The C-type lectin receptor Dectin-1 plays a pivotal role in antifungal immunity. In this study, the recently characterized human DECTIN1 Y238X early stop codon polymorphism leading to diminished Dectin-1 receptor activity was studied in relation to invasive aspergillosis susceptibility and severity in patients receiving hematopoietic stem cell transplantation. We found that the presence of the DECTIN1 Y238X polymorphism in either donors or recipients of hematopoietic stem cell transplantation increased susceptibility to aspergillosis, with the risk being highest when the polymorphism was present simultaneously in both donors and recipients (adjusted hazard ratio ‫؍‬ 3.9; P ‫؍‬ .005). Functionally, the Y238X polymorphism impaired the production of interferon-␥ and interleukin-10 (IL-10), in addition to IL-1␤, IL-6, and IL-17A, by human peripheral mononuclear cells and Dectin-1 on human epithelial cells contributed to fungal recognition. Mechanistically, studies on preclinical models of infection in intact or bone marrow-transplanted Dectin-1 knockout mice revealed that protection from infection requires a distinct, yet complementary, role of both donor and recipient Dectin-1. This study discloses Dectin-1 deficiency as a novel susceptibility factor for aspergillosis in high-risk patients and identifies a previously unsuspected role for Dectin-1 in antifungal immunity that is the ability to control both resistance and tolerance to the fungus contingent on hematopoietic/ nonhematopoietic compartmentalization. (Blood. 2010;116(24):5394-5402) IntroductionAspergillus spp are ubiquitous in nature, and the spectrum of diseases they cause is myriad, including saprophytic colonization of preexisting cavities (aspergilloma), allergic asthma, hypersensitivity pneumonitis, allergic bronchopulmonary aspergillosis occurring as a complication of bronchial asthma or cystic fibrosis, and disseminated disease associated with high mortality rates in patients with hematologic malignancies and recipients of solid organs and stem cell transplantations. 1 Immunocompetent and nonatopic subjects are relatively resistant to infections, and disease occurs in the setting of host damage. 2 The association of persistent inflammation with intractable infection is common in nonneutropenic patients after hematopoietic stem cell transplantation (HSCT) as well as in allergic fungal diseases. 2 The current understanding of the pathophysiology underlying Aspergillus infection and disease highlights a truly bipolar nature of the inflammatory process in infection. Early inflammation prevents or limits infection, but an uncontrolled response may eventually oppose disease eradication. This condition is crucially exemplified in mice with chronic granulomatous disease, in which an intrinsic, genetically determined failure to control inflammation to sterile fungal components determines the animals' inability to resolve an actual infection with A fumigatus. 3 A main implication of these findings is that, at least in specific clinical settings, it is an exag...

Dendritic cells (DCs) show a remarkable functional plasticity in the recognition of Aspergillus fumigatus and orchestrate the antifungal immune resistance in the lungs. Here, we show that thymosin alpha 1, a naturally occurring thymic peptide, induces functional maturation and interleukin-12 production by fungus-pulsed DCs through the p38 mitogen-activated protein kinase/nuclear factor (NF)-kappaB-dependent pathway. This occurs by signaling through the myeloid differentiation factor 88-dependent pathway, involving distinct Toll-like receptors. In vivo, the synthetic peptide activates T-helper (Th) cell 1-dependent antifungal immunity, accelerates myeloid cell recovery, and protects highly susceptible mice that received hematopoietic transplants from aspergillosis. By revealing the unexpected activity of an old molecule, our finding provides the rationale for its therapeutic utility and qualify the synthetic peptide as a candidate adjuvant promoting the coordinated activation of the innate and adaptive Th immunity to the fungus.

Polymorphonuclear neutrophils (PMNs) are essential in initiation and execution of the acute inflammatory response and subsequent resolution of fungal infection. PMNs, however, may act as double-edged swords, as the excessive release of oxidants and proteases may be responsible for injury to organs and fungal sepsis. To identify regulatory mechanisms that may balance PMN-dependent protection and immunopathology in fungal infections, the involvement of different TLR-activation pathways was evaluated on human PMNs exposed to the fungus Aspergillus fumigatus. Recognition of Aspergillus and activation of PMNs occurred through the involvement of distinct members of the TLR family, each likely activating specialized antifungal effector functions. By affecting the balance between fungicidal oxidative and nonoxidative mechanisms, pro- and anti-inflammatory cytokine production, and apoptosis vs necrosis, the different TLRs ultimately impacted on the quality of microbicidal activity and inflammatory pathology. Signaling through TLR2 promoted the fungicidal activity of PMNs through oxidative pathways involving extracellular release of gelatinases and proinflammatory cytokines while TLR4 favored the oxidative pathways through the participation of azurophil, myeloperoxidase-positive, granules and IL-10. This translated in vivo in the occurrence of different patterns of fungal clearance and inflammatory pathology. Both pathways were variably affected by signaling through TLR3, TLR5, TLR6, TLR7, TLR8, and TLR9. The ability of selected individual TLRs to restore antifungal functions in defective PMNs suggests that the coordinated outputs of activation of multiple TLRs may contribute to PMN function in aspergillosis.

Toll-like receptors (TLRs) are important components of innate immunity. We investigated the association between polymorphisms in the TLR2, TLR4, and TLR9 genes and susceptibility to noninvasive forms of pulmonary aspergillosis. A significant association was observed between allele G on Asp299Gly (TLR4) and chronic cavitary pulmonary aspergillosis (odds ratio [OR], 3.46; P =.003). Susceptibility to allergic bronchopulmonary aspergillosis was associated with allele C on T-1237C (TLR9) (OR, 2.49; P =. 043). No particular polymorphism was associated with severe asthma with fungal sensitization. These findings reinforce the importance of innate immunity in the pathogenesis of different forms of aspergillosis.

The inherent resistance to diseases caused by Aspergillus fumigatus suggests the occurrence of regulatory mechanisms that provide the host with adequate defense without necessarily eliminating the fungus or causing unacceptable levels of host damage. In this study, we show that a division of labor occurs between functionally distinct regulatory T cells (Treg) that are coordinately activated by a CD28/B-7-dependent costimulatory pathway after exposure of mice to Aspergillus conidia. Early in infection, inflammation is controlled by the expansion, activation and local recruitment of CD4+CD25+ Treg capable of suppressing neutrophils through the combined actions of IL-10 and CTLA-4 on indoleamine 2,3-dioxygenase. The levels of IFN-γ produced in this early phase set the subsequent adaptive stage by conditioning the indoleamine 2,3-dioxygenase-dependent tolerogenic program of dendritic cells and the subsequent activation and expansion of tolerogenic Treg, which produce IL-10 and TGF-β, inhibit Th2 cells, and prevent allergy to the fungus. The coordinate activation of Treg may, however, be subverted by the fungus, as germinating conidia are capable of interfering with anti-inflammatory and tolerogenic Treg programs. Thus, regulation is an essential component of the host response in infection and allergy to the fungus, and its manipulation may allow the pathogen to overcome host resistance and promote disease.

IntroductionInvasive aspergillosis (IA) is the leading cause of both nosocomial pneumonia and death in recipients of allogeneic hematopoietic stem cell (HSC) transplants given myeloablative 1,2 or nonmyeloablative 3 conditioning. Despite advances in early diagnosis 4 and new antifungal agents, 2 the majority of cases of IA remain undiagnosed and untreated at the individual's death. The most important risk factor for IA has historically been neutropenia. However, recent studies on the epidemiology of IA in recipients of an HSC transplant indicate a reduced neutropenia-related infection and an increased "late-onset" infection, in concomitance with the occurrence of graft-versus-host disease. 5,6 These findings, together with the occurrence in nonneutropenic patients, 2,7 attest to the importance of specific defects in both the innate and adaptive immune effector mechanisms in the pathogenesis of the disease. [8][9][10][11][12][13] The recent evidence that, in healthy individuals and in patients surviving IA, a significant antigen-specific proliferation of interferon ␥ (IFN-␥)-producing T cells occurred 14 confirms the crucial role of a T helper (T H )1 reactivity in the control of infection. 9,10,15 Dendritic cells (DCs) orchestrate the overall antifungal immune resistance in the lungs. 10,16,17 A dense network of DCs has been described in the respiratory tracts. 18 The evidence that pulmonary DCs, through production of interleukin 10 (IL-10), mediate unresponsiveness to respiratory antigens, 19 suggests that the ability of DCs to instruct the appropriate T-cell responses to the invading pathogens may be affected by local immunoregulatory events. In the case of Aspergillus, by using distinct pattern recognition receptors, including Toll-like receptors (TLRs), murine pulmonary DCs were able to finely discriminate between conidia and hyphae of Aspergillus in terms of induction of adaptive T H responses. 16,17 A protective T H 1-mediated resistance was induced on vaccination with Aspergillus antigens and the TLR-9 ligand CpG oligodeoxynucleotide as adjuvant. 17 These results suggest that the proper manipulation of DC functioning in vivo may translate into beneficial effects in fungal infections.Recent evidence suggests the utility of DCs pulsed with Candida albicans in adoptive transfer experiments. The ability of Candida-pulsed DCs to prime for T H 1-and T H 2-cell activation correlated with the occurrence of resistance and susceptibility to the fungus. 20 In addition, transfecting DCs with fungal RNA was also an effective way to induce antifungal protective immunity in vivo. 21 In the current study we assessed the utility of Aspergilluspulsed DCs in conferring antifungal resistance in vivo. Ex vivo DCs were pulsed with conidia, hyphae, or fungal RNA and adoptively transferred into mice that were recipients of allogeneic HSC transplants. 22 Parameters of infection and immunity were then evaluated on the infection with the fungus. Materials and methods MiceFemale, 8-to 10-week-old inbred BALB/c, C57BL6, and C3H/HeJ m...

Reactivation of latent human cytomegalovirus (HCMV) following allogeneic transplantation is a major cause of morbidity and mortality and predisposes to severe complications, including superinfection by Aspergillus species (spp). Antimicrobial polypeptides, including defensins and mannan-binding lectin, are known to block viral fusion by cross-linking sugars on cell surface. Pentraxin 3 (PTX3), a member of the long pentraxin family, successfully restored antifungal immunity in experimental hematopoietic transplantation. We assessed here whether PTX3 binds HCMV and murine virus (MCMV) and the impact on viral infectivity and superinfection in vivo. We found that PTX3 bound both viruses, reduced viral entry and infectivity in vitro, and protected from MCMV primary infection and reactivation as well IntroductionHuman cytomegalovirus (HCMV), a member of the Herpesviridae family, is a ubiquitous opportunistic pathogen that has an intimate lifelong relationship with its human host and establishes latency after clearance of primary infection. 1-3 Reactivation of latent virus following allogeneic transplantation immune responses results in progressive tissue damage manifesting as overt HCMV disease or complications of this infection, including acute and chronic graft rejection, graft-versus-host disease, and superinfection by other viruses, bacteria, and fungi, particularly Aspergillus species (spp). 3 Efforts have focused on the development of adoptive immunotherapeutic strategies to hasten host immune reconstruction, and cellular immunotherapy appears to be an attractive approach. 4,5 The immune control of murine CMV (MCMV) infection requires elements from both innate and adaptive immune systems. [6][7][8] Through the participation of members of the Toll-like receptors (TLRs) 9-11 and interferon (IFN) regulatory factor 3 (IRF) families, IRF3 in particular, 12,13 MCMV induces early dendritic cell (DC)-dependent type I IFN and interleukin-12 (IL-12) responses essential for mouse resistance to MCMV. 10,11,[14][15][16] The TLR9/ MyD88 signaling pathway mediates antiviral cytokine responses by plasmacytoid DCs (pDCs) that, through their unique capacity to secrete IFN-␣, and to a lesser extent IL-12 and other innate cytokines, are a cornerstone in the initiation of both innate and adaptive immune responses to MCMV. [15][16][17][18][19] However, conventional CD11b ϩ DCs also produce IFN-␣ independently of TLR9 and MyD88. 10,20 In addition to directly interfering with viral replication through ubiquitous cellular mechanisms, IFN-␣ controls natural killer (NK) cell cytotoxic activity 15 and regulates T-cell functions by activating classical DCs to more efficiently present antigens (Ags). 15 IL-12 and IL-18 secretion are instead required to prime a strong NK cell-dependent IFN-␥ response, 17,21,22 a process that is essential to counteract MCMV infection in the liver, in contrast to a perforin-dependent mechanism in the spleen. 23 Pentraxin 3 (PTX3) is a member of a superfamily of conserved proteins characterized by a cyclic mul...

By mediating tryptophan catabolism, the enzyme indoleamine 2,3-dioxygenase (IDO) has a complex role in immunoregulation in infection, pregnancy, autoimmunity, transplantation, and neoplasia. We hypothesized that IDO might affect the outcome of the infection in mice infected with Candida albicans by virtue of its potent regulatory effects on inflammatory and T cell responses. IDO expression was examined in mice challenged with the fungus along with the consequences of its blockade by in vivo treatment with an enzyme inhibitor. We found that IDO activity was induced at sites of infection as well as in dendritic cells and effector neutrophils via IFN-γ- and CTLA-4-dependent mechanisms. IDO inhibition greatly exacerbated infection and associated inflammatory pathology as a result of deregulated innate and adaptive/regulatory immune responses. However, a role for tryptophan catabolism was also demonstrated in a fungus-autonomous fashion; its blockade in vitro promoted yeast-to-hyphal transition. These results provide novel mechanistic insights into complex events that, occurring at the fungus/pathogen interface, relate to the dynamics of host adaptation to the fungus. The production of IFN-γ may be squarely placed at this interface, where IDO activation probably exerts a fine control over fungal morphology as well as inflammatory and adaptive antifungal responses.