Lung Immunopathology Research Articles

Severe acute respiratory syndrome-associated coronavirus vaccines formulated with delta inulin adjuvants provide enhanced protection while ameliorating lung eosinophilic immunopathology.

Although the severe acute respiratory syndrome-associated coronavirus (SARS-CoV) epidemic was controlled by nonvaccine measures, coronaviruses remain a major threat to human health. The design of optimal coronavirus vaccines therefore remains a priority. Such vaccines present major challenges: coronavirus immunity often wanes rapidly, individuals needing to be protected include the elderly, and vaccines may exacerbate rather than prevent coronavirus lung immunopathology. To address these issues, we compared in a murine model a range of recombinant spike protein or inactivated whole-virus vaccine candidates alone or adjuvanted with either alum, CpG, or Advax, a new delta inulin-based polysaccharide adjuvant. While all vaccines protected against lethal infection, addition of adjuvant significantly increased serum neutralizing-antibody titers and reduced lung virus titers on day 3 postchallenge. Whereas unadjuvanted or alum-formulated vaccines were associated with significantly increased lung eosinophilic immunopathology on day 6 postchallenge, this was not seen in mice immunized with vaccines formulated with delta inulin adjuvant. Protection against eosinophilic immunopathology by vaccines containing delta inulin adjuvants correlated better with enhanced T-cell gamma interferon (IFN-γ) recall responses rather than reduced interleukin-4 (IL-4) responses, suggesting that immunopathology predominantly reflects an inadequate vaccine-induced Th1 response. This study highlights the critical importance for development of effective and safe coronavirus vaccines of selection of adjuvants based on the ability to induce durable IFN-γ responses. Coronaviruses such as SARS-CoV and Middle East respiratory syndrome-associated coronavirus (MERS-CoV) cause high case fatality rates and remain major human public health threats, creating a need for effective vaccines. While coronavirus antigens that induce protective neutralizing antibodies have been identified, coronavirus vaccines present a unique problem in that immunized individuals when infected by virus can develop lung eosinophilic pathology, a problem that is further exacerbated by the formulation of SARS-CoV vaccines with alum adjuvants. This study shows that formulation of SARS-CoV spike protein or inactivated whole-virus vaccines with novel delta inulin-based polysaccharide adjuvants enhances neutralizing-antibody titers and protection against clinical disease but at the same time also protects against development of lung eosinophilic immunopathology. It also shows that immunity achieved with delta inulin adjuvants is long-lived, thereby overcoming the natural tendency for rapidly waning coronavirus immunity. Thus, delta inulin adjuvants may offer a unique ability to develop safer and more effective coronavirus vaccines.

Recombinant influenza virus expressing a fusion protein neutralizing epitope of respiratory syncytial virus (RSV) confers protection without vaccine-enhanced RSV disease

Respiratory syncytial virus (RSV) is the leading cause of viral bronchiolitis in both children and the elderly. There is no vaccine available for the prevention of RSV infection. Here, we generated recombinant influenza virus (PR8/RSV.HA-F) expressing an RSV F243–294 neutralizing epitope in the hemagglutinin (HA) as a chimeric protein. Neutralizing antibodies specific for both RSV and influenza virus were induced by a single intranasal immunization of mice with PR8/RSV.HA-F. Mice that were immunized with PR8/RSV.HA-F were protected against RSV infection comparable with live RSV as evidenced by significant reduction of RSV lung viral loads, as well as the absence of lung eosinophilia and RSV-specific cellular immune responses. In contrast, formalin-inactivated RSV-immunized mice showed severe disease and high cellular immune responses in lungs after RSV infection. These findings support a concept that recombinant influenza virus carrying the RSV F243–294 neutralizing epitope can be developed as a promising RSV vaccine candidate which induces protective neutralizing antibodies but avoids lung immunopathology.

Antigen-pulsed bone marrow-derived and pulmonary dendritic cells promote Th2 cell responses and immunopathology in lungs during the pathogenesis of murine Mycoplasma pneumonia.

Mycoplasmas are a common cause of pneumonia in humans and animals, and attempts to create vaccines have not only failed to generate protective host responses, but they have exacerbated the disease. Mycoplasma pulmonis causes a chronic inflammatory lung disease resulting from a persistent infection, similar to other mycoplasma respiratory diseases. Using this model, Th1 subsets promote resistance to mycoplasma disease and infection, whereas Th2 responses contribute to immunopathology. The purpose of the present study was to evaluate the capacity of cytokine-differentiated dendritic cell (DC) populations to influence the generation of protective and/or pathologic immune responses during M. pulmonis respiratory disease in BALB/c mice. We hypothesized that intratracheal inoculation of mycoplasma Ag-pulsed bone marrow-derived DCs could result in the generation of protective T cell responses during mycoplasma infection. However, intratracheal inoculation (priming) of mice with Ag-pulsed DCs resulted in enhanced pathology in the recipient mice when challenged with mycoplasma. Inoculation of immunodeficient SCID mice with Ag-pulsed DCs demonstrated that this effect was dependent on lymphocyte responses. Similar results were observed when mice were primed with Ag-pulsed pulmonary, but not splenic, DCs. Lymphocytes generated in uninfected mice after the transfer of either Ag-pulsed bone marrow-derived DCs or pulmonary DCs were shown to be IL-13(+) Th2 cells, known to be associated with immunopathology. Thus, resident pulmonary DCs most likely promote the development of immunopathology in mycoplasma disease through the generation of mycoplasma-specific Th2 responses. Vaccination strategies that disrupt or bypass this process could potentially result in a more effective vaccination.

Lysosomal phospholipase A2: A novel player in host immunity to Mycobacterium tuberculosis

Phospholipases catalyze the cleavage of membrane phospholipids into smaller bioactive molecules. The lysosomal phospholipase A2 (LPLA2 ) is specifically expressed in macrophages. LPLA2 gene deletion in mice causes lysosomal phospholipid accumulation in tissue macrophages leading to phospholipidosis. This phenotype becomes most prominent in alveolar macrophages where LPLA2 contributes to surfactant phospholipid degradation. High expression of LPLA2 in alveolar macrophages prompted us to investigate its role in host immunity against the respiratory pathogen Mycobacterium tuberculosis, the causative agent of tuberculosis. Here we report that adaptive immune responses to M. tuberculosis were impaired in LPLA2 deficient mice. Upon aerosol infection with M. tuberculosis, LPLA2 deficient mice showed enhanced mycobacterial counts but less lung immunopathology and pulmonary inflammatory responses. Compromised T-cell priming in the lymph nodes was associated with impaired pulmonary T-cell recruitment and activation. Together with reduced Th1 type cytokine production, these results indicate that LPLA2 is indispensable for the induction of adaptive T-cell immunity to M. tuberculosis. Taken together, we identified an unexpected and novel function of a lysosomal phospholipid-degrading enzyme.

TREM2 differential expression is associated with BAL cells that express high levels of ARG1 (VET2P.1044)

Abstract Lung immunopathology is the major cause of influenza induced morbidity and mortality. DAP12 (DNAX-Activating Protein of 12kDa) is a membrane adaptor protein associated with varied surface receptors, and it is known to regulate influenza induced lung immunopathology. DAP12 associated receptor expression and their association with phenotype of bronchoalveolar lavage fluid (BAL) cells during swine influenza virus (SIV) infection in pigs is unknown. Since pig is a suitable large animal model for influenza research, our aim was to understand the effects of zoonotic SIV H1N1 infection on expression of DAP12 associated MDL-1, TREM-1 and TREM-2 receptors in pig BAL cells through qRT-PCR profiling. Results indicated that DAP12, MDL-1 and TREM-1 were constitutively expressed, but TREM-2 was upregulated. Stimulation of uninfected pig BAL cells in vitro using the cytokines IFNgamma and IL4 revealed that upregulation of TREM-2 was associated with enhanced expression of ARG1, but not with high levels of expression of TNFalpha and iNOS. ARG1 is a phenotype of alternatively activated alveolar macrophages that are known to protect lung tissue; hence TREM2 upregulation in pig BAL cells with similar phenotype suggests the possible beneficial role of these molecules in preventing the lung immunopathology. In conclusion, DAP12 and associated receptors are differentially expressed in pig BAL cells and TREM2 appears to have a beneficial role in regulating the lung immunopathology in SIV infection.

Effect of asthma on susceptibility to influenza A virus infection (VIR2P.1029)

Abstract Asthma exacerbations by respiratory infections are a serious health issue and, thus significant effort has been put into understanding the impact of respiratory infections on asthma. However little remains known about how the asthmatic phenotype could influence immune responses against respiratory viral pathogens. Using a mouse model of allergic lung inflammtion, we investigated the susceptibility of asthmatics to influenza A virus challenge. Unexpectedly, asthmatic mice were highly resistant to primary infection with the H1N1 2009 pandemic strain. However, the increased survival rate did not correlate with viral lung titers. Strikingly, asthmatic mice exhibited a significant reduction in cytokine storm, total protein levels and cytotoxicity in the airway; observations that indicate minimal tissue injury. Further, we found that asthmatic mice had significantly increased levels of TGF-β1 and that the heightened resistance of asthmatic mice was abrogated in the absence of TGF-β receptor II. Thus, we conclude that a transient increase in TGF-β expression following an asthma attack is responsible for temporary protection against influenza virus challenge via suppression of lung immunopathology.

Adjunctive Corticosteroid Therapy Improves Lung Immunopathology and Survival During Severe Secondary Pneumococcal Pneumonia in Mice

Secondary bacterial pneumonia is a significant cause of morbidity and mortality during influenza, despite routine use of standard antibiotics. Antibiotic-induced immunopathology associated with bacterial cell wall lysis has been suggested to contribute to these poor outcomes. Using Streptococcus pneumoniae in a well-established murine model of secondary bacterial pneumonia (SBP) following influenza, we stratified disease severity based on pneumococcal load in the lungs via in vivo bioluminescence imaging. Ampicillin treatment cured mice with mild pneumonia but was ineffective against severely pneumonic mice, despite effective bacterial killing. Adjunctive dexamethasone therapy improved ampicillin-induced immunopathology and improved outcomes in mice with severe SBP. However, early dexamethasone therapy during primary influenza infection impaired lung adaptive immunity as manifest by increased viral titers, with an associated loss of its protective functions in SBP. These data support adjunctive clinical use of corticosteroids in severe cases of community-acquired pneumonia.

IL-33 Signaling Regulates Innate and Adaptive Immunity to Cryptococcus neoformans

Susceptibility to progressive infection with the fungus Cryptococcus neoformans is associated with an allergic pattern of lung inflammation, yet the factors that govern this host response are not clearly understood. Using a clinically relevant mouse model of inhalational infection with virulent C. neoformans H99, we demonstrate a role for IL-33-dependent signaling in host immune defense. Infection of BALB/c mice with 10(4) CFU of C. neoformans H99 caused a time-dependent induction of IL-33 with accumulation of type 2 pulmonary innate lymphoid cells and alternatively activated macrophages in the lungs as well as Th2-polarized CD4(+) T cells in draining lymph nodes. IL-33R subunit T1/ST2-deficient (T1/ST2(-/-)) mice infected with C. neoformans H99 had improved survival with a decreased fungal burden in the lungs, spleen, and brain, compared with wild-type mice. Signaling through T1/ST2 was required for the accumulation and early production of IL-5 and IL-13 by lung type 2 pulmonary innate lymphoid cells. Further analysis of T1/ST2(-/-) mice revealed increased fungicidal exudate macrophages in the lungs and decreased C. neoformans-specific Th2 cells in the mediastinal lymph nodes. T1/ST2 deficiency also diminished goblet cell hyperplasia, mucus hypersecretion, bronchoalveolar lavage eosinophilia, alternative activation of macrophages, and serum IgE. These observations demonstrate that IL-33-dependent signaling contributes to the expansion of innate type 2 immunity and subsequent Th2-biased lung immunopathology that facilitates C. neoformans growth and dissemination.

81 : Regulation of CD4 T cell derived IL-10 during influenza infection

Effector CD4 T cells play an important role in limiting pulmonary inflammation and tissue damage during primary influenza infection by producing the immunosuppressive cytokine IL-10. The regulation of IL-10 in these cells is critical for efficient viral clearance and reducing lung immunopathology. Evidence suggests that IL-27 plays a role in eliciting IL-10 from CD4 T cells. However, the molecular signals required to regulate IL-10 in CD4 T cells during respiratory viral infection are not well defined. Data from our in vivo studies using IL-10 GFP reporter mice show that CD4 T cells in a secondary response to influenza express less IL-10 than during a primary infection. Through in vitro studies we demonstrate that the IL-27 receptor modulates IL-10 expression through down regulation of its gp130 subunit. The resulting loss of IL-27 responsiveness in memory type CD4 T cells leads to decreased IL-10 expression in the secondary response. Together our data identify the IL-27 receptor as a critical determinant of IL-10 expression in CD4 T cells. Our study provides new insight into the mechanisms that balance effective immunity and immunopathology during respiratory disease.

Interleukin‐6 limits influenza‐induced inflammation and protects against fatal lung pathology

Balancing the generation of immune responses capable of controlling virus replication with those causing immunopathology is critical for the survival of the host and resolution of influenza-induced inflammation. Based on the capacity of interleukin-6 (IL-6) to govern both optimal T-cell responses and inflammatory resolution, we hypothesised that IL-6 plays an important role in maintaining this balance. Comparison of innate and adaptive immune responses in influenza-infected wild-type control and IL-6-deficient mice revealed striking differences in virus clearance, lung immunopathology and generation of heterosubtypic immunity. Mice lacking IL-6 displayed a profound defect in their ability to mount an anti-viral T-cell response. Failure to adequately control virus was further associated with an enhanced infiltration of inflammatory monocytes into the lung and an elevated production of the pro-inflammatory cytokines, IFN-α and TNF-α. These events were associated with severe lung damage, characterised by profound vascular leakage and death. Our data highlight an essential role for IL-6 in orchestrating anti-viral immunity through an ability to limit inflammation, promote protective adaptive immune responses and prevent fatal immunopathology.

Marked Improvement of Severe Lung Immunopathology by Influenza-Associated Pneumococcal Superinfection Requires the Control of Both Bacterial Replication and Host Immune Responses

Bacterial superinfection and associated lung immunopathology are major contributors to hospitalizations and mortality after influenza. However, the underlying mechanisms and effective intervention strategies remain poorly defined. By using a model of influenza and pneumococcal superinfection, we found that dual-infected animals experienced rapid weight loss and succumbed to infection. Bacterial outgrowth, dysregulated cytokines, including keratinocyte-derived chemokine and macrophage inflammatory protein 2, and severe lung neutrophilia and immunopathology were linked to the poor clinical outcome. Invivo neutralization of highly induced macrophage inflammatory protein 2 did not affect clinical outcome, bacterial loads, or lung immunopathology. On the other hand, invivo neutrophil depletion did not alter the clinical outcome and bacterial burden, although it moderately improved lung immunopathology. Treatment with a bacteriostatic antibiotic, azithromycin, alone significantly improved clinical outcome and bacterial clearance, but failed to reduce lung immunopathology. In comparison, treatment with a global inflammation inhibitor, dexamethasone, alone failed to alter clinical outcome, bacterial infection, and immunopathology, despite its moderate reducing effects on neutrophilic and cytokine responses. In contrast, combined treatment with both azithromycin and dexamethasone best improved clinical outcome, bacterial clearance, lung cellular and cytokine responses, and immunopathology. Our study suggests that marked improvement of clinical outcome and lung immunopathology caused by bacterial superinfection requires the control of both bacterial infection and aberrant host immune responses. Our findings hold implications in clinical management for influenza-associated bacterial superinfections.

A mycobacteriophage‐derived trehalose‐6,6'‐dimycolate‐binding peptide containing both antimycobacterial and anti‐inflammatory abilities

Bacteriophages, the viruses of eubacteria, have developed unique mechanisms to interact with their host bacteria. They have been viewed as potential antibacterial therapeutics. Mycobacteriophage-derived compounds may interact with Mycobacterium tuberculosis (MTB) and/or its components, such as the cord factor, trehalose-6,6'-dimycolate (TDM), which is the most abundant glycolipid produced on the surface of MTB. TDM emulsion injected intravenously into mice induces lung immunopathology that mimics many aspects of MTB infection. Thus, TDM is an important target for anti-MTB agent development. On the basis of genomics information of mycobacteriophages, 200 peptides were synthesized. Their effects on MTB, their interactions with TDM, and anti-inflammatory activities were tested. One of them (PK34) showed MTB-killing activity with a minimal inhibitory concentration of 50 μg/ml and TDM-binding ability. In a mouse model, PK34 showed comparable ability to clear MTB as rifampin did in vivo. It also exerted strong activity to inhibit MTB or TDM-induced inflammation in vivo. PK34 significantly inhibited inflammatory cytokines secretions by inactivating MAPK and PKB signals while it maintained certain proinflammatory cytokine production. It is possible to prospect for TDM-binding and/or anti-MTB peptides by mining the mycobacteriophages genome. In addition to its direct MTB-killing ability, PK34 might be a useful adjunct in the treatment of granulomatous inflammation occurring during mycobacterial infection or a template for developing antituberculosis (TB) agents because of its immunoregulative effects. As a TDM-binding peptide, PK34 may be a promising tool to study TDM's interactions with corresponding receptors and signal pathways.

Efficacy and safety of an intranasal virosomal respiratory syncytial virus vaccine adjuvanted with monophosphoryl lipid A in mice and cotton rats

Respiratory syncytial virus infection remains a serious health problem, not only in infants but also in immunocompromised adults and the elderly. An effective and safe vaccine is not available due to several obstacles: non-replicating RSV vaccines may prime for excess Th2-type responses and enhanced respiratory disease (ERD) upon natural RSV infection of vaccine recipients. We previously found that inclusion of the Toll-like receptor 4 (TLR4) ligand monophosphoryl lipid A (MPLA) in reconstituted RSV membranes (virosomes) potentiates vaccine-induced immunity and skews immune responses toward a Th1-phenotype, without priming for ERD. As mucosal immunization is an attractive approach for induction of RSV-specific systemic and mucosal antibody responses and TLR ligands could potentiate such responses, we explored the efficacy and safety of RSV-MPLA virosomes administered intranasally (IN) to mice and cotton rats. In mice, we found that incorporation of MPLA in IN-administered RSV virosomes increased both systemic IgG and local secretory-IgA (S-IgA) antibody levels and resulted in significantly reduced lung viral titers upon live virus challenge. Also, RSV MPLA virosomes induced more Th1–skewed responses compared to responses induced by FI-RSV. Antibody responses and Th1/Th2-cytokine responses induced by RSV-MPLA virosomes were comparable to those induced by live RSV infection. By comparison, formalin-inactivated RSV (FI-RSV) induced serum IgG that inhibited viral shedding upon challenge, but also induced Th2-skewed responses. In cotton rats, similar effects of incorporation of MPLA in virosomes were observed with respect to induction of systemic antibodies and inhibition of lung viral shedding upon challenge, but mucosal sS-IgA responses were only moderately enhanced. Importantly, IN immunization with RSV-MPLA virosomes, like live virus infection, did not lead to any signs of ERD upon live virus challenge of vaccinated animals, whereas IM immunization with FI-RSV did induce severe lung immunopathology under otherwise comparable conditions. Taken together, these data show that mucosally administered RSV-MPLA virosomes hold promise for a safe and effective vaccine against RSV.

The signalling imprints of nanoparticle uptake by bone marrow derived dendritic cells

Nanoparticles (NP) possess remarkable adjuvant and carrier capacity, therefore are used in the development of various vaccine formulations. Our previous studies demonstrated that inert non-toxic 40–50nm polystyrene NP (PS-NP) can promote strong CD8 T cell and antibody responses to the antigen, in the absence of observable inflammatory responses. Furthermore, instillation of PS-NP inhibited the development of allergic airway inflammation by induction of an immunological imprint via modulation of dendritic cell (DC) function without inducing oxidative stress in the lungs in mice. This is in contrast to many studies which show that a variety of ambient and man-made NP promote lung immunopathology, raising concerns generally about the safe use of NPs in biomedicine. Most NPs are capable of inducing inflammatory pathways in DC largely mediated by signalling via the extracellular signal-regulated kinase 1/2 (ERK). Herein, we investigate whether PS-NPs also activate ERK in DC in vitro. Our data show that PS-NP do not induce ERK activation in two different types of bone marrow derived (BM) DC cultures (expanded with GM-CSF or with GM-CSF together with IL-4). The absence of such signalling was not due to lack of PS-NP uptake by BM-DC as confirmed by confocal microscopy and flow cytometry. The process of NP uptake by DC usually initiates ERK signalling, suggesting an unusual uptake pathway may be engaged by PS-NPs. Indeed, data herein showns that uptake of PS-NP by BM-DC was substantially inhibited by phorbol myristate acetate (PMA) but not cytochalasin D (CCD), suggesting an uptake pathway utilising caveole for PS-NP. Together these data show that BM-DC take up PS-NP via a caveole-dependent pathway which does not trigger ERK signalling which may explain their efficient uptake by DC, without the concomitant activation of conventional inflammatory pathways.

An adjuvanted respiratory syncytial virus fusion protein induces protection in aged BALB/c mice

BackgroundRespiratory Syncytial Virus (RSV) causes significant disease in the elderly, in part, because immunosenescence impairs protective immune responses to infection in this population. Despite previous and current efforts, there is no RSV vaccine currently licensed in infants or elderly adults. Adjuvanted RSV subunit vaccines have the potential to boost waning immune responses and reduce the burden of RSV disease in the elderly population.ResultsWe used an aged BALB/c mouse model to evaluate immune responses to RSV Fusion (F) protein in the absence and presence of an alum adjuvant. We demonstrate that aged BALB/c mice immunized with alum-adjuvanted RSV F protein had significantly reduced lung viral titers at day 4 following challenge with wild-type (wt) RSV. Serum neutralizing antibody titers measured on day 27 correlated with protection in both young and aged vaccinated mice, although the magnitude of antibody titers was lower in aged mice. Unlike young mice, in aged mice, alum-adjuvanted RSV F did not induce lung TH2-type cytokines or eosinophil infiltration compared to non-adjuvanted F protein following wt RSV challenge.ConclusionOur studies demonstrate that neutralizing anti-RSV antibody titers correlate with protection in both young and aged BALB/c mice vaccinated with RSV F protein vaccines. The F + alum formulation mediated greater protection compared to the non-adjuvanted F protein in both young and aged mice. However, while alum can boost F-specific antibody responses in aged mice, it does not completely overcome the reduced ability of a senescent immune system to respond to the RSV F antigen. Thus, our data suggest that a stronger adjuvant may be required for the prevention of RSV disease in immunosenescent populations, to achieve the appropriate balance of protective neutralizing antibodies and effective TH1-type cytokine response along with minimal lung immunopathology.

The β-Glucan Receptor Dectin-1 Promotes Lung Immunopathology during Fungal Allergy via IL-22

Sensitization to fungi, such as the mold Aspergillus fumigatus, is increasingly becoming linked with asthma severity. We have previously shown that lung responses generated via the β-glucan receptor Dectin-1 are required for lung defense during acute, invasive A. fumigatus infection. Unexpectedly, in an allergic model of chronic lung exposure to live A. fumigatus conidia, β-glucan recognition via Dectin-1 led to the induction of multiple proallergic (Muc5ac, Clca3, CCL17, CCL22, and IL-33) and proinflammatory (IL-1β and CXCL1) mediators that compromised lung function. Attenuated proallergic and proinflammatory responses in the absence of Dectin-1 were not associated with changes in Ido (IDO), Il12p35/Ebi3 (IL-35), IL-10, or TGF-β levels. Assessment of Th responses demonstrated that purified lung CD4(+) T cells produced IL-4, IL-13, IFN-γ, and IL-17A, but not IL-22, in a Dectin-1-dependent manner. In contrast, we observed robust, Dectin-1-dependent IL-22 production by unfractionated lung digest cells. Intriguingly, the absence of IL-22 alone mimicked the attenuated proallergic and proinflammatory responses observed in the absence of Dectin-1, suggesting that Dectin-1-mediated IL-22 production potentiated responses that led to decrements in lung function. To this end, neutralization of IL-22 improved lung function in normal mice. Collectively, these results indicate that the β-glucan receptor Dectin-1 contributes to lung inflammation and immunopathology associated with persistent fungal exposure via the production of IL-22.

I12 Interferons: Overlooked regulators of innate and adaptive immune responses

Type I IFNs, that include the IFN- α s and IFN-s, are critical effectors of the host antiviral immune response. Much attention has focused on their direct antiviral effects against viruses in infected tissues, examining specific signaling cascades and effector molecules induced by these IFNs that mediate virus inhibition and protection from infection. To date, their activities as modulators of innate and adaptive immune responses have been largely overlooked, perhaps contributing to their limited clinical application as therapeutics for acute viral infections. Our early studies using IFN-s -/- mice revealed diverse consequences of IFN-s for both the innate and adaptive arms of immunity: altered splenic architecture in IFN-s -/- mice and a reduction in resident macrophages; a defect in B cell maturation; circulating IgM-, Mac-1-, and Gr-1-positive cells are also substantially decreased in IFN-s -/- mice. The decrease in the numbers of circulating macrophages and granulocytes likely reflects defective maturation of primitive BM hematopoiesis in mice, shown by a reduction in CFU-GM. Viewed together, these data indicate that IFN-s is required during different stages of maturation in the development of the immune system. Constant immunosurveillance in the host requires that lymphocytes traffic through lymph nodes (LNs) to sample antigen. pDCs and lymphocytes use similar mechanisms for retention within LNs and we provided evidence that these processes are influenced by IFN-s, even in the absence of viral infection, implicating IFN-s in affecting critical APC-T cell interactions that determine a robust immune response to pathogen infection. Most recently, we have conducted studies in mice infected intranasally with different strains of influenza A virus and show that clearance of virus mediated by IFN-s is associated with polarization of CD4 and CD8 T cells to a Th1 phenotype, increased DC infiltrates into the lungs and dLNs of infected mice, reduced eosinophilia, and a reduction in a novel antigen presenting cell population associated with type 2 immunity and lung immunopathology. Viewed in conjunction with data that suggest an adjuvant role for IFNs in vaccination, accumulating evidence suggests that the immunomodulatory activities of type I IFNs contribute significantly to their antiviral effectiveness.

Immunopathology in influenza virus infection: Uncoupling the friend from foe

Influenza epidemics and pandemics cause significant morbidity and mortality worldwide associated with severe immunopathology in the lung, and the mechanisms of such immunopathogenesis still remain poorly understood. While human studies help to understand influenza immunopathology, they provide only limited mechanistic information. On the other hand, recent studies using experimental animal models have significantly enhanced our understanding of the complex mechanisms involved in the immunopathogenesis during primary influenza or influenza-associated bacterial superinfection. This includes the involvement of acute inflammatory responses (macrophages, neutrophils, dendritic cells, toll-like receptors, cytokines, chemokines), CD4 and CD8 T cells, tissue remodeling processes, and contribution of bacterial superinfection. In particular, progress has been made in uncoupling the mechanisms that are involved in both anti-viral host defense and in immunopathogenesis from those that solely contribute to lung immunopathology. Uncoupling such events will facilitate the discovery of new intervention strategies to treat pulmonary immunopathology associated with influenza infection.

Critical Role of Natural Killer Cells in Lung Immunopathology During Influenza Infection in Mice

Influenza viral infection results in excessive pulmonary inflammation that has been linked to the damage caused by immune responses and viral replication. The multifunctional cytokine interleukin (IL-15), influences the proliferation and maintenance of immune cells such as CD8(+) T cells and natural killer (NK) cells. Here we show that IL-15(-/-) mice are protected from lethal influenza infection. Irrespective of the mouse strains, the protection observed was linked to the lack of NK cells. Increased survival in the IL-15(-/-) or NK1.1(+) cell-depleted wild-type mice was associated with significantly lower lung lesions as well as decreased mononuclear cells and neutrophils in the airway lumen. Levels of interleukin 10 were significantly higher and levels of proinflammatory cytokines, including interleukin 6 and interleukin 12, were significantly lower in the bronchoalveolar lavage fluid from IL-15(-/-) and NK1.1(+) cell-depleted wild-type mice than in that from control mice. Our data suggest that NK cells significantly augment pulmonary inflammation, contributing to the pathogenesis of influenza infection.

Pulmonary immunopathology mediated by proteolytic processing of TNF-α by antiviral CD8+ T cells (105.11)

Abstract Infection with influenza virus evokes a potent CD8+ T cell response that may contribute to lung immunopathology. Recognition of alveolar anitgen by CD8+ T cells results in significant injury that is critically dependent on TNF-α expressed by CD8+ T cells and is largely mediated by the secretion of chemokines by alveolar epithelial cells and the subsequent pulmonary infiltration. TNF-α is expressed as a transmembrane (tm) protein that is proteolytically processed by TNF-α converting enzyme (TACE) to produce soluble (s) TNF-α. To determine the roles of tmTNF-α and sTNF-α in CD8+ T cell mediated lung injury, we generated CD8+ T cells that express either a noncleavable tmTNF-α or have a deletion in the catalytic domain of TACE. Both T cells are impaired in sTNF-α production but exhibited no defect in other effector functions. In vitro analysis indicated that proteolytic processing of tmTNF-α to sTNF-α was required to induce alveolar epithelial cell expression of macrophage inflammatory protein-2. Furthermore, inhibition of proteolytic processing of TNF-α resulted in reduced induction of alveolar epithelial cell chemokines, less neutrophil infiltration and milder lung injury, though these T cells exhibited no defect in antiviral activity in vivo. This indicates that proteolytic processing of tmTNF-α to sTNF-α is a critical event in the initiation of CD8+ T cell mediated lung injury and represents a potential mechanism of regulation of immunopathology during an influenza infection.