John Goulding

Sustained desensitization to bacterial Toll-like receptor ligands after resolution of respiratory influenza infection

The World Health Organization estimates that lower respiratory tract infections (excluding tuberculosis) account for ∼35% of all deaths caused by infectious diseases. In many cases, the cause of death may be caused by multiple pathogens, e.g., the life-threatening bacterial pneumonia observed in patients infected with influenza virus. The ability to evolve more efficient immunity on each successive encounter with antigen is the hallmark of the adaptive immune response. However, in the absence of cross-reactive T and B cell epitopes, one lung infection can modify immunity and pathology to the next for extended periods of time. We now report for the first time that this phenomenon is mediated by a sustained desensitization of lung sentinel cells to Toll-like receptor (TLR) ligands; this is an effect that lasts for several months after resolution of influenza or respiratory syncytial virus infection and is associated with reduced chemokine production and NF-κB activation in alveolar macrophages. Although such desensitization may be beneficial in alleviating overall immunopathology, the reduced neutrophil recruitment correlates with heightened bacterial load during secondary respiratory infection. Our data therefore suggests that post-viral desensitization to TLR signals may be one possible contributor to the common secondary bacterial pneumonia associated with pandemic and seasonal influenza infection.

Immune homeostasis in the respiratory tract and its impact on heterologous infection

Innate immunity at mucosal surfaces requires additional restraint to prevent inflammation to innocuous antigens or commensal microorganisms. The threshold above which airway macrophages become activated is raised by site-specific factors including the receptors for transforming growth factor beta, interleukin 10 and CD200; the ligands for which are produced by, or expressed on, respiratory epithelium. We discuss such site-specific regulation and how this is continually altered by prior infections. Resetting of innate reactivity represents a strategy for limiting excessive inflammation, but in some may pre-dispose to secondary bacterial pneumonia.

The impact of successive infections on the lung microenvironment

The effect of infection history on the immune response is ignored in most models of infectious disease and in preclinical vaccination studies. No one, however, is naïve and repeated microbial exposure, in particular during childhood, shapes the immune system to respond more efficiently later in life. Concurrent or sequential infections influence the immune response to secondary unrelated pathogens. The involvement of cross‐reactive acquired immunity, in particular T‐cell responses, is extensively documented. In this review, we discuss the impact of successive infections on the infected tissue itself, with a particular focus on the innate response of the respiratory tract, including a persistent alteration of (1) epithelial or macrophage expression of Toll‐like receptors or adherence molecules used by subsequent bacteria to invade the host, (2) the responsiveness of macrophages and neutrophils and (3) the local cytokine milieu that affects the activation of local antigen‐presenting cells and hence adaptive immunity to the next infection. We emphasize that such alterations not only occur during coinfection, but are maintained long after the initial pathogen is cleared. As innate responses are crucial to the fight against local pathogens but are also involved in the maintenance of the homeostasis of mucosal tissues, dysregulation of these responses by repeated infections is likely to have a major impact on the outcome of infectious or allergic disease.

Lowering the Threshold of Lung Innate Immune Cell Activation Alters Susceptibility to Secondary Bacterial Superinfection

BACKGROUND Previous studies have shown that the interaction of CD200R, a myeloid inhibitory receptor, with its ligand, CD200, is critical in the control of innate immune activation in the lung. METHODS AND RESULTS Using a mouse model of bacterial superinfection following influenza, we show that an absence of CD200R (a negative regulator highly expressed by macrophages and dendritic cells), restricts commensal and exogenous bacterial invasiveness and completely prevents the mortality observed in wild-type mice. This benefit is due to a heightened innate immune response to influenza virus in cd200r knockout mice that limits immune pathogenesis and viral load. In wild-type mice, apoptotic cells expressing CD200 that we believe contribute to the suppressed innate immune response to bacteria dominate during the resolution phase of influenza-induced inflammation. We also show for the first time the presence of a variety of previously unidentified bacterial species in the lower airways that are significantly adjusted by influenza virus infection and may contribute to the pathophysiology of disease. CONCLUSIONS The interaction of CD200 with CD200R therefore contributes to the hyporesponsive innate immune state following influenza virus infection that predisposes to secondary bacterial infection, a phenomenon that has the potential for immune modulation.

Benzodiazepine Augmented γ-amino-butyric Acid Signaling Increases Mortality From Pneumonia in Mice*

Objectives:Benzodiazepines are used for treating anxiety, epilepsy, muscle spasm, alcohol withdrawal, palliation, insomnia, and sedation as they allosterically modulate &ggr;-amino-butyric acid type A (GABAA) receptors. Despite widespread use, the importance and mechanism of their immune side-effects are poorly understood. Herein we sought to elucidate the impact and mechanism of benzodiazepine-induced susceptibility to infection at anxiolytic doses in mice. Design:Animal randomized controlled trial. Setting:Laboratory. Subjects:Adult female C57BL/6 and BALB/c mice. Interventions:The effect of a subsedative, anxiolytic dose of diazepam (2 mg kg−1 intraperitoneal) was investigated in a murine Streptococcus pneumoniae pneumonia model. Measurement and Main Results:Mortality, bacterial and cytokine load, cell recruitment, and intracellular pH were measured. Diazepam treatment did not affect immune homeostasis in the lung. However, diazepam increased mortality and bacterial load from S. pneumoniae pneumonia. The increases in mortality and bacterial load were reversed by a GABAA antagonist, bicuculline, indicating dependence on GABAA receptor signaling. While cell recruitment was unaltered by diazepam, the cytokine response to infection was affected, suggesting that local responses to the pathogen were perturbed. Macrophage and monocytes expressed benzodiazepine sensitive (&agr;1-&ggr;2) GABAA receptors. Interestingly macrophage GABAA receptor expression was regulated by bacterial toll-like receptor agonists and cytokines indicating an endogenous role in the immune response. Functionally diazepam appeared to counteract the endogenous down-regulation of GABAA signaling during infection. Consistent with augmented GABAA signaling, diazepam provoked intracellular acidosis in macrophage, leading to impaired cytokine production, bacterial phagocytosis and killing. In contrast, selective benzodiazepines that do not target the &agr;1 GABAA subunit did not affect macrophage function ex vivo or increase susceptibility to pneumonia in vivo. Conclusions:Our data highlight the regulation of macrophage function by GABAA receptor signaling and the potential harm of benzodiazepine exposure during pneumonia. Therapeutically, selective drugs may improve the safety profile of benzodiazepines.

Altered regulation of Toll-like receptor responses impairs antibacterial immunity in the allergic lung

The lung is colonized by commensal bacteria, some of which are associated with asthma exacerbations. Using the intranasal house-dust mite–sensitized mouse model of allergic airway disease, we show an imbalance in novel antibacterial pathways that culminates in a reduction in neutrophil recruitment to the airspaces and leads to bacterial invasion and dissemination. The expression of TREM (Triggering Receptor Expressed on Myeloid cells)-1 that amplifies Toll-like receptor (TLR) signaling and TREM-2 that inhibits this process is reversed. Furthermore, endogenous TLR inhibitors (A20, Tollip, SOCS1, and IRAK-M) and proteins involved in receptor recycling (TRIAD3) are raised. Consequently, the production of neutrophil chemoattractants is reduced. Intranasal administration of either chemokine restores the ability to recruit neutrophils, which prevents bacterial invasion. A background of allergic airway disease therefore exacerbates bacterial infection by altering key antibacterial innate immune pathways that are amenable to therapeutic intervention.

IL-9 Regulates Pathology during Primary and Memory Responses to Respiratory Syncytial Virus Infection

IL-9 is a cytokine of great current interest associated with allergic/Th2 responses. High levels of IL-9 are present in bronchial secretions from infants with respiratory syncytial virus (RSV) bronchiolitis. To test its effects in RSV disease with a Th2 profile, BALB/c mice were vaccinated with recombinant vaccinia virus expressing the RSV G protein. On RSV challenge, immunized mice developed augmented disease characterized by enhanced pulmonary Th2 and local IL-9 production peaking on days 7–10 of RSV infection. Depletion with anti-IL-9 Ab at vaccination or RSV challenge enhanced viral clearance. Depletion only at challenge had no effect on disease severity, whereas depletion at immunization and challenge enhanced Th1 responses, inhibited virus-specific IgG1 production, and enhanced disease severity. By contrast, depletion of IL-9 at immunization boosted IgG2a and inhibited the Th2 response and disease during subsequent infection without a concomitant increase in type 1 cytokines. Adoptive transfer of secondary memory CD4 T cells from the spleens of IL-9-depleted mice into naive recipients replicated many of the effects of depletion, indicating that IL-9 acts via CD4 T cells. Therefore, IL-9 is a previously unknown but key modulator of antiviral immunity, regulating T and B cell responses and having potent and specific effects on viral lung disease.

Structured regulation of inflammation during respiratory viral infection.

Innate immune cells including macrophages, dendritic cells, and granulocytes are resident within or patrol very different microenvironments in the host. Their activity or responsiveness to antigen is dictated by site-specific factors. Because of the constant exposure to environmental antigens and commensal microorganisms, mucosal immunity needs to be more constrained than peripheral counterparts to prevent unnecessary inflammation. The epithelial surfaces that dominate all mucosal tissues provide an ideal regulator since innate immune cells are often in intimate contact with, or lie immediately beneath, them and a breach in epithelial integrity would signal a damaging event and release innate immunity from their influence. We discuss the role of the respiratory epithelium in raising the threshold of innate immune cell activation at homoeostasis, how its absence triggers innate immunity, and how inflammatory resolution often produces an altered homoeostatic environment that can affect the next inflammatory event at this site.

CD8 T Cells Are Essential for Recovery from a Respiratory Vaccinia Virus Infection

The precise immune components required for protection against a respiratory Orthopoxvirus infection, such as human smallpox or monkeypox, remain to be fully identified. In this study, we used the virulent Western Reserve strain of vaccinia virus (VACV-WR) to model a primary respiratory Orthopoxvirus infection. Naive mice infected with VACV-WR mounted an early CD8 T cell response directed against dominant and subdominant VACV-WR Ags, followed by a CD4 T cell and Ig response. In contrast to other VACV-WR infection models that highlight the critical requirement for CD4 T cells and Ig, we found that only mice deficient in CD8 T cells presented with severe cachexia, pulmonary inflammation, viral dissemination, and 100% mortality. Depletion of CD8 T cells at specified times throughout infection highlighted that they perform their critical function between days 4 and 6 postinfection and that their protective requirement is critically dictated by initial viral load and virulence. Finally, the ability of adoptively transferred naive CD8 T cells to protect RAG−/− mice against a lethal VACV-WR infection demonstrated that they are both necessary and sufficient in protecting against a primary VACV-WR infection of the respiratory tract.

Bacterial complications during pandemic influenza infection

Evaluation of: Morens DM, Taubenberger JK, Fauci AS: Predominant role of bacterial pneumonia as a cause of death in pandemic influenza: implications for pandemic influenza preparedness. J. Infect. Dis. 198(7), 962-970 (2008). Secondary bacterial pneumonia is a common occurrence following lung influenza virus infection and leads to a significantly worse prognosis. This recent re-analysis of postmortem specimens and a vast number of reports from past influenza pandemics shows an extremely high frequency of lung colonization by bacterial species that are commonly found in the nasopharynx. This polymicrobial condition occurred in the preantibiotic era 1918-1919 influenza pandemic, but there is also evidence of bacterial co-infections in those outbreaks that occurred after antibiotic introduction. As such, antibiotic treatment should be included in any pandemic preparedness strategy. However, the choice of which antibiotic to use is important since some may even heighten morbidity and mortality.