Organ-specific mechanisms linking innate and adaptive antifungal immunity.

Fungal infections remain a significant global health problem in humans. Fungi infect millions of people worldwide and cause from acute superficial infections to life-threatening systemic disease to chronic illnesses. Trying to decipher the complex innate and adaptive immune mechanisms that protect humans from pathogenic fungi is therefore a key research goal that may lead to immune-based therapeutic strategies and improved patient outcomes. In this review, we summarize how the cells and molecules of the innate immune system activate the adaptive immune system to elicit long-term immunity to fungi. We present current knowledge and exciting new advances in the context of organ-specific immunity, outlining the tissue-specific tropisms for the major pathogenic fungi of humans, the antifungal functions of tissue-resident myeloid cells, and the adaptive immune responses required to protect specific organs from fungal challenge.

The domestic pig as human-relevant large animal model to study adaptive antifungal immune responses against airborne Aspergillus fumigatus.

Pulmonary mucosal immune response is critical for preventing opportunistic Aspergillus fumigatus infections. Although fungus-specific CD4+ T cells in blood are described to reflect the actual host-pathogen interaction status, little is known about Aspergillus-specific pulmonary T-cell responses. Here, we exploit the domestic pig as human-relevant large animal model and introduce antigen-specific T-cell enrichment in pigs to address Aspergillus-specific T cells in the lung compared to peripheral blood. In healthy, environmentally Aspergillus-exposed pigs, the fungus-specific T cells are detectable in blood in similar frequencies as observed in healthy humans and exhibit a Th1 phenotype. Exposing pigs to 106 cfu/m3 conidia induces a long-lasting accumulation of Aspergillus-specific Th1 cells locally in the lung and also systemically. Temporary immunosuppression during Aspergillus-exposure showed a drastic reduction in the lung-infiltrating antifungal T-cell responses more than 2 weeks after abrogation of the suppressive treatment. This was reflected in blood, but to a much lesser extent. In conclusion, by using the human-relevant large animal model the pig, this study highlights that the blood clearly reflects the mucosal fungal-specific T-cell reactivity in environmentally exposed as well as experimentally exposed healthy pigs. But, immunosuppression significantly impacts the mucosal site in contrast to the initial systemic immune response.

Caspofungin Increases Fungal Chitin and Eosinophil and γδ T Cell-Dependent Pathology in Invasive Aspergillosis.

The polysaccharide-rich fungal cell wall provides pathogen-specific targets for antifungal therapy and distinct molecular patterns that stimulate protective or detrimental host immunity. The echinocandin antifungal caspofungin inhibits synthesis of cell wall ß-1,3-glucan and is used for prophylactic therapy in immune-suppressed individuals. However, breakthrough infections with fungal pathogen Aspergillus fumigatus are associated with caspofungin prophylaxis. In this study, we report in vitro and in vivo increases in fungal surface chitin in A. fumigatus induced by caspofungin that was associated with airway eosinophil recruitment in neutropenic mice with invasive pulmonary aspergillosis (IA). More importantly, caspofungin treatment of mice with IA resulted in a pattern of increased fungal burden and severity of disease that was reversed in eosinophil-deficient mice. Additionally, the eosinophil granule proteins major basic protein and eosinophil peroxidase were more frequently detected in the bronchoalveolar lavage fluid of lung transplant patients diagnosed with IA that received caspofungin therapy when compared with azole-treated patients. Eosinophil recruitment and inhibition of fungal clearance in caspofungin-treated mice with IA required RAG1 expression and γδ T cells. These results identify an eosinophil-mediated mechanism for paradoxical caspofungin activity and support the future investigation of the potential of eosinophil or fungal chitin-targeted inhibition in the treatment of IA.

Identification and characterization of a novel antimicrobial protein from the housefly Musca domestica.

Antimicrobial peptides/proteins are immune-related molecules that are widely distributed in bacteria, fungi, plants, invertebrates and higher animals. They have exhibited great potential to be developed into antimicrobial drugs. The housefly, Musca domestica, lives in a highly contaminated environment and has adapted a robust immune system against various pathogens. As an effort to search for new antimicrobial molecules in the housefly, we investigated the function of an uncharacterized gene firstly by confirming that its expression was induced by infection in M. domestica. The corresponding protein was then shown to have potent antimicrobial activity. Scanning Electron Microscopy data showed that treatment of C. albicans cells with the protein caused cell size decreasing and cell elongation. The results here suggest the protein a novel class of antimicrobial protein and provide new insights into the immunological mechanisms by which M. domestica combats invading C. albicans.

Induced accumulation of tyramine, serotonin, and related amines in response to Bipolaris sorokiniana infection in barley.

The inducible metabolites were analyzed in barley leaves inoculated with Bipolaris sorokiniana, the causal agent of spot blotch of barley. HPLC analysis revealed that B. sorokiniana-infected leaves accumulated 4 hydrophilic compounds. They were purified by ODS column chromatography and preparative HPLC. Spectroscopic analyses revealed that they were tyramine (1), 3-(2-aminoethyl)-3-hydroxyindolin-2-one (2), serotonin (3), and 5,5'-dihydroxy-2,4'-bitryptamine (4). Among these, 2 and 4 have not been reported as natural products. They showed antifungal activity in an assay of inhibition of B. sorokiniana conidia germination, suggesting that they play a role in the chemical defense of barley as phytoalexins. The accumulation of 1-4 was examined also in the leaves of rice and foxtail millet. Rice leaves accumulated 2, 3, and 4, whereas foxtail millet leaves accumulated 3 and 4 in response to pathogen attack, suggesting the generality of accumulation of 3 and 4 in the Poaceae species.

Role of Granulocyte-Macrophage Colony-Stimulating Factor Signaling in Regulating Neutrophil Antifungal Activity and the Oxidative Burst During Respiratory Fungal Challenge.

Granulocyte-macrophage colony-stimulating factor (GM-CSF) is a pleiotropic cytokine that plays a critical role in regulating myeloid cell host defense. In this study, we demonstrated that GM-CSF signaling plays an essential role in antifungal defense against Aspergillus fumigatus. Mice that lack the GM-CSF receptor ß chain (GM-CSFRß) developed invasive hyphal growth and exhibited impaired survival after pulmonary challenge with A. fumigatus conidia. GM-CSFRß signaling regulated the recruitment of inflammatory monocytes to infected lungs, but not the recruitment of effector neutrophils. Cell-intrinsic GM-CSFRß signaling mediated neutrophil and inflammatory monocyte antifungal activity, because lung GM-CSFRß(-/-) leukocytes exhibited impaired conidial killing compared with GM-CSFRß(+/+) counterparts in mixed bone marrow chimeric mice. GM-CSFRß(-/-) neutrophils exhibited reduced (hydrogenated) nicotinamide adenine dinucleotide phosphate (NADPH) oxidase activity in vivo. Conversely, administration of recombinant GM-CSF enhanced neutrophil NADPH oxidase function, conidiacidal activity, and lung fungal clearance in A. fumigatus-challenged mice. Thus, our study illustrates the functional role of GM-CSFRß signaling on lung myeloid cell responses against inhaled A. fumigatus conidia and demonstrates a benefit for systemic GM-CSF administration.

Antifungal Activity of Plasmacytoid Dendritic Cells against Cryptococcus neoformans In Vitro Requires Expression of Dectin-3 (CLEC4D) and Reactive Oxygen Species.

Conventional dendritic cells (cDCs) are critical for protection against pulmonary infection with the opportunistic fungal pathogen Cryptococcus neoformans; however, the role of plasmacytoid dendritic cells (pDCs) is unknown. We show for the first time that murine pDCs have direct activity against C. neoformans via reactive oxygen species (ROS), a mechanism different from that employed to control Aspergillus fumigatus infections. The anticryptococcal activity of murine pDCs is independent of opsonization but appears to require the C-type lectin receptor Dectin-3, a receptor not previously evaluated during cryptococcal infections. Human pDCs can also inhibit cryptococcal growth by a mechanism similar to that of murine pDCs. Experimental pulmonary infection of mice with a C. neoformans strain that induces protective immunity demonstrated that recruitment of pDCs to the lungs is CXCR3 dependent. Taken together, our results show that pDCs inhibit C. neoformans growth in vitro via the production of ROS and that Dectin-3 is required for optimal growth-inhibitory activity.

Peanut defensins: Novel allergens isolated from lipophilic peanut extract.

BACKGROUND: Peanut is one of the most hazardous sources of food allergens. Unknown allergens are still hidden in the complex lipophilic matrix. These allergens need to be discovered to allow estimation of the allergenic risk for patients with peanut allergy and to further improve diagnostic measures. OBJECTIVE: We performed detection, isolation, and characterization of novel peanut allergens from lipophilic peanut extract. METHODS: Extraction of roasted peanuts were performed under defined extraction conditions and examined by means of 2-dimensional PAGE. Subsequently, chromatographic methods were adapted to isolate low-molecular-weight components. Proteins were studied by using SDS-PAGE and immunoblotting with sera from patients with peanut allergy. For allergen identification protein sequencing, homology search and mass spectrometry were applied. Functional characterization for allergenicity was performed by using the basophil activation assay and for antimicrobial activity by using inhibition assays of different bacteria and fungi. RESULTS: IgE-reactive proteins of 12, 11, and 10 kDa were first detected after chloroform/methanol extraction in the flow through of hydrophobic interaction chromatography. The proteins were able to activate basophils of patients with peanut allergy. N-terminal sequencing and homology search in the expressed sequence tag database identified the allergens as peanut defensins, which was confirmed by using mass spectrometry. On microbial cell cultures, the peanut defensins showed inhibitory effects on the mold strains of the genera Cladosporium and Alternaria but none on bacteria. CONCLUSIONS: We identified defensins as novel peanut allergens (Ara h 12 and Ara h 13) that react in particular with IgE of patients with severe peanut allergy. Their antimicrobial activity is solely antifungal.

Synthesis and immunological studies of linear oligosaccharides of ß-glucan as antigens for antifungal vaccine development.

Antifungal vaccines have recently engendered considerable excitement for counteracting the resurgence of fungal infections. In this context, ß-glucan, which is abundantly expressed on all fungal cell surfaces, functionally necessary for fungi, and immunologically active, is an attractive target antigen. Aiming at the development of effective antifungal vaccines based on ß-glucan, a series of its oligosaccharide derivatives was designed, synthesized, and coupled with a carrier protein, keyhole limpet hemocyanin (KLH), to form new semisynthetic glycoconjugate vaccines. In this article, a convergent and effective synthetic strategy using preactivation-based iterative glycosylation was developed for the designed oligosaccharides. The strategy can be widely useful for rapid construction of large oligo-ß-glucans with shorter oligosaccharides as building blocks. The KLH conjugates of the synthesized ß-glucan hexa-, octa-, deca-, and dodecasaccharides were demonstrated to elicit high titers of antigen-specific total and IgG antibodies in mice, suggesting the induction of functional T cell-mediated immunity. Moreover, it was revealed that octa-, deca-, and dodeca-ß-glucans were much more immunogenic than the hexamer and that the octamer was the best among these. The results suggested that the optimal oligosaccharide sequence of ß-glucan required for exceptional immunogenicity was a hepta- or octamer and that longer glucans are not necessarily better antigens, a finding that may be of general importance. Most importantly, the octa-ß-glucan-KLH conjugate provoked protective immunity against Candida albicans infection in a systemic challenge model in mice, suggesting the great potential of this glycoconjugate as a clinically useful immunoprophylactic antifungal vaccine.

ß-Microseminoprotein endows post coital seminal plasma with potent candidacidal activity by a calcium- and pH-dependent mechanism.

The innate immune factors controlling Candida albicans are mostly unknown. Vulvovaginal candidiasis is common in women and affects approximately 70-75% of all women at least once. Despite the propensity of Candida to colonize the vagina, transmission of Candida albicans following sexual intercourse is very rare. This prompted us to investigate whether the post coital vaginal milieu contained factors active against C. albicans. By CFU assays, we found prominent candidacidal activity of post coital seminal plasma at both neutral and the acid vaginal pH. In contrast, normal seminal plasma did not display candidacidal activity prior to acidification. By antifungal gel overlay assay, one clearing zone corresponding to a protein band was found in both post coital and normal seminal plasma, which was subsequently identified as ß-microseminoprotein. At neutral pH, the fungicidal activity of ß-microseminoprotein and seminal plasma was inhibited by calcium. By NMR spectroscopy, amino acid residue E(71) was shown to be critical for the calcium coordination. The acidic vaginal milieu unleashed the fungicidal activity by decreasing the inhibitory effect of calcium. The candidacidal activity of ß-microseminoprotein was mapped to a fragment of the C-terminal domain with no structural similarity to other known proteins. A homologous fragment from porcine ß-microseminoprotein demonstrated calcium-dependent fungicidal activity in a CFU assay, suggesting this may be a common feature for members of the ß-microseminoprotein family. By electron microscopy, ß-microseminoprotein was found to cause lysis of Candida. Liposome experiments demonstrated that ß-microseminoprotein was active towards ergosterol-containing liposomes that mimic fungal membranes, offering an explanation for the selectivity against fungi. These data identify ß-microseminoprotein as an important innate immune factor active against C. albicans and may help explain the low sexual transmission rate of Candida.

Melanocytes and melanin represent a first line of innate immunity against Candida albicans.

Melanocytes are dendritic cells located in the skin and mucosae that synthesize melanin. Some infections induce hypo- or hyperpigmentation, which is associated with the activation of Toll-like receptors (TLRs), especially TLR4. Candida albicans is an opportunist pathogen that can switch between blastoconidia and hyphae forms; the latter is associated with invasion. Our objectives in this study were to ascertain whether C. albicans induces pigmentation in melanocytes and whether this process is dependent on TLR activation, as well as relating this with the antifungal activity of melanin as a first line of innate immunity against fungal infections. Normal human melanocytes were stimulated with C. albicans supernatants or with crude extracts of the blastoconidia or hyphae forms, and pigmentation and TLR2/TLR4 expression were measured. Expression of the melanosomal antigens Melan-A and gp100 was examined for any correlation with increased melanin levels or antifungal activity in melanocyte lysates. Melanosomal antigens were induced earlier than cell pigmentation, and hyphae induced stronger melanization than blastoconidia. Notably, when melanocytes were stimulated with crude extracts of C. albicans, the cell surface expression of TLR2/TLR4 began at 48 h post-stimulation and peaked at 72 h. At this time, blastoconidia induced both TLR2 and TLR4 expression, whereas hyphae only induced TLR4 expression. Taken together, these results suggest that melanocytes play a key role in innate immune responses against C. albicans infections by recognizing pathogenic forms of C. albicans via TLR4, resulting in increased melanin content and inhibition of infection.

Designing a new antifungal glycoconjugate vaccine.

Bacterial capsular polysaccharides have been used as effective vaccines for adults but infants and seniors respond poorly to these immunogens because pure polysaccharides are unable to activate T-cells resulting in antibodies of low affinity and poor immunological memory. These deficiencies are addressed by conjugate vaccines composed of bacterial polysaccharide covalently attached to protein carriers such as tetanus or diphtheria toxoids. These vaccines activate T-cells and have been hugely effective in reducing the incidence of dangerous infectious diseases such as bacterial meningitis in infants and adults. The methods of conjugation often produce conjugate vaccines that contain polysaccharides with several cross links to one or more protein carrier molecules. Synthesis of large oligosaccharides derivatized to achieve single site attachment to protein has established the minimal size of an oligosaccharide that is required to produce protective antibody. Several examples are described where such oligosaccharides range in size from 8 to 16 monosaccharides. A more limited set of examples show that protective antibodies may be elicited by conjugate vaccines composed of tri and tetrasaccharide epitopes. One example is the ß-mannan present in the phosphomannan glycoprotein of Candida albicans. Reverse engineering a protective antibody that recognizes this oligosaccharide revealed that a disaccharide was the minimal epitope. Further epitope mapping by functional group modification established that the internal mannose residue of a disaccharide was involved in the most important antibody-disaccharide interactions. NMR binding studies in combination with homology modeling of the bound ß-mannan antigen suggested an optimum oligosaccharide for inclusion in a conjugate vaccine. Several conjugate vaccines were developed to test these conclusions. Immunization of mice or rabbits with conjugates containing disaccharide or trisaccharide conjugated to immunogenic proteins followed by live challenge experiments showed that the vaccines reduced fungal burden. The results of these and other studies suggest new approaches to novel synthetic conjugate vaccines.

Autoimmune polyendocrinopathy candidiasis ectodermal dystrophy and other primary immunodeficiency diseases help to resolve the nature of protective immunity against chronic mucocutaneous candidiasis.

PURPOSE OF REVIEW: This review summarizes and discusses the most recent and important publications describing Mendelian diseases associated with susceptibility to chronic mucocutaneous candidiasis (CMC) as a means of gaining insight into the pathogenesis of this immunodeficiency. RECENT FINDINGS: Impairment to T helper 17 (Th17) cell-associated signalling pathways are common in immunodeficiency syndromes associated with CMC infections. Mutations in CARD9, STAT3, IL17RA, IL17F, STAT1, and IL12RB and polymorphisms in Dectin 1 and interleukin-22 (IL-22) encoding genes have been shown to impair the development or function of Th17 cells and are associated with susceptibility to candidiasis. Studies on autoimmune polyendocrinopathy candidiasis ectodermal dystrophy have revealed autoimmunity to Th17 cytokines and cells as the basis for CMC. IL-17A, IL-17F, and IL-22 induce production of antimicrobial peptides and chemoattractants that recruit neutrophils in response to invading fungi. Th17 cell-associated cytokines may play a role in shaping the host's microbiome (that competes with C. albicans) preventing overgrowth of this pathogen. Recent evidence also suggests that IL-22 together with IL-17F might be the most important Th17 cytokine in protection against Candida. SUMMARY: Dissection of critical molecular and immunological mechanisms will allow the development of new treatments for primary and secondary immunodeficiency disorders resulting in chronic Candida infections.

Prokaryotic expression of pathogenesis related protein 1 gene from Nicotiana benthamiana: antifungal activity and preparation of its polyclonal antibody.

The nucleotide sequence of the pathogenesis-related protein 1(PR-1) gene was obtained from Nicotiana benthamiana using RT-PCR. Restriction enzyme cutting sites of EcoRI and NotI were introduced to the ORF fragments of PR-1, they were then linked together with pET-30a (+) and transformed into E. coli BL21 (DE3). The target protein was induced by 1.5 mM IPTG, at 37°C for 4 h. The expressed protein was purified by Ni-NTA and an anti-NbPR-1 polyclonal antibody was prepared using rabbits. The antibody could detect the expression of PR-1 in N. benthamiana and other Nicotiana plants. NbPR-1 protein has four α-helices and two ß-sheets by homology modeling. Furthermore, the purified NbPR-1 protein exhibited a broad-spectrum antifungal activity.

Effects of interferon-γ and granulocyte colony-stimulating factor on antifungal activity of human polymorphonuclear neutrophils against Candida albicans grown as biofilms or planktonic cells.

Candida albicans is a leading cause of biofilm-related infections. As Candida biofilms are recalcitrant to host defenses, we sought to determine the effects of interferon-γ and granulocyte colony-stimulating factor, two pro-inflammatory cytokines, on the antifungal activities of human polymorphonuclear neutrophils (PMNs) against C. albicans biofilms, using an in vitro biofilm model. Priming of PMNs by these cytokines augmented fungal damage of planktonic cells; however, priming of PMNs did not have the same effect against Candida biofilms. Biofilm phenotype appears to play an important role in protecting C. albicans from the innate immune system.

Immunomodulatory properties of antifungal agents on phagocytic cells.

Phagocytic cells, particularly neutrophils and monocytes/macrophages, are the first line and the most effective form of innate host defence against pathogenic fungi. During antifungal therapy these phagocytic cells are also exposed to antifungal agents. In the phagocyte-fungus-antifungal agent interplay, drugs may directly interact with phagocytes through specific pattern recognition receptors, leading to altered antifungal activities. Antifungal agents, through modulation of fungal virulence, may initiate different immune response programs in the phagocytes, leading to antifungal synergism/antagonism or up-regulation of gene expression for a pro-inflammatory response. Additionally, indirect modulation of phagocyte behavior by pretreatment of neutrophils, monocytes, and macrophages with cytokines and exposure to antifungal agents have shown promising findings for combined drug-cytokine therapy that may improve treatment of life-threatening fungal diseases. In this review, we discuss the main in vitro and in vivo immunomodulatory effects of antifungal agents on phagocytes in response to pathogenic fungi, as well as we address underlying immunopharmacologic mechanisms and their potential impact on clinical outcome.

Micafungin-induced thrombotic thrombocytopenic purpura: a case report and review of the literature.

Thrombotic thrombocytopenic purpura (TTP) is a life-threatening multisystem disorder characterized by microangiopathic hemolytic anemia and thrombocytopenia accompanied by microvascular thrombosis that causes variable degrees of tissue ischemia and infarction leading to organ dysfunction. Drug-associated thrombotic thrombocytopenic purpura and hemolytic uremic syndrome has been recognized for several years. The most commonly implicated drugs are mitomycin C, cyclosporine, quinine, clopidogrel, and ticlopidine. Recent advances have suggested that like in idiopathic TTP, the most likely pathogenesis for drug-induced TTP is either an immune-mediated phenomenon involving the ADAMTS13 metalloprotease or direct endothelial toxicity. In this communication, we report a case of micafungin-induced TTP. Micafungin is a new antifungal drug of the Echinocandins group. Whether micafungin induces autoantibodies against ADAMTS13 or not, this needs further evaluation, but TTP should be recognized as a possible complication of micafungin. Clinicians should be alert to this adverse effect of micafungin and monitor platelet counts in patients receiving this drug.

An altered camelid-like single domain anti-idiotypic antibody fragment of HM-1 killer toxin: acts as an effective antifungal agent.

Phage-display and competitive panning elution leads to the identification of minimum-sized antigen binders together with conventional antibodies from a mouse cDNA library constructed from HM-1 killer toxin neutralizing monoclonal antibody (nmAb-KT). Antigen-specific altered camelid-like single-domain heavy chain antibody (scFv K2) and a conventional antibody (scFv K1) have been isolated against the idiotypic antigen nmAb-KT. The objectives of the study were to examine (1) their properties as compared to conventional antibodies and also (2) their antifungal activity against different pathogenic and non-pathogenic fungal species. The alternative small antigen-binder, i.e., the single-domain heavy chain antibody, was originated from a conventional mouse scFv phage library through somatic hyper-mutation while selection against antigen. This single-domain antibody fragment was well expressed in bacteria and specifically bound with the idiotypic antigen nmAb-KT and had a high stability and solubility. Experimental data showed that the binding affinity for this single-domain antibody was 272-fold higher (K(d)=1.07×10(-10) M) and antifungal activity was three- to fivefold more efficient (IC(50)=0.46×10(-6) to 1.17×10(-6) M) than that for the conventional antibody (K(d)=2.91×10(-8) M and IC(50)=2.14×10(-6) to 3.78×10(-6) M). The derived single-domain antibody might be an ideal scaffold for anti-idiotypic antibody therapy and the development of smaller peptides or peptide mimetic drugs due to their less complex antigen-binding site. We expect that such single-domain synthetic antibodies will find their way into a number of biotechnological or medical applications.

Protection of immunocompromised mice from fungal infection with a thymus growth-stimulatory component from Selaginella involvens, a fern.

CONTEXT: Recent studies have shown that the water extract of Selaginella involvens (Sw.) Spring, a wild fern, exhibits thymus growth-stimulatory activity in adult mice (reversal of involution of thymus) and remarkable anti-lipid peroxidation activity. Follow-up studies were carried out in the present study. MATERIALS AND METHODS: Activity-guided isolation of the active component (AC) was carried out. The effect of AC on immune function was studied using fungal (Aspergillus fumigatus) challenge in cortisone-treated mice. The in vitro antifungal activity of AC was assayed using disc diffusion assay. In vitro and in vivo effect of AC on DNA synthesis in thymus was studied using (3)H-thymidine incorporation. In in vitro anti-lipid peroxidation, hydroxyl radical scavenging and inhibition of superoxide production were assayed. RESULTS: The active principle/component (AC) was isolated in a chromatographically pure form from the water extract of S. involvens. AC showed positive reaction to glycosides. AC possessed both thymus growth-stimulatory and antioxidant properties. It protected cortisone-treated mice from A. fumigatus challenge. It did not exhibit in vitro antifungal activity. Increased (3)H-thymidine incorporation was observed in the reticuloepithelium of thymus obtained from AC-treated mice. However, in vitro AC treatment to thymus for 5 h did not result in an increase in (3)H-thymidine incorporation. DISCUSSION AND CONCLUSION: AC (named as Selagin), from S. involvens, could reverse involution of thymus to a large extent, exhibit remarkable antioxidant activity, and protect immunocompromised mice from fungal infection. Therefore, it is very promising for the development of a drug to ameliorate old age-related health problems and prolong lifespan.

Antifungal activity of dendritic cell lysosomal proteins against Cryptococcus neoformans.

Cryptococcal meningitis is a life-threatening disease among immune compromised individuals that is caused by the opportunistic fungal pathogen Cryptococcus neoformans. Previous studies have shown that the fungus is phagocytosed by dendritic cells (DCs) and trafficked to the lysosome where it is killed by both oxidative and non-oxidative mechanisms. While certain molecules from the lysosome are known to kill or inhibit the growth of C. neoformans, the lysosome is an organelle containing many different proteins and enzymes that are designed to degrade phagocytosed material. We hypothesized that multiple lysosomal components, including cysteine proteases and antimicrobial peptides, could inhibit the growth of C. neoformans. Our study identified the contents of the DC lysosome and examined the anti-cryptococcal properties of different proteins found within the lysosome. Results showed several DC lysosomal proteins affected the growth of C. neoformans in vitro. The proteins that killed or inhibited the fungus did so in a dose-dependent manner. Furthermore, the concentration of protein needed for cryptococcal inhibition was found to be non-cytotoxic to mammalian cells. These data show that many DC lysosomal proteins have antifungal activity and have potential as immune-based therapeutics.