Pulmonary Host Defense Research Articles

MerTK negatively regulates Staphylococcus aureus induced inflammatory response via SOCS1/SOCS3 and Mal.

Staphylococcus aureus (S. aureus), one of Gram-positive pathogen, is frequently associated with acute lung inflammation. The central feature of S. aureus acute lung inflammation are pulmonary dysfunctioning and impeded host defence response, which cause failure in inflammatory cytokines homeostasis and leads to serious tissue damage. However, the role of the Mer receptor tyrosine kinase (MerTK) in the lung following S. aureus infection remains elusive. Here, we investigate whether MerTK alleviates S. aureus induced uncontrolled inflammation through negatively regulating toll-like receptor 2 and 6 (TLR2/ TLR6) via suppressor of cytokine signalling 1, 3 (SOCS1/SOCS3). We found in mice lung tissues and RAW 264.7 macrophages upon S. aureus infection activates TLR2 and TLR6 driven mitogen-activated protein kinases (MAPKs) and nuclear factor kappa B (NF-κB) signalling pathways, resulting in production of inflammatory cytokines including tumour necrosis factor-α (TNF-α), interleukin 1β (IL-1β), interleukin 6 (IL-6). Furthermore, S. aureus-infection groups showed a significant up-regulation of MerTK which serves as mediator of SOCS1 and SOCS3. Subsequently, through feedback mechanism SOCS1/3 degrade Mal, resulting in inhibition of downstream TLR mediated inflammatory pathways. Moreover, MerTK-/- mice lung tissues and silencing MerTK in RAW 264.7 inhibited the S. aureus-induced activation of MerTK, which significantly upregulated the phosphorylation of crucial protein in MAPKs (ERK, JNK, p38) and NF-κB (IĸBα, p65) signalling pathways, as well as the production of pro-inflammatory cytokines. Collectively, these findings indicate the important role of MerTK in self-regulatory resolution of S. aureus-induced inflammatory pathways and cytokines through intrinsic SOCS1 and SOCS3 repressed feedback on TLR2, TLR6 both in vivo and in vitro.

Caspase-11 contributes to pulmonary host defense against Klebsiella pneumoniae and local activation of coagulation.

Klebsiella (K.) pneumoniae is a common cause of gram-negative pneumonia and sepsis. Caspase-11 is an intracellular receptor for lipopolysaccharide and regulates pyroptosis, a specific form of inflammatory cell death, which aids in host defense against intracellular gram-negative bacteria. Recently, caspase-11 has also been implicated in blood coagulation. Previously, we found that local fibrin formation contributes to protective immunity against Klebsiella infection of the lung. The aim of the present study was to determine the role of caspase-11 in host defense during K. pneumoniae-evoked pneumonia and sepsis. Therefore, we infected wild-type and caspase-11-deficient (Casp11-/-) mice with a low-dose K. pneumoniae via the airways to induce a gradually evolving pneumosepsis. Casp11-/- mice displayed increased bacterial numbers in the lung 12 h and 48 h after inoculation. Analysis of pulmonary IL-1α, IL-1β, and TNF levels showed reduced IL-1α levels in bronchoalveolar lavage fluid and increased TNF levels in the lung of Casp11-/- mice at 48 h after inoculation. Lung γH2AX staining (marker for cell death), lung pathology and neutrophil influx in the lung, as well as bacterial dissemination and organ damage, however, were not altered in Casp11-/- mice after Klebsiella infection. Strikingly, analysis of cross-linked fibrin and D-dimer (markers for coagulation) revealed significantly less fibrin formation in the lungs of Casp11-/- mice at either time point after Klebsiella infection. These data reveal that caspase-11 contributes to protective immunity against K. pneumoniae possibly by activation of blood coagulation in the lung.

Co-expression of surfactant protein A and chicken lung lectin in chicken respiratory system

Chicken surfactant protein A (cSP-A) and chicken lung lectin (cLL) are C-type lectins that play important roles in pulmonary host defense responses. Herein, we explored the localization of cSP-A and cLL in the chicken respiratory system. Six tissues from 30-days-old SPF chickens were used to quantify the expression of cSP-A and cLL using the quantitative real-time reverse transcriptional polymerase chain reaction (qRT-PCR) and fluorescence multiplex immunohistochemistry staining (fluorescence mIHC staining). Results showed that cSP-A and cLL mRNA were highly expressed in lungs compared to other tissues. cSP-A mRNA expression levels in all tissues were higher compared with cLL expression levels as analyzed using qRT-PCR. Fluorescence mIHC co-expression of cSP-A and cLL were mainly detected in lung parabronchial epithelia, and mucosal epithelia of larynx, trachea, syrinx, bronchus and air sac, with cSP-A showing a stronger positive staining compared with cLL. cLL is expressed on both mucosal surfaces, some individual lung epithelial cells and cartilage cells, while cSP-A is mainly restricted to mucosal surfaces of the respiratory tract. These histological findings may be useful for understanding the biological significance of this pulmonary lectins in future studies.

Deletion of MCP-1 enhances susceptibility to carbapenem-resistant Klebsiella pneumoniae via modulating neutrophil and macrophage function

Abstract Monocyte chemoattractant protein-1 (MCP-1) is important for monocyte recruitment to the lungs in response to bacterial infection. MCP-1 is also essential for neutrophil-dependent host defense during Escherichia coli infection. Carbapenem-Resistant Klebsiella pneumoniae (CRKP) infection has been rapidly growing as a life-threatening nosocomial disease in the U.S. and world. However, the role of chemokines and cellular responses in protective immunity to pulmonary infection with CRKP is largely unknown. Here, we investigated the role of MCP-1 in pulmonary host defense against CRKP infection using MCP-1 knockout (KO) mice. We determined survival, bacterial burden, and neutrophil influx, cytokine/chemokine expression as well as cellular functions, such as formation of neutrophil extracellular traps (NETs) and macrophage extracellular traps (METs). MCP-1 is essential for survival and restricting bacterial outgrowth organs. Compared to the C57Bl/6 (control) mice, KO mice showed impaired influx of neutrophils in the airways as assessed by nuclear cell morphology in BALF and by myeloperoxidase activity (MPO) in lung parenchyma. Moreover, the NETs and METs induced by CRKP were reduced in KO mice in both in vitro or in vivo. Intriguingly, subsequent to CRKP infection, KO mice demonstrated increased pyroptosis through enhanced activation of caspase-1, higher expression of IL-1β and cleaved gasdermin-D in the lungs. These findings enhance our understanding of the critical role of MCP-1 in modulating neutrophil function, such as NETosis, METosis and pyroptosis and suggest that therapies targeting modulation of NETs, METs and pyroptosis during CRKP infection can be explored through clinical trials.

Epithelial membrane protein 2 regulates transepithelial migration of neutrophils into the inflamed airspace

Abstract Neutrophil (PMN) migration from the circulation to the airway lumen is critical for pulmonary host defense but also causes bystander tissue injury that underlies a wide array of acute and chronic lung diseases. Whereas good evidence supports a role for intestinal epithelial cells in control of transmigration of extravasated PMNs into the intestinal lumen, little is known about whether and how respiratory epithelial cells regulate PMN influx into the airspace. Here, we identify epithelial membrane protein 2 (EMP2), a tetraspan membrane protein highly expressed in alveolar epithelial type 1 (AT1) cells but of unknown function in lung biology, as a master regulator of the terminal entry of PMNs into the airspace. EMP2−/− mice had reduced PMN influx into the airspace after multiple inhaled exposures and as a result exhibited attenuated lung injury and improved survival during bacterial pneumonia. Bone marrow chimeras, intravital PMN labelling, and in vitro assays suggest epithelial dysfunction in EMP2−/− lungs was responsible for deficient transepithelial PMN transit into airspace. We provide evidence that EMP2 supports epithelial surface display of multiple adhesion molecules that are required for cognate interactions with migrating PMNs and that it does so through suppression of epithelial caveolins. Moreover, EMP2 supports epithelial lipid raft mass in a manner dependent on putative cholesterol-binding motifs in its sequence. Taken together, we propose that EMP2-dependent membrane organization ensures proper display on AT1 cells of a suite of proteins required to instruct paracellular PMN traffic into the alveolus. Our findings identify EMP2 as a potential target for treatment of neutrophilic lung diseases.

Macrophage FABP4 is required for neutrophil recruitment and bacterial clearance in Pseudomonas aeruginosa pneumonia.

Fatty acid binding protein 4 (FABP4), an intracellular lipid chaperone and adipokine, is expressed by lung macrophages, but the function of macrophage-FABP4 remains elusive. We investigated the role of FABP4 in host defense in a murine model of Pseudomonas aeruginosa pneumonia. Compared with wild-type (WT) mice, FABP4-deficient (FABP4-/-) mice exhibited decreased bacterial clearance and increased mortality when challenged intranasally with P. aeruginosa. These findings in FABP4-/- mice were associated with a delayed neutrophil recruitment into the lungs and were followed by greater acute lung injury and inflammation. Among leukocytes, only macrophages expressed FABP4 in WT mice with P. aeruginosa pneumonia. Chimeric FABP4-/- mice with WT bone marrow were protected from increased mortality seen in chimeric WT mice with FABP4-/- bone marrow during P. aeruginosa pneumonia, thus confirming the role of macrophages as the main source of protective FABP4 against that infection. There was less production of C-X-C motif chemokine ligand 1 (CXCL1) in FABP4-/- alveolar macrophages and lower airway CXCL1 levels in FABP4-/- mice. Delivering recombinant CXCL1 to the airways protected FABP4-/- mice from increased susceptibility to P. aeruginosa pneumonia. Thus, macrophage-FABP4 has a novel role in pulmonary host defense against P. aeruginosa infection by facilitating crosstalk between macrophages and neutrophils via regulation of macrophage CXCL1 production.-Liang, X., Gupta, K., Rojas Quintero, J., Cernadas, M., Kobzik, L., Christou, H., Pier, G. B., Owen, C. A., Çataltepe, S. Macrophage FABP4 is required for neutrophil recruitment and bacterial clearance in Pseudomonas aeruginosa pneumonia.

STAT2 Signaling Regulates Macrophage Phenotype During Influenza and Bacterial Super-Infection.

Influenza is a common respiratory virus that infects between 5 and 20% of the US population and results in 30,000 deaths annually. A primary cause of influenza-associated death is secondary bacterial pneumonia. We have previously shown that influenza induces type I interferon (IFN)-mediated inhibition of Type 17 immune responses, resulting in exacerbation of bacterial burden during influenza and Staphylococcus aureus super-infection. In this study, we investigated the role of STAT2 signaling during influenza and influenza-bacterial super-infection in mice. Influenza-infected STAT2−/− mice had increased morbidity, viral burden, and inflammation when compared to wild-type mice. Despite an exaggerated inflammatory response to influenza infection, we found increased bacterial control and survival in STAT2 deficient mice during influenza-MRSA super-infection compared to controls. Further, we found that increased bacterial clearance during influenza-MRSA super-infection is not due to rescue of Type 17 immunity. Absence of STAT2 was associated with increased accumulation of M1, M2 and M1/M2 co-expressing macrophages during influenza-bacterial super-infection. Neutralization of IFNγ (M1) and/or Arginase 1 (M2) impaired bacterial clearance in Stat2−/− mice during super-infection, demonstrating that pulmonary macrophages expressing a mixed M1/M2 phenotype promote bacterial control during influenza-bacterial super-infection. Together, these results suggest that the STAT2 signaling is involved in suppressing macrophage activation and bacterial control during influenza-bacterial super-infection. Further, these studies reveal novel mechanistic insight into the roles of macrophage subpopulations in pulmonary host defense.

NLRP6 negatively regulates pulmonary host defense in Gram-positive bacterial infection through modulating neutrophil recruitment and function.

Gram-positive bacteria, including Staphylococcus aureus are endemic in the U.S., which cause life-threatening necrotizing pneumonia. Neutrophils are known to be critical for clearance of S. aureus infection from the lungs and extrapulmonary organs. Therefore, we investigated whether the NLRP6 inflammasome regulates neutrophil-dependent host immunity during pulmonary S. aureus infection. Unlike their wild-type (WT) counterparts, NLRP6 knockout (KO) mice were protected against pulmonary S. aureus infection as evidenced by their higher survival rate and lower bacterial burden in the lungs and extrapulmonary organs. In addition, NLRP6 KO mice displayed increased neutrophil recruitment following infection, and when neutrophils were depleted the protective effect was lost. Furthermore, neutrophils from the KO mice demonstrated enhanced intracellular bacterial killing and increased NADPH oxidase-dependent ROS production. Intriguingly, we found higher NK cell-mediated IFN-γ production in KO mouse lungs, and treatment with IFN-γ was found to enhance the bactericidal ability of WT and KO neutrophils. The NLRP6 KO mice also displayed decreased pyroptosis and necroptosis in the lungs following infection. Blocking of pyroptosis and necroptosis in WT mice resulted in increased survival, reduced bacterial burden in the lungs, and attenuated cytokine production. Taken together, these novel findings show that NLRP6 serves as a negative regulator of neutrophil-mediated host defense during Gram-positive bacterial infection in the lungs through regulating both neutrophil influx and function. These results also suggest that blocking NLRP6 to augment neutrophil-associated bacterial clearance should be considered as a potential therapeutic intervention strategy for treatment of S. aureus pneumonia.

Iron restriction inside macrophages regulates pulmonary host defense against Rhizopus species

Mucormycosis is a life-threatening respiratory fungal infection predominantly caused by Rhizopus species. Mucormycosis has incompletely understood pathogenesis, particularly how abnormalities in iron metabolism compromise immune responses. Here we show how, as opposed to other filamentous fungi, Rhizopus spp. establish intracellular persistence inside alveolar macrophages (AMs). Mechanistically, lack of intracellular swelling of Rhizopus conidia results in surface retention of melanin, which induces phagosome maturation arrest through inhibition of LC3-associated phagocytosis. Intracellular inhibition of Rhizopus is an important effector mechanism, as infection of immunocompetent mice with swollen conidia, which evade phagocytosis, results in acute lethality. Concordantly, AM depletion markedly increases susceptibility to mucormycosis. Host and pathogen transcriptomics, iron supplementation studies, and genetic manipulation of iron assimilation of fungal pathways demonstrate that iron restriction inside macrophages regulates immunity against Rhizopus. Our findings shed light on the pathogenetic mechanisms of mucormycosis and reveal the role of macrophage-mediated nutritional immunity against filamentous fungi.

Antibiotic therapy-induced collateral damage: IgA takes center stage in pulmonary host defense.

The use of broad-spectrum antibiotics in empirical antimicrobial therapy is a lifesaving strategy for patients in intensive care. At the same time, antibiotics dramatically increase the risk for nosocomial infections, such as hospital‑acquired pneumonia caused by Pseudomonas aeruginosa, and other antibiotic-resistant bacteria. In this issue of the JCI, Robak and colleagues identified a mechanism by which depletion of resident gut and lung microbiota by antibiotic treatment results in secondary IgA deficiency and impaired anti-P. aeruginosa host defense. Impaired defenses could be improved by substitution of polyclonal IgA via the intranasal route in a mouse model of pneumonia. Importantly, antibiotic treatment caused lung IgA deficiency that involved reduced TLR-dependent production of a proliferation-inducing ligand (APRIL) and B cell-activating factor (BAFF) in intensive care unit patients. These patients might therefore benefit from future strategies to increase pulmonary IgA levels.

NKG2D Regulation of Lung Pathology and Dendritic Cell Function Following Respiratory Syncytial Virus Infection.

Respiratory syncytial virus (RSV) is a common cause of respiratory tract infection in vulnerable populations. Natural killer (NK) cells and dendritic cells (DC) are important for the effector functions of both cell types following infection. Wild-type and NKG2D-deficient mice were infected with RSV. Lung pathology was assessed by histology. Dendritic cell function and phenotype were evaluated by enzyme-linked immunosorbent assay and flow cytometry. The expression of NKG2D ligands on lung and lymph node DCs was measured by immunostaining and flow cytometry. Adoptive transfer experiments were performed to assess the importance of NKG2D-dependent DC function in RSV infection. NKG2D-deficient mice exhibited greater lung pathology, marked by the accumulation of DCs following RSV infection. Dendritic cells isolated from NKG2D-deficient mice had impaired responses toward Toll-like receptor ligands. Dendritic cells expressed NKG2D ligands on their surface, which was further increased in NKG2D-deficient mice and during RSV infection. Adoptive transfer of DCs isolated from wild-type mice into the airways of NKG2D-deficient mice ameliorated the enhanced inflammation in NKG2D-deficient mice after RSV infection. NKG2D-dependent interactions with DCs control the phenotype and function of DCs and play a critical role in pulmonary host defenses against RSV infection.

Nociceptor sensory neurons suppress neutrophil and γδ T cell responses in bacterial lung infections and lethal pneumonia.

Lung-innervating nociceptor sensory neurons detect noxious or harmful stimuli and consequently protect organisms by mediating coughing, pain, and bronchoconstriction. However, the role of sensory neurons in pulmonary host defense is unclear. Here, we found that TRPV1+ nociceptors suppressed protective immunity against lethal Staphylococcus aureus pneumonia. Targeted TRPV1+-neuron ablation increased survival, cytokine induction, and lung bacterial clearance. Nociceptors suppressed the recruitment and surveillance of neutrophils, and altered lung γδ T cell numbers, which are necessary for immunity. Vagal ganglia TRPV1+ afferents mediated immunosuppression through release of the neuropeptide calcitonin gene-related peptide (CGRP). Targeting neuroimmunological signaling may be an effective approach to treat lung infections and bacterial pneumonia.

Toll-like receptor 2 has a prominent but nonessential role in innate immunity to Staphylococcus aureus pneumonia.

Staphylococcus aureus is an important cause of acute bacterial pneumonia. Toll‐like receptor 2 (TLR2) recognizes multiple components of the bacterial cell wall and activates innate immune responses to gram‐positive bacteria. We hypothesized that TLR2 would have an important role in pulmonary host defense against S. aureus. TLR null (TLR2−/−) mice and wild type (WT) C57BL/6 controls were challenged with aerosolized S. aureus at a range of inocula for kinetic studies of cytokine and antimicrobial peptide expression, lung inflammation, bacterial killing by alveolar macrophages, and bacterial clearance. Survival was measured after intranasal infection. Pulmonary induction of most pro‐inflammatory cytokines was significantly blunted in TLR2−/− mice 4 and 24 h after infection in comparison with WT controls. Bronchoalveolar concentrations of cathelicidin‐related antimicrobial peptide also were reduced in TLR2−/− mice. Lung inflammation, measured by enumeration of bronchoalveolar neutrophils and scoring of histological sections, was significantly blunted in TLR2−/− mice. Phagocytosis of S. aureus by alveolar macrophages in vivo after low‐dose infection was unimpaired, but viability of ingested bacteria was significantly greater in TLR2−/− mice. Bacterial clearance from the lungs was slightly impaired in TLR2−/− mice after low‐dose infection only; bacterial elimination from the lungs was slightly accelerated in the TLR2−/− mice after high‐dose infection. Survival after high‐dose intranasal challenge was 50–60% in both groups. TLR2 has a significant role in early innate immune responses to S. aureus in the lungs but is not required for bacterial clearance and survival from S. aureus pneumonia.

Serum surfactant protein D as a marker for bronchopulmonary dysplasia

Background: Lung epithelial cells express surfactant protein D (SP-D), a calcium-dependent lectin that plays an important role in antibody-independent pulmonary host defense. Previous studies have shown that it is found in the peripheral circulation in patients with pulmonary disease, likely because of translocation into the blood when lung epithelial barriers are disrupted by inflammation or acute injury. In adults, serum SP-D levels are biomarkers for the progression and severity of chronic lung disease. In neonates, elevated SP-D levels in cord blood and on day 1 have been associated with prenatal risk factors and with an increased risk of respiratory distress syndrome and infections. It is not known whether serum SP-D during the first week of life is a marker for bronchopulmonary dysplasia (BPD), a form of chronic lung disease of prematurity that is associated with lung parenchymal maldevelopment and injury.Objective: The goal of this study is to determine whether serum SP-D on days 3 and 7 of life are associated with the development of BPD in preterm infants.Design/methods: Serum samples were obtained on postnatal days 3 and 7 from 106 preterm infants (500–2000 g birth weight, 23–32-week gestation). SP-D was quantified by Western blot. BPD was determined at 36 weeks PMA using NICHD criteria.Results: The mean birth weight was 1145 ± 347 g and gestational age 29.2 ± 7.4 weeks. BPD was diagnosed in 7 and “BPD or death” in 16 infants. Days 3 and 7 values tracked significantly (r = 0.648), and did not correlate with birth weight or gestational age. Contrary to expectations, serum SP-D was not associated with BPD. Significant gender differences were noted, with SP-D dropping from day 3 to day 7 in males, while increasing in females (p < .05).Conclusion: Elevated serum SP-D does not appear to be a useful marker for BPD. Decreasing serum SP-D levels in males, as compared to females, during the first week of life are likely related to gender differences in lung maturation, consistent with the higher incidence of BPD in males.

Endurance exercise training decreased serum levels of surfactant protein D and improved aerobic fitness of obese women with type-2 diabetes

BackgroundSurfactant protein D (SP-D) is a member of the collectin family and is an important component of the pulmonary innate host defense. To find the relationship between exercise training and SP-D in diabetes, we examined the possible effects of a 10-week endurance exercise-training program on serum levels of SP-D, leptin, lipid profile and insulin resistance in obese women with type-2 diabetes mellitus (T2DM).MethodsTwenty-two obese women with T2DM were randomly assigned to either exercise training (ET) or control (C) group. A subject dropped from ET group due to personal reasons and 1 subject dropped from C group due to commitment to experiments conditions. A total of twenty obese women with T2DM were randomly assigned into endurance exercise training (ET = 10) and control (C = 10) groups. The training group underwent a progressive endurance-training program for 10 weeks (running on a treadmill for 30–55 min/day at 50–75% heart rate reserve) and the control group did not participate in any exercise program. Venous blood samples were collected from both groups before and 72 h after the last session of exercise training for analysis of serum SP-D, leptin, lipid profile, glucose and insulin. Data were analyzed using 2 (group: control, endurance training) × 2 (time: pre, post) ANOVA with repeated measures on the second factor. Absolute changes from rest (∆ baseline) values were calculated according to the following formula: ((measure-baseline)·baseline-1)·100. Percent change between groups was analyzed using independent t-tests (p < 0.05). All analyses were completed using SPSS 19.ResultsThe serum SP-D levels were decreased after exercise training in ET (∆ = − 78.78 ± 17.14%, p = 0.001) when compare to C (∆ = 9.41 ± 4.75%). Obese diabetic women in the ET group showed significantly lower serum leptin levels (8053.27 ± 878.7 pg/ml, ∆ = − 26.97 ± 16.41%) when compared with women in the control group (9885.5 ± 696 pg/ml, ∆ = 7.02 ± 3.46%, p = 0.003). Fasting glucose was favorably and significantly affected by the intervention (ET ∆ = − 17.01 ± 12.74%, Control ∆ = 15.47 ± 7.32%, p = 0.011). VO2max as an index of aerobic fitness was significantly increased after 10-weeks of endurance exercise training (ET ∆ = 19.29 ± 6.18%).ConclusionsEndurance exercise training with improvement in aerobic fitness induced a significant reduction of serum SP-D levels in obese women with T2DM.

Alcohol-associated intestinal dysbiosis impairs pulmonary host defense against Klebsiella pneumoniae.

Chronic alcohol consumption perturbs the normal intestinal microbial communities (dysbiosis). To investigate the relationship between alcohol-mediated dysbiosis and pulmonary host defense we developed a fecal adoptive transfer model, which allows us to investigate the impact of alcohol-induced gut dysbiosis on host immune response to an infectious challenge at a distal organ, independent of prevailing alcohol use. Male C57BL/6 mice were treated with a cocktail of antibiotics (ampicillin, gentamicin, neomycin, vancomycin, and metronidazole) via daily gavage for two weeks. A separate group of animals was fed a chronic alcohol (or isocaloric dextrose pair-fed controls) liquid diet for 10 days. Microbiota-depleted mice were recolonized with intestinal microbiota from alcohol-fed or pair-fed (control) animals. Following recolonization groups of mice were sacrificed prior to and 48 hrs. post respiratory infection with Klebsiella pneumoniae. Klebsiella lung burden, lung immunology and inflammation, as well as intestinal immunology, inflammation, and barrier damage were examined. Results showed that alcohol-associated susceptibility to K. pneumoniae is, in part, mediated by gut dysbiosis, as alcohol-naïve animals recolonized with a microbiota isolated from alcohol-fed mice had an increased respiratory burden of K. pneumoniae compared to mice recolonized with a control microbiota. The increased susceptibility in alcohol-dysbiosis recolonized animals was associated with an increase in pulmonary inflammatory cytokines, and a decrease in the number of CD4+ and CD8+ T-cells in the lung following Klebsiella infection but an increase in T-cell counts in the intestinal tract following Klebsiella infection, suggesting intestinal T-cell sequestration as a factor in impaired lung host defense. Mice recolonized with an alcohol-dysbiotic microbiota also had increased intestinal damage as measured by increased levels of serum intestinal fatty acid binding protein. Collectively, these results suggest that alterations in the intestinal immune response as a consequence of alcohol-induced dysbiosis contribute to increased host susceptibility to Klebsiella pneumonia.

STAT2-mediated regulation of Macrophage Phenotype during Influenza and Bacterial Super-infection

Abstract Influenza is a common contagious respiratory illness that affects people worldwide. Secondary bacterial infection is a major complication during influenza infection that causes significant morbidity and mortality. We have previously shown that influenza induces type I interferon (IFN)-mediated inhibition of Type17 immune responses, resulting in exacerbation of bacterial burden during influenza and Staphylococcus aureus super-infection in mice. Here, we investigated the role of the signaling mediator, STAT2, downstream of type I IFN signaling in both influenza infection and bacterial super-infection. Influenza-infected STAT2−/− mice had increased weight loss, increased viral burden, and increased inflammatory responses when compared to influenza-infected WT mice. Despite an exaggerated inflammatory response to influenza infection alone, influenza and methicillin resistant S. aureus (MRSA) super-infected STAT2−/− mice had lower bacterial burden, morbidity, and mortality than WT mice. STAT2−/− mice had an increased expression of markers associated with M1 (classical) and M2 (alternative) macrophages in lung during influenza or influenza-bacterial super infection compared to WT mice. Next, we found neutralization of IFNγ or Arginase1 decreased bacterial clearance in STAT2−/− mice in influenza-bacterial super-infection, suggesting that both M1 and M2 phenotype mediate host defense during influenza-bacterial super-infection. These data demonstrate a clear role for type I IFN induced STAT2 signaling in suppression of macrophage function during influenza, MRSA super-infection. Further, these studies reveal novel mechanistic insight into the roles of macrophage subpopulations in pulmonary host defense.

Roles of the Mevalonate Pathway and Cholesterol Trafficking in Pulmonary Host Defense.

The mevalonic acid synthesis pathway, cholesterol, and lipoproteins play fundamental roles in lung physiology and the innate immune response. Recent literature investigating roles for cholesterol synthesis and trafficking in host defense against respiratory infection was critically reviewed. The innate immune response and the cholesterol biosynthesis/trafficking network regulate one another, with important implications for pathogen invasion and host defense in the lung. The activation of pathogen recognition receptors and downstream cellular host defense functions are critically sensitive to cellular cholesterol. Conversely, microorganisms can co-opt the sterol/lipoprotein network in order to facilitate replication and evade immunity. Emerging literature suggests the potential for harnessing these insights towards therapeutic development. Given that >50% of adults in the U.S. have serum cholesterol abnormalities and pneumonia remains a leading cause of death, the potential impact of cholesterol on pulmonary host defense is of tremendous public health significance and warrants further mechanistic and translational investigation.

Regulatory T cells and IL10 suppress pulmonary host defense during early-life exposure to radical containing combustion derived ultrafine particulate matter

BackgroundExposure to elevated levels of particulate matter (PM) is associated with increased risk of morbidity and mortality due to respiratory tract viral infections in infants. Recent identification of environmentally persistent free radicals (EPFRs) in the PM from a variety of combustion sources suggests its role in the enhancement of disease severity of lower respiratory tract infections (LRTI). Our previous studies demonstrated that acute exposure to EPFRs induces pulmonary immunosuppression allowing for enhanced influenza disease severity. Here, we determine the mechanism of EPFR-induced immunosuppression and its impact on the immune response towards influenza infection.MethodsNeonatal mice (3 days old) were acutely exposed to DCB (combustion derived PM with chemisorbed EPFR) for seven consecutive days. Four days post-exposure (dpe), mice were infected with influenza virus. Pulmonary T cell phenotypes including regulatory T cells (Tregs) were analyzed by flow cytometry. The role of IL10 in EPFR-induced exacerbation of influenza disease severity was determined by administering recombinant IL10 (rIL10) to wild type mice or by using IL10 deficient (IL10−/−) neonatal mice. Mice were assessed for morbidity by measuring percent weight change and pulmonary viral load.ResultsNeonatal mice exposed to EPFRs had a significant increase in pulmonary Tregs and the immunosuppressive cytokine IL10 following influenza infection, which coincided with decreased protective T cell responses to influenza infection at 6 dpi. Depletion of Tregs in EPFR-exposed neonatal mice resulted in increased protective, adaptive T cell responses, whereas adoptive transfer of Tregs from EPFR-exposed neonates to air-exposed neonatal mice suppressed adaptive T cell responses towards influenza infection. Further, treatment with rIL10 could recapitulate EPFR-induced exacerbation of morbidity and pulmonary viral load compared to air exposed and influenza infected mice, whereas, EPFR-exposed IL10−/− neonates exhibited significant reductions in morbidity, pulmonary viral load and adaptive T cell responses following influenza infection.ConclusionsNeonatal exposure to EPFRs induced Tregs and IL10 resulting in suppressed adaptive T cell responses and enhanced influenza disease severity in neonatal mice. Depletion of Tregs increased adaptive T cell responses and deficiency of IL10 reduced morbidity and conferred enhanced protection against influenza virus.

IL-17 Receptor Signaling in the Lung Epithelium Is Required for Mucosal Chemokine Gradients and Pulmonary Host Defense against K.pneumoniae.

The cytokine IL-17, and signaling via its heterodimeric IL-17RA/IL-17RC receptor, is critical for host defense against extracellular bacterial and fungal pathogens. Polarized lung epithelial cells express IL-17RA and IL-17RC basolaterally. However, their contribution to IL-17-dependent pulmonary defenses invivo remains to be determined. To address this, we generated mice with conditional deletion of Il17ra or Il17rc in Scgb1a1-expressing club cells, a major component of the murine bronchiolar epithelium. These mice displayed an impaired ability to recruit neutrophils into the airway lumen in response to IL-17, a defect in bacterial clearance upon mucosal challenge with the pulmonary pathogen Klebsiella pneumoniae, and substantially reduced epithelial expression of the chemokine Cxcl5. Neutrophil recruitment and bacterial clearance were restored by intranasal administration of recombinant CXCL5. Our data show that IL-17R signaling in the lung epithelium plays a critical role in establishing chemokine gradients that are essential for mucosal immunity against pulmonary bacterial pathogens.