Jane C. Deng

Type I IFNs mediate development of postinfluenza bacterial pneumonia in mice

Influenza-related complications continue to be a major cause of mortality worldwide. Due to unclear mechanisms, a substantial number of influenza-related deaths result from bacterial superinfections, particularly secondary pneumococcal pneumonia. Here, we report what we believe to be a novel mechanism by which influenza-induced type I IFNs sensitize hosts to secondary bacterial infections. Influenza-infected mice deficient for type I IFN-alpha/beta receptor signaling (Ifnar-/- mice) had improved survival and clearance of secondary Streptococcus pneumoniae infection from the lungs and blood, as compared with similarly infected wild-type animals. The less effective response in wild-type mice seemed to be attributable to impaired production of neutrophil chemoattractants KC (also known as Cxcl1) and Mip2 (also known as Cxcl2) following secondary challenge with S. pneumoniae. This resulted in inadequate neutrophil responses during the early phase of host defense against secondary bacterial infection. Indeed, influenza-infected wild-type mice cleared secondary pneumococcal pneumonia after pulmonary administration of exogenous KC and Mip2, whereas neutralization of Cxcr2, the common receptor for KC and Mip2, reversed the protective phenotype observed in Ifnar-/- mice. These data may underscore the importance of the type I IFN inhibitory pathway on CXC chemokine production. Collectively, these findings highlight what we believe to be a novel mechanism by which the antiviral response to influenza sensitizes hosts to secondary bacterial pneumonia.

Early recruitment of neutrophils determines subsequent T1/T2 host responses in a murine model of Legionella pneumophila pneumonia

The contribution of neutrophils to lethal sensitivity and cytokine balance governing T1 and T2 host responses was assessed in a murine model of Legionella pneumophila pneumonia. Neutrophil depletion by administration of granulocyte-specific mAb RB6-8C5 at 1 day before infection rendered mice ∼100-fold more susceptible to lethal pneumonia induced by L. pneumophila. However, this treatment did not alter early bacterial clearance, despite a substantial decrease in neutrophil influx at this time point. Cytokine profiles in the lungs of control mice demonstrated strong T1 responses, characterized by an increase of IFN-γ and IL-12. In contrast, neutrophil-depleted mice exhibited significantly lower levels of IFN-γ and IL-12, and elevation of T2 cytokines, IL-4 and IL-10. Immunohistochemistry of bronchoalveolar lavage cells demonstrated the presence of IL-12 in neutrophils, but not alveolar macrophages. Moreover, IL-12 was detected in lavage cell lysates by ELISA, which was paralleled to neutrophil number. However, intratracheal administration of recombinant murine IL-12 did not restore resistance, whereas reconstitution of IFN-γ drastically improved bacterial clearance and survival in neutrophil-depleted mice. Taken together, these data demonstrated that neutrophils play crucial roles in primary L. pneumophila infection, not via direct killing but more immunomodulatory effects. Our results suggest that the early recruitment of neutrophils may contribute to T1 polarization in a murine model of L. pneumophila pneumonia.

Chemokine-dependent neutrophil recruitment in a murine model of Legionella pneumonia: Potential role of neutrophils as immunoregulatory cells

ABSTRACT The roles of CXC chemokine-mediated host responses were examined with an A/J mouse model of Legionella pneumophilapneumonia. After intratracheal inoculation of 106 CFU ofL. pneumophila, the bacterial numbers in the lungs increased 10-fold by day 2; this increase was accompanied by the massive accumulation of neutrophils. Reverse transcription-PCR data demonstrated the up-regulation of CXC chemokines, such as keratinocyte-derived chemokine, macrophage inflammatory protein 2 (MIP-2), and lipopolysaccharide-induced CXC chemokine (LIX). Consistent with these data, increased levels of KC, MIP-2, and LIX proteins were observed in the lungs and peaked at days 1, 2, and 2, respectively. Although the administration of anti-KC or anti–MIP-2 antibody resulted in an approximately 20% decrease in neutrophil recruitment on day 2, no increase in mortality was observed. In contrast, the blockade of CXC chemokine receptor 2 (CXCR2), a receptor for CXC chemokines, including KC and MIP-2, strikingly enhanced mortality; this effect coincided with a 67% decrease in neutrophil recruitment. Interestingly, anti-CXCR2 antibody did not affect bacterial burden by day 2, even in the presence of a lethal challenge of bacteria. Moreover, a significant decrease in interleukin-12 (IL-12) levels, in contrast to the increases in KC, MIP-2, and LIX levels, was demonstrated for CXCR2-blocked mice. These data indicated that CXCR2-mediated neutrophil accumulation may play a crucial role in host defense against L. pneumophilapneumonia in mice. The increase in lethality without a change in early bacterial clearance suggested that neutrophils may exert their protective effect not through direct killing but through more immunomodulatory actions in L. pneumophila pneumonia. We speculate that a decrease in the levels of the protective cytokine IL-12 may explain, at least in part, the high mortality in the setting of reduced neutrophil recruitment.

CpG Oligodeoxynucleotides Stimulate Protective Innate Immunity against Pulmonary Klebsiella Infection

Bacterial pneumonia is a leading cause of mortality in the United States. Innate immune responses, including type-1 cytokine production, are critical to the effective clearance of bacterial pathogens from the lung. Synthetic oligodeoxynucleotides (ODN) containing unmethylated CpG dinucleotide motifs (CpG ODN), which mimic the effects of bacterial DNA, have been shown to enhance type-1 cytokine responses during infection due to intracellular pathogens, resulting in enhanced microbial clearance. The role of CpG ODN in modulating protective innate immunity against extracellular pathogens is unknown. Using a murine model of Gram-negative pneumonia, we found that CpG ODN administration stimulated protective immunity against Klebsiella pneumoniae. Specifically, intratracheal (i.t.) administration of CpG ODN (30 μg) 48 h before i.t. K. pneumoniae challenge resulted in increased survival, compared with animals pretreated with control ODN or saline. Pretreatment with CpG ODN resulted in enhanced bacterial clearance in lung and blood, and higher numbers of pulmonary neutrophils, NKT cells, γδ-T cells, and activated NK1.1+ cells and γδ-T lymphocytes during infection. Furthermore, pretreatment with CpG ODN enhanced the production of TNF-α, and type-1 cytokines, including IL-12, IFN-γ, and the IFN-γ-dependent ELR− CXC chemokines IFN-γ-inducible protein-10 and monokine induced by IFN-γ in response to Klebsiella challenge, compared with control mice. These findings indicate that i.t. administration of CpG ODN can stimulate multiple components of innate immunity in the lung, and may form the basis for novel therapies directed at enhancing protective immune responses to severe bacterial infections of the lung.

Hyperoxia Mediates Acute Lung Injury and Increased Lethality in Murine Legionella Pneumonia: The Role of Apoptosis

Legionella pneumophila is a major cause of life-threatening pneumonia, which is characterized by a high incidence of acute lung injury and resultant severe hypoxemia. Mechanical ventilation using high oxygen concentrations is often required in the treatment of patients with L. pneumophila pneumonia. Unfortunately, oxygen itself may propagate various forms of tissue damage, including acute lung injury. The effect of hyperoxia as a cofactor in the course of L. pneumophila pneumonia is poorly understood. In this study, we show that exposure to hyperoxic conditions during the evolution of pneumonia results in a marked increase in lethality in mice with Legionella pneumonia. The enhanced lethality was associated with an increase in lung permeability, but not changes in either lung bacterial burden or leukocyte accumulation. Interestingly, accelerated apoptosis as evidenced by assessment of histone-DNA fragments and caspase-3 activity were noted in the infected lungs of mice exposed to hyperoxia. TUNEL staining of infected lung sections demonstrated increased apoptosis in hyperoxic mice, predominantly in macrophages and alveolar epithelial cells. In vitro exposure of primary murine alveolar epithelial cells to Legionella in conjunction with hyperoxia accelerated apoptosis and loss of barrier function. Fas-deficient mice demonstrated partial resistance to the lethal effects of Legionella infection induced by hyperoxia, which was associated with attenuated apoptosis in the lung. These results demonstrate that hyperoxia serves as an important cofactor for the development of acute lung injury and lethality in L. pneumophila pneumonia. Exaggerated apoptosis, in part through Fas-mediated signaling, may accelerate hyperoxia-induced acute lung injury in Legionella pneumonia.

Transient transgenic expression of gamma interferon promotes legionella pneumophila clearance in immunocompetent hosts

ABSTRACT Gamma interferon (IFN-γ) and T1-phenotype immune responses are important components of host defense against a variety of intracellular pathogens, including Legionella pneumophila. The benefit of intrapulmonary adenovirus-mediated IFN-γ gene therapy was investigated in a nonlethal murine model of experimental L. pneumophilapneumonia. Intratracheal (i.t.) administration of 106 CFU of L. pneumophila induced the expression of T1 phenotype cytokines, such as IFN-γ and interleukin-12 (IL-12). Natural killer cells were identified as the major cellular source of IFN-γ. To determine if enhanced expression of IFN-γ in the lung could promote pulmonary clearance of L. pneumophila, we i.t. administered 5 × 108 PFU of a recombinant adenovirus vector containing the murine IFN-γ cDNA (AdmIFN-γ) concomitant with L. pneumophila. We observed a 10-fold decrease in lung bacterial CFU at day 2 in the AdmIFN-γ-treated group compared to controls (P < 0.01). Alveolar macrophages isolated from AdmIFN-γ-treated animals displayed enhanced killing of intracellular L. pneumophila organisms ex vivo. Similar improvements in bacterial clearance were observed with i.t. recombinant IFN-γ treatment. The transient transgenic expression of IL-12, a known inducer of IFN-γ and promoter of T1-type immune responses, resulted in more modest improvement in bacterial clearance (sixfold reduction;P < 0.05). These results demonstrate that, even in immunocompetent hosts, exogenous administration or transient transgenic expression of IFN-γ, and to a lesser extent IL-12, may be of potential therapeutic benefit in the treatment of patients withLegionella pneumonia.

Impaired Pulmonary Host Defense in Mice Lacking Expression of the CXC Chemokine Lungkine

Lungkine (CXCL15) is a novel CXC chemokine that is highly expressed in the adult mouse lung. To determine the biologic function of Lungkine, we generated Lungkine null mice by targeted gene disruption. These mice did not differ from wild-type mice in their hematocrits or in the relative number of cells in leukocyte populations of peripheral blood or other tissues, including lung and bone marrow. However, Lungkine null mice were more susceptible to Klebsiella pneumonia infection, with a decreased survival and increased lung bacterial burden compared with infected wild-type mice. Histologic analysis of the lung and assessment of leukocytes in the bronchioalveolar lavage revealed that neutrophil numbers were normal in the lung parenchyma, but reduced in the airspace. The production of other neutrophil chemoattractants in the Lungkine null mice did not differ from that in wild-type mice, and neutrophil migration into other tissues was normal. Taken together, these findings demonstrate that Lungkine is an important mediator of neutrophil migration from the lung parenchyma into the airspace.

Respiratory Viral Infections in Hematopoietic Stem Cell and Solid Organ Transplant Recipients

Respiratory viral infections (RVIs) are common causes of mild illness in immunocompetent children and adults with rare occurrences of significant morbidity or mortality. Complications are more common in the very young, very old, and those with underlying lung diseases. However, RVIs are increasingly recognized as a cause of morbidity and mortality in recipients of hematopoietic stem cell transplants (HSCT) and solid organ transplants (SOTs). Diagnostic techniques for respiratory syncytial virus (RSV), parainfluenza, influenza, and adenovirus have been clinically available for decades, and these infections are known to cause serious disease in transplant recipients. Modern molecular technology has now made it possible to detect other RVIs including human metapneumovirus, coronavirus, and bocavirus, and the role of these viruses in causing serious disease in transplant recipients is still being worked out. This article reviews the current information regarding epidemiology, pathogenesis, clinical presentation, diagnosis, and treatment of these infections, as well as the aspects of clinical significance of RVIs unique to HSCT or SOT.

Interferon-Inducible Protein 10, but Not Monokine Induced by Gamma Interferon, Promotes Protective Type 1 Immunity in Murine Klebsiella pneumoniae Pneumonia

ABSTRACT CXC chemokines that lack the ELR motif, including interferon-inducible protein 10 [IP-10 (CXCL10)] and monokine induced by gamma interferon (IFN-γ) [MIG (CXCL9)], have been shown to mediate the generation of type 1 immune responses. In this study, we found that intrapulmonary administration of the gram-negative bacterium Klebsiella pneumoniae resulted in the local and systemic expression of IP-10, followed sequentially by MIG expression. MIG mRNA expression in the lungs of Klebsiella-infected mice required the endogenous production of IFN-γ, whereas IP-10 was expressed in both an IFN-γ-dependent and an IFN-γ-independent fashion. Antibody-mediated neutralization of IP-10 resulted in reduced bacterial clearance and decreased survival, whereas bacterial clearance was unaltered in mice treated with anti-MIG antibody. Impaired bacterial clearance in anti-IP-10 antibody-treated mice was associated with significant reductions in the number and/or activational status of NK and NK-T cells, CD4+ T cells, and γδ T cells, as well as a reduction in the expression of IFN-γ. Conversely, the transient transgenic expression of murine IP-10 using adenovirus-mediated gene transfer resulted in improved bacterial clearance when IP-10 adenovirus was given concomitant with intrapulmonary bacterial challenge. These results indicate that IP-10 is an important component of innate immunity against extracellular bacterial pathogens of the lung and may represent a candidate molecule for immunotherapy in the setting of severe respiratory tract infection.

Subsequent infections in survivors of sepsis: epidemiology and outcomes.

Purpose: Sepsis is a devastating condition with considerable mortality. The causes of long-term mortality are poorly understood. To test the hypothesis that patients with sepsis are more susceptible to recurrent infections and death due to infectious complications, we investigated the outcomes of patients who survived sepsis, with regard to the incidence of recurrent infections and mortality. Materials and Methods: A retrospective study of the patients admitted to the intensive care unit (ICU) for sepsis from 2001 to 2002 who achieved 30-day survival (sepsis survivors [SSs], N = 78) and a control group of patients admitted to the ICU for noninfectious conditions with a similar severity of illness (N = 50) was performed. The primary end point was the number of recurrent infections in the first year posthospitalization. Results: The SSs group had higher rates of infections following hospital discharge compared to controls. Using a multivariable model, having survived sepsis was the strongest predictor of the development of subsequent infections (rate ratio [RR]: 2.83, P= .0006), the need for rehospitalization for infection in the year after the initial hospitalization (RR: 3.78, P = .0009), and postdischarge mortality (hazard ratio = 3.61, P = .003). Conclusions: Critically ill patients who survive sepsis have an increased risk of recurrent infections in the year following their septic episode that is associated with increased mortality.