The WI-1 adhesin blocks phagocyte TNF-alpha production, imparting pathogenicity on Blastomyces dermatitidis

The WI-1 adhesin is indispensable for pathogenicity of Blastomyces dermatitidis and is thought to promote pulmonary infection by fixing yeast to lung tissue and cells. Recent findings suggest that WI-1 confers pathogenicity by mechanisms in addition to adherence. Here, we investigated whether WI-1 modulates host immunity by altering production of pro-inflammatory cytokines. Production of TNF-alpha in lung alveolar fluids of mice infected with B. dermatitidis was severalfold higher for WI-1 knockout yeast compared with wild-type yeast, and in vitro coculture of unseparated lung cells with these isogenic yeast disclosed similar differences. Upon coculture with purified macrophages and neutrophils, wild-type yeast blocked TNF-alpha production, yet WI-1 knockout yeast stimulated production. Coating knockout yeast with purified WI-1 converted them from stimulating TNF-alpha production to inhibiting production. Addition of purified WI-1 into stimulated phagocyte cultures led to concentration-dependent inhibition of TNF-alpha production. Neutralization of TNF-alpha in vivo exacerbated experimental pulmonary infection, particularly for the nonpathogenic WI-1 knockout yeast. Inducing increased TNF-alpha levels in the lung by adenovirus-vectored gene therapy controlled infection with wild-type yeast. Thus, the WI-1 adhesin on yeast modulates host immunity through blocking TNF-alpha production by phagocytes, which fosters progression of pulmonary infection.     

Interleukin-22 is produced by invariant natural killer T lymphocytes during influenza A virus infection: potential role in protection against lung epithelial damages

Invariant natural killer T (iNKT) cells are non-conventional lipid-reactive αβ T lymphocytes that play a key role in host responses during viral infections, in particular through the swift production of cytokines. Their beneficial role during experimental influenza A virus (IAV) infection has recently been proposed, although the mechanisms involved remain elusive. Here we show that during in vivo IAV infection, mouse pulmonary iNKT cells produce IFN-γ and IL-22, a Th17-related cytokine critical in mucosal immunity. Although permissive to viral replication, IL-22 production by iNKT cells is not due to IAV infection per se of these cells but is indirectly mediated by IAV-infected dendritic cells (DCs). We show that activation of the viral RNA sensors TLR7 and RIG-I in DCs is important for triggering IL-22 secretion by iNKT cells, whereas the NOD-like receptors NOD2 and NLRP3 are dispensable. Invariant NKT cells respond to IL-1β and IL-23 provided by infected DCs independently of the CD1d molecule to release IL-22. In vitro, IL-22 protects IAV-infected airway epithelial cells against mortality but has no role on viral replication. Finally, during early IAV infection, IL-22 plays a positive role in the control of lung epithelial damages. Overall, IAV infection of DCs activates iNKT cells, providing a rapid source of IL-22 that might be beneficial to preserve the lung epithelium integrity.     

Early production of IL-17 protects against acute pulmonary Pseudomonas aeruginosa infection in mice

Interleukin-17 (IL-17) is involved in protection against extracellular bacteria. However, IL-17 is likely to be deleterious to a host with chronic pulmonary Pseudomonas aeruginosa infection. The role of IL-17 during acute pulmonary P. aeruginosa infection remains unknown. Here, we evaluated the role that IL-17 plays in acute pulmonary P. aeruginosa infection and the source of the interleukin. The production of IL-17 increased rapidly after acute pulmonary P. aeruginosa infection in mice. We subsequently examined the role of IL-17 in acute infection and found 100 times more bacteria in the bronchoalveolar lavage fluid of mice treated with an IL-17-neutralizing antibody compared with the IgG(2a) -treated mice after 16 h of infection. The main infiltrating cells in the anti-IL-17-treated mice were lymphocytes rather than neutrophils. Consistently, more tissue damage and pathological changes in the lung were observed in the anti-IL-17-treated mice. We also found that Th17 cells are one of the sources of IL-17. We conclude that the early production of IL-17 plays a protective role during acute pulmonary P. aeruginosa infection in mice and that Th17 cells are one of the sources of IL-17 during acute pulmonary P. aeruginosa infection. Altogether, IL-17 and Th17 cells contribute to the pathogenesis of acute pulmonary P. aeruginosa infection in vivo.     

Aryl hydrocarbon receptor activation during influenza virus infection unveils a novel pathway of IFN-gamma production by phagocytic cells

The contribution of environmental factors is important as we consider reasons that underlie differential susceptibility to influenza virus. Aryl hydrocarbon receptor (AhR) activation by the pollutant dioxin during influenza virus infection decreases survival, which correlates with a 4-fold increase in pulmonary IFN-gamma levels. We report here that the majority of IFN-gamma-producing cells in the lung are neutrophils and macrophages not lymphocytes, and elevated IFN-gamma is associated with increased pulmonary inducible NO synthase (iNOS) levels. Moreover, we show that even in the absence of dioxin, infection with influenza virus elicits IFN-gamma production by B cells, gammadelta T cells, CD11c(+) cells, macrophages and neutrophils, as well as CD3(+) and NK1.1(+) cells in the lung. Bone marrow chimeric mice reveal that AhR-mediated events external to hemopoietic cells direct dioxin-enhanced IFN-gamma production. We also show that AhR-mediated increases in IFN-gamma are dependent upon iNOS, but elevated iNOS in lung epithelial cells is not driven by AhR-dependent signals from bone marrow-derived cells. Thus, the lung contains important targets of AhR regulation, which likely influence a novel iNOS-mediated mechanism that controls IFN-gamma production by phagocytic cells. This suggests that AhR activation changes the response of lung parenchymal cells, such that regulatory pathways in the lung are cued to respond inappropriately during infection. These findings also imply that environmental factors may contribute to differential susceptibility to influenza virus and other respiratory pathogens.     

Pulmonary immunity to viral infection: adenovirus infection of lung dendritic cells renders T cells nonresponsive to interleukin-2

Adenovirus (Ad) infection has been identified as predisposing hosts to the development of pulmonary disease through unknown mechanisms. Lung dendritic cells (DCs) are vital for initiating pulmonary immune responses; however, the effects of Ad infection on primary lung DC have not been studied. In contrast to the effects on bone marrow- and monocyte-derived DCs, the current study shows that Ad infection of murine BALB/c lung DCs in vitro and in vivo suppresses DC-induced T-cell proliferation. The effect of Ad on DCs was not due to a downregulation of major histocompatibility complex or costimulatory molecules. Analysis of the production of interleukin-12 (IL-12), alpha interferon (IFN-alpha), and IFN-gamma by the Ad-infected DCs shows no significant differences over noninfected control lung DCs. Ad-induced suppression was not due to a deficiency of IL-2 or other DC-secreted factors and was dependent on viral protein synthesis, as UV irradiation of Ad abrogated the suppressive effect. Results suggest that Ad-infected DCs induce T cells to be nonresponsive to IL-2 during primary coculture, as the addition of IL-2 in secondary cultures recovered T-cell proliferation. In vivo studies supported in vitro results showing that Ad infection resulted in lung T cells with decreased proliferative ability. This study demonstrates that Ad infection induces local immunoincompetence by altering DC-T-cell interactions.     

Early IL-17A production helps establish Mycobacterium intracellulare infection in mice

Nontuberculous mycobacteria (NTM) infection is common in patients with structural lung damage. To address how NTM infection is established and causes lung damage, we established an NTM mouse model by intranasal inoculation of clinical isolates of M. intracellulare. During the 39-week course of infection, the bacteria persistently grew in the lung and caused progressive granulomatous and fibrotic lung damage with mortality exceeding 50%. Lung neutrophils were significantly increased at 1 week postinfection, reduced at 2 weeks postinfection and increased again at 39 weeks postinfection. IL-17A was increased in the lungs at 1–2 weeks of infection and reduced at 3 weeks postinfection. Depletion of neutrophils during early (0–2 weeks) and late (32–34 weeks) infection had no effect on mortality or lung damage in chronically infected mice. However, neutralization of IL-17A during early infection significantly reduced bacterial burden, fibrotic lung damage, and mortality in chronically infected mice. Since it is known that IL-17A regulates matrix metalloproteinases (MMPs) and that MMPs contribute to the pathogenesis of pulmonary fibrosis, we determined the levels of MMPs in the lungs of M. intracellulare-infected mice. Interestingly, MMP-3 was significantly reduced by anti-IL-17A neutralizing antibody. Moreover, in vitro data showed that exogenous IL-17A exaggerated the production of MMP-3 by lung epithelial cells upon M. intracellulare infection. Collectively, our findings suggest that early IL-17A production precedes and promotes organized pulmonary M. intracellulare infection in mice, at least in part through MMP-3 production.     

Andes Hantavirus-Infection of a 3D Human Lung Tissue Model Reveals a Late Peak in Progeny Virus Production Followed by Increased Levels of Proinflammatory Cytokines and VEGF-A

Andes virus (ANDV) causes hantavirus pulmonary syndrome (HPS), a severe acute disease with a 40% case fatality rate. Humans are infected via inhalation, and the lungs are severely affected during HPS, but little is known regarding the effects of ANDV-infection of the lung. Using a 3-dimensional air-exposed organotypic human lung tissue model, we analyzed progeny virus production and cytokine-responses after ANDV-infection. After a 7–10 day period of low progeny virus production, a sudden peak in progeny virus levels was observed during approximately one week. This peak in ANDV-production coincided in time with activation of innate immune responses, as shown by induction of type I and III interferons and ISG56. After the peak in ANDV production a low, but stable, level of ANDV progeny was observed until 39 days after infection. Compared to uninfected models, ANDV caused long-term elevated levels of eotaxin-1, IL-6, IL-8, IP-10, and VEGF-A that peaked 20–25 days after infection, i.e., after the observed peak in progeny virus production. Notably, eotaxin-1 was only detected in supernatants from infected models. In conclusion, these findings suggest that ANDV replication in lung tissue elicits a late proinflammatory immune response with possible long-term effects on the local lung cytokine milieu. The change from an innate to a proinflammatory response might be important for the transition from initial asymptomatic infection to severe clinical disease, HPS.     

Dual function of the long pentraxin PTX3 in resistance against pulmonary infection with Klebsiella pneumoniae in transgenic mice

The long pentraxin PTX3 is expressed during acute inflammation and appears to control nitric oxide (NO) and tumor necrosis factor (TNF)-alpha production. In the present study, the physiological function of PTX3 was investigated in a model of pulmonary infection caused by the Gram-negative bacterium Klebsiella pneumoniae. Transgenic mice expressing multiple copies of PTX3 under the control of its own promoter were used to assess lethality rates, bacterial counts and inflammatory indices following pulmonary infection of mice. Expression of PTX3 is enhanced during pulmonary infection in wild-type mice. In transgenic mice given a high inoculum, overt PTX3 expression was associated with faster lethality. Faster lethality correlated with enhanced nitrate in plasma, an inability of neutrophils to migrate to lung tissue and greater dissemination of bacteria to blood at 20h after infection. In contrast, transgenic PTX3 expression conferred protection to mice given lower pulmonary inocula. In the latter experiments, there was enhanced TNF-alpha production, greater neutrophil influx and phagocytosis of bacteria by migrated neutrophils. By controlling the production of TNF-alpha and NO, and depending on the intensity of the inflammatory response induced by a given inoculum, the expression of PTX3 may favor or disfavor the influx of neutrophils and the ability of the murine host to deal with pulmonary infection with K. pneumoniae. These experiments highlight the delicate balance that exists among the various mediators that control the inflammatory response and suggest that PTX3 is an essential part of the ability of a host to deal with bacterial infection.     

Interleukin-12p40 modulates human metapneumovirus-induced pulmonary disease in an acute mouse model of infection

The mechanisms that regulate the host immune response induced by human metapneumovirus (hMPV), a newly-recognized member of the Paramyxoviridae family, are largely unknown. Cytokines play an important role in modulating inflammatory responses during viral infections. IL-12p40, a known important mediator in limiting lung inflammation, is induced by hMPV and its production is sustained after the resolution phase of infection suggesting that this cytokine plays a role in the immune response against hMPV. In this work, we demonstrated that in mice deficient in IL-12p40, hMPV infection induced an exacerbated pulmonary inflammatory response and mucus production, altered cytokine response, and decreased lung function. However, hMPV infection in these mice does not have an effect on viral replication. These results identify an important regulatory role of IL-12p40 in hMPV infection.     

Eosinophils mediate SIgA production triggered by TLR2 and TLR4 to control Ascaris suum infection in mice

Human ascariasis is the most prevalent but neglected tropical disease in the world, affecting approximately 450 million people. The initial phase of Ascaris infection is marked by larval migration from the host’s organs, causing mechanical injuries followed by an intense local inflammatory response, which is characterized mainly by neutrophil and eosinophil infiltration, especially in the lungs. During the pulmonary phase, the lesions induced by larval migration and excessive immune responses contribute to tissue remodeling marked by fibrosis and lung dysfunction. In this study, we investigated the relationship between SIgA levels and eosinophils. We found that TLR2 and TLR4 signaling induces eosinophils and promotes SIgA production during Ascaris suum infection. Therefore, control of parasite burden during the pulmonary phase of ascariasis involves eosinophil influx and subsequent promotion of SIgA levels. In addition, we also demonstrate that eosinophils also participate in the process of tissue remodeling after lung injury caused by larval migration, contributing to pulmonary fibrosis and dysfunction in re-infected mice. In conclusion, we postulate that eosinophils play a central role in mediating host innate and humoral immune responses by controlling parasite burden, tissue inflammation, and remodeling during Ascaris suum infection. Furthermore, we suggest that the use of probiotics can induce eosinophilia and SIgA production and contribute to controlling parasite burden and morbidity of helminthic diseases with pulmonary cycles.     

Interleukin 10 modulation of neutrophil subsets infiltrating lungs during Streptococcus pneumoniae infection

Interleukin-10 production and lung neutrophil infiltration are two essential components of the balanced immune response to pneumonia caused by Streptococcus pneumoniae. Here we describe the existence of two neutrophil subsets in lungs during experimental S. pneumoniae infection in mice, which have different size, granularity and expression of activation markers. During infection, both neutrophils subsets were increased in the lungs of IL-10 producing mice, however this increment was significantly higher in the absence of this cytokine. These results suggest that IL-10 is a key cytokine that regulates lung inflammation during bacterial infection caused by specific neutrophil subsets infiltrating the lungs.     

A dual role for TGF-beta1 in the control and persistence of fungal pneumonia

TGF-beta1 (TGF) has been implicated in the pathogenesis of several chronic infections and is thought to promote microbial persistence by interfering with macrophage function. In rats with experimental pulmonary cryptococcosis, increased lung levels of TGF were present at 12 mo of infection. Within the lung, expression of TGF localized to epithelioid cells and foamy macrophages in areas of inflammation. Increased TGF expression was also observed in the lungs of experimentally infected mice and a patient with pulmonary cryptococcosis. TGF reduced Ab and serum-mediated phagocytosis of Cryptococcus neoformans by rat alveolar macrophages (AM) and peripheral blood monocytes, and this was associated with decreased chemokine production and oxidative burst. Interestingly, TGF-treated rat AM limited both intracellular and extracellular growth of C. neoformans. Control of C. neoformans growth by TGF-treated rat AM was due to increased secretion of lysozyme, a protein with potent antifungal activity. The effects of TGF on the course of infection were dependent on the timing of TGF administration relative to the time of infection. TGF treatment of chronically infected rats resulted in reduced lung fungal burden, while treatment early in the course of infection resulted in increased fungal burden. In summary, our studies suggest a dual role for TGF in persistent fungal pneumonia whereby it contributes to the local control of infection by enhancing macrophage antifungal efficacy through increased lysozyme secretion, while limiting inflammation by inhibiting macrophage/monocyte phagocytosis and reducing associated chemokine production and oxidative burst.     

Depletion of alveolar macrophages exerts protective effects in pulmonary tuberculosis in mice

Mycobacterium tuberculosis bacilli are intracellular organisms that reside in phagosomes of alveolar macrophages (AMs). To determine the in vivo role of AM depletion in host defense against M. tuberculosis infection, mice with pulmonary tuberculosis induced by intranasal administration of virulent M. tuberculosis were treated intranasally with either liposome-encapsulated dichloromethylene diphosphonate (AM(-) mice), liposomes, or saline (AM(+) mice). AM(-) mice were completely protected against lethality, which was associated with a reduced outgrowth of mycobacteria in lungs and liver, and a polarized production of type 1 cytokines in lung tissue, and by splenocytes stimulated ex vivo. AM(-) mice displayed deficient granuloma formation, but were more capable of attraction and activation of T cells into the lung and had increased numbers of pulmonary polymorphonuclear cells. These data demonstrate that depletion of AMs is protective during pulmonary tuberculosis.     

Local CD11c+ MHC class II- precursors generate lung dendritic cells during respiratory viral infection, but are depleted in the process

Increases in numbers of lung dendritic cells (DC) observed during respiratory viral infections are assumed to be due to recruitment from bone marrow precursors. No local production has been demonstrated. In this study, we isolated defined populations of murine lung cells based on CD11c and MHC class II (MHC II) expression. After culture for 12 days with GM-CSF, we analyzed cell numbers, DC surface markers, and Ag-presenting capacity. Only CD11c+ MHC II- cells from naive mice proliferated, yielding myeloid DC, which induced Ag-specific proliferation of naive T cells. After respiratory syncytial virus (RSV) infection, numbers of pulmonary CD11c+ MHC II- precursor cells were significantly reduced and DC could not be generated. Moreover, RSV infection prevented subsequent in vivo expansion of pulmonary DC in response to influenza infection or LPS treatment. These results provide direct evidence of local generation of fully functional myeloid DC in the lung from CD11c+ MHC II(-) precursor cells that are depleted by RSV infection, leading to an inability to expand lung DC numbers in response to subsequent viral infection or exposure to bacterial products. This depletion of local DC precursors in respiratory viral infections may be important in explaining complex interactions between multiple and intercurrent pulmonary infections.     

IL-12-independent Th1-type immune responses to respiratory viral infection: requirement of IL-18 for IFN-gamma release in the lung but not for the differentiation of viral-reactive Th1-type lymphocytes

We demonstrated that IL-12 was induced during primary or secondary pulmonary adenoviral infection in wild-type (wt) mice. However, cellular responses were not compromised in the lungs of IL-12-/- mice. The level of IFN-gamma in the lung was similar in wt and IL-12-/- mice during pulmonary viral infection. Upon Ag stimulation in vitro, lymphocytes from draining lymph nodes or spleen of infected IL-12-/- mice released large amounts of IFN-gamma, but not IL-4, which were comparable to those released by wt lymphocytes. Furthermore, a predominantly IgG2a response to adenoviral infection was unimpaired in IL-12-/- mice. These significant anti-adenoviral Th1-type responses in IL-12-/- mice led to an efficient clearance of virus-infected cells in the lung. Whether IL-18 was involved in IL-12-independent anti-adenoviral immune responses was investigated. Abrogation of endogenous IL-18 by an Ab resulted in diminished IFN-gamma release and lymphocytic infiltrate in the lung during adenoviral infection. Nevertheless, the development of lymphocytes of the Th1 phenotype was unimpaired in the absence of both IL-12 and IL-18. In contrast to their intact ability to mount Th1-type responses to viral infection, IL-12-/- mice suffered impaired Th1-type immune responses to pulmonary mycobacterial infection. Our findings suggest that IL-12, although induced, is not required for Th1-type responses to respiratory viral infection, in contrast to mycobacterial infection. IL-18 is required for the optimal release of IFN-gamma in the lung during viral infection, but is not required for the generation of virus-reactive Th1-type lymphocytes. Th1 differentiation during respiratory adenoviral infection may involve molecules different from IL-12 or IL-18.     

Inhibition of pulmonary antibacterial defense by interferon-gamma during recovery from influenza infection

Secondary bacterial infection often occurs after pulmonary virus infection and is a common cause of severe disease in humans, yet the mechanisms responsible for this viral-bacterial synergy in the lung are only poorly understood. We now report that pulmonary interferon-gamma (IFN-gamma) produced during T cell responses to influenza infection in mice inhibits initial bacterial clearance from the lung by alveolar macrophages. This suppression of phagocytosis correlates with lung IFN-gamma abundance, but not viral burden, and leads to enhanced susceptibility to secondary pneumococcal infection, which can be prevented by IFN-gamma neutralization after influenza infection. Direct inoculation of IFN-gamma can mimic influenza infection and downregulate the expression of the class A scavenger receptor MARCO on alveolar macrophages. Thus, IFN-gamma, although probably facilitating induction of specific anti-influenza adaptive immunity, suppresses innate protection against extracellular bacterial pathogens in the lung.     

Kinase suppressor of Ras-1 protects against pulmonary Pseudomonas aeruginosa infections

Pseudomonas aeruginosa is a Gram-negative pathogen that causes severe infections in immunocompromised individuals and individuals with cystic fibrosis or chronic obstructive pulmonary disease (COPD). Here we show that kinase suppressor of Ras-1 (Ksr1)-deficient mice are highly susceptible to pulmonary P. aeruginosa infection accompanied by uncontrolled pulmonary cytokine release, sepsis and death, whereas wild-type mice clear the infection. Ksr1 recruits and assembles inducible nitric oxide (NO) synthase (iNOS) and heat shock protein-90 (Hsp90) to enhance iNOS activity and to release NO upon infection. Ksr1 deficiency prevents lung alveolar macrophages and neutrophils from activating iNOS, producing NO and killing bacteria. Restoring NO production restores the bactericidal capability of Ksr1-deficient lung alveolar macrophages and neutrophils and rescues Ksr1-deficient mice from P. aeruginosa infection. Our findings suggest that Ksr1 functions as a previously unknown scaffold that enhances iNOS activity and is therefore crucial for the pulmonary response to P. aeruginosa infections.     

Pulmonary immune cells and inflammatory cytokine dysregulation are associated with mortality of IL-1R1-/-mice infected with influenza virus (H1 N1)

Respirovirus infection can cause viral pneumonia and acute lung injury (ALI).The interleukin-1 (IL-1) family consists of proinflammatory cytokines that play essential roles in regulating immune and inflammatory responses in vivo.IL-1 signaling is associated with protection against respiratory influenza virus infection by mediation of the pulmonary anti-viral immune response and inflammation.We analyzed the infiltration lung immune leukocytes and cytokines that contribute to inflammatory lung pathology and mortality of fatal H1N1 virus-infected IL-1 receptor 1 (IL-1R1) deficient mice.Results showed that early innate immune cells and cytokine/chemokine dysregulation were observed with significantly decreased neutrophil infiltration and IL-6,TNF-α,G-CSF,KC,and MIP-2 cytokine levels in the bronchoalveolar lavage fluid of infected IL-1R1-/-mice in comparison with that of wild type infected mice.The adaptive immune response against the H1N1 virus in IL-1R1-/-mice was impaired with downregulated anti-viral Th1 cell,CD8+ cell,and antibody functions,which contributes to attenuated viral clearance.Histological analysis revealed reduced lung inflammation during early infection but severe lung pathology in late infection in IL-1R1-/-mice compared with that in WT infected mice.Moreover,the infected IL-1R1-/-mice showed markedly reduced neutrophil generation in bone marrow and neutrophil recruitment to the inflamed lung.Together,these results suggest that IL-1 signaling is associated with pulmonary anti-influenza immune response and inflammatory lung injury,particularly via the influence on neutrophil mobilization and inflammatory cytokine/chemokine production.     

Respiratory syncytial virus infection suppresses lung CD8+ T-cell effector activity and peripheral CD8+ T-cell memory in the respiratory tract.

Respiratory syncytial virus (RSV) is a major cause of morbidity from respiratory infection in infants, young children and the elderly. No effective vaccine against RSV is currently available and studies of the natural history of RSV infection suggest repeated infections with antigenically related virus strains are common throughout an individual's lifetime. We have studied the CD8+ T-cell response during experimental murine RSV infection and found that RSV inhibits the expression of effector activity by activated RSV-specific CD8+ T cells infiltrating the lung parenchyma and the development of pulmonary CD8+ T-cell memory by interfering with TCR-mediated signaling. These data suggest a possible mechanism to explain the limited duration of protective immunity in RSV infection.