract and also is found on the mucocutaneous membranes of the healthy host. Candida is the leading cause of invasive fungal disease in premature infants, diabetics, and surgical patients, and of oropharyngeal disease in AIDS patients. As the induction of cell-mediated immunity to Candida is of critical importance in host defense, we sought to determine whether human dendritic cells (DC) could phagocytose and degrade Candida and subsequently present Candida antigens to T cells. Immature DC obtained by culture of human monocytes in the presence of granulocyte-macrophage colony-stimulating factor and interleukin-4 phagocytosed unopsonized Candida in a time-dependent manner, and phagocytosis was not enhanced by opsonization of Candida in serum. Like macrophages (M), DC recognized Candida by the mannose-fucose receptor. Upon ingestion, DC killed Candida as efficiently as human M, and fungicidal activity was not enhanced by the presence of fresh serum. Although phagocytosis of Candida by DC stimulated the production of superoxide anion, inhibitors of the respiratory burst (or NO production) did not inhibit killing of Candida, even when phagocytosis was blocked by preincubation of DC with cytochalasin D. Further, although apparently only modest phagolysosomal fusion occurred upon DC phagocytosis of Candida, killing of Candida under anaerobic conditions was almost equivalent to killing under aerobic conditions. Finally, DC stimulated Candida-specific lymphocyte proliferation in a concentration-dependent manner after phagocytosis of both viable and heat-killed Candida cells. These data suggest that, in vivo, such interactions between DC and C. albicans may facilitate the induction of cell-mediated immunity.
Candida albicans Is Phagocytosed, Killed, and Processed for Antigen Presentation by Human Dendritic Cells
Simon L. Newman and Angela Holly
- The fungus Candida albicans behaves as a commensal as well as a true pathogen of areas highly enriched in dendritic cells, such as skin and mucosal surfaces. The ability of the fungus to reversibly switch between unicellular yeast to filamentous forms is thought to be important for virulence. However, whether it is the yeast or the hyphal form that is responsible for pathogenicity is still a matter of debate. Here we show the interaction, and consequences, of different forms of C. albicans with dendritic cells. Immature myeloid dendritic cells rapidly and efficiently phagocytosed both yeasts and hyphae of the fungus. Phagocytosis occurred through different phagocytic morphologies and receptors, resulting in phagosome formation. However, hyphae escaped the phagosome and were found lying free in the cytoplasm of the cells. In vitro, ingestion of yeasts activated dendritic cells for interleukin (IL)-12 production and priming of T helper type 1 (Th1) cells, whereas ingestion of hyphae inhibited IL-12 and Th1 priming, and induced IL-4 production. In vivo, generation of antifungal protective immunity was induced upon injection of dendritic cells ex vivo pulsed with Candida yeasts but not hyphae. The immunization capacity of yeast-pulsed dendritic cells was lost in the absence of IL-12, whereas that of hypha-pulsed dendritic cells was gained in the absence of IL-4. These results indicate that dendritic cells fulfill the requirement of a cell uniquely capable of sensing the two forms of C. albicans in terms of type of immune responses elicited. By the discriminative production of IL-12 and IL-4 in response to the nonvirulent and virulent forms of the fungus, dendritic cells appear to meet the challenge of Th priming and education in C. albicans saprophytism and infections.
Dendritic Cells Discriminate between Yeasts and Hyphae of the Fungus Candida albicans: Implications for Initiation of T Helper Cell Immunity in Vitro and in Vivo
Cristiana Fè d’Ostiani, Giuseppe Del Sero, Angela Bacci, Claudia Montagnoli, Antonio Spreca, Antonella Mencacci, Paola Ricciardi-Castagnoli, and Luigina Romani
- The ability of Candida albicans to convert from the yeast (Y) form to mycelial forms through germ tube (GT) formation is considered a key feature of the transition of the organism from commensalism to virulence. We show here that human monocytes cultured with granulocyte-macrophage colony-stimulating factor and interleukin-4 (IL-4) after phagocytosis of Y forms did not differentiate into dendritic cells (DCs); they retained CD14, did not acquire CD1a, and were unable to express the maturation markers CD83 and CCR7. Moreover, they did not produce IL-12p70 but secreted IL-10. In addition, they spontaneously expressed high levels of tumor necrosis factor alpha (TNF-α), IL-6, and IL-8 mRNA transcripts and were able to induce proliferation of alloreactive memory but not naïve T lymphocytes. Conversely, monocytes that had phagocytosed GT forms differentiated into mature CD83+ and CCR7+ DCs; however, there was no up-regulation of CD40, CD80, and major histocompatibility complex class II, irrespective of lipopolysaccharide (LPS) treatment. In addition, these cells were unable to produce IL-12 even after LPS stimulation, but they were not functionally exhausted, as shown by their capacity to express TNF-α and IL-8 mRNA transcripts. These cells were able to prime naïve T cells but not to induce their functional polarization into effector cells. These data indicate that phagocytosis of Y and GT forms has profound and distinct effects on the differentiation pathway of monocytes. Thus, the differentiation of human monocytes into DCs appears to be tunable and exploitable by C. albicans to elude immune surveillance.
Candida albicans Yeast and Germ Tube Forms Interfere Differently with Human Monocyte Differentiation into Dendritic Cells: a Novel Dimorphism-Dependent Mechanism To Escape the Host’s Immune Response
Antonella Torosantucci, Giulia Romagnoli, Paola Chiani, Annarita Stringaro, Pasqualina Crateri, Sabrina Mariotti, Raffaela Teloni, Giuseppe Arancia, Antonio Cassone, and Roberto Nisini
- A new study has uncovered the genetic wiring diagram underlying the infectiousness of Candida albicans, a fungus that causes thrush in babies, vaginal infections in women, and life-threatening infections in chemotherapy and AIDS patients. The study, led by Dr. Gerald R. Fink, Director of the Whitehead Institute for Biomedical Research, reveals that one key to Candida’s infectiousness lies in its ability to switch from a rounded form to filamentous forms. Fungal infections in hospitalized patients have almost doubled throughout the 1980s, often with life-threatening results in individuals with weakened immune systems. Candida, in particul