B. Paige Lawrence

Activation of the aryl hydrocarbon receptor is essential for mediating the anti-inflammatory effects of a novel low-molecular-weight compound

VAF347 is a low-molecular-weight compound that inhibits allergic lung inflammation in vivo. This effect is likely the result of a block of dendritic cell (DC) function to generate proinflammatory T-helper (Th) cells because VAF347 inhibits interleukin (IL)-6, CD86, and human leukocyte antigen (HLA)-DR expression by human monocyte-derived DC, 3 relevant molecules for Th-cell generation. Here we demonstrate that VAF347 interacts with the aryl hydrocarbon receptor (AhR) protein, resulting in activation of the AhR signaling pathway. Functional AhR is responsible for the biologic activity of VAF347 because (1) other AhR agonists display an identical activity profile in vitro, (2) gene silencing of wild-type AhR expression or forced overexpression of a trans-dominant negative AhR ablates VAF347 activity to inhibit cytokine induced IL-6 expression in a human monocytic cell line, and (3) AhR-deficient mice are resistant to the compounds ability to block allergic lung inflammation in vivo. These data identify the AhR protein as key molecular target of VAF347 and its essential role for mediating the anti-inflammatory effects of the compound in vitro and in vivo.

Neonatal Hyperoxia Enhances the Inflammatory Response in Adult Mice Infected with Influenza A Virus

RATIONALE Lungs of adult mice exposed to hyperoxia as newborns are simplified and exhibit reduced function much like that observed in people who had bronchopulmonary dysplasia (BPD) as infants. Because survivors of BPD also show increased risk for symptomatic respiratory infections, we investigated how neonatal hyperoxia affected the response of adult mice infected with influenza A virus infection. OBJECTIVES To determine whether neonatal hyperoxia increased the severity of influenza A virus infection in adult mice. METHODS Adult female mice exposed to room air or hyperoxia between Postnatal Days 1 and 4 were infected with a sublethal dose of influenza A virus. MEASUREMENTS AND MAIN RESULTS The number of macrophages, neutrophils, and lymphocytes observed in airways of infected mice that had been exposed to hyperoxia as neonates was significantly greater than in infected siblings that had been exposed to room air. Enhanced inflammation correlated with increased levels of monocyte chemotactic protein-1 (CCL2) in lavage fluid, whereas infection-associated changes in IFN-gamma, IL-1beta, IL-6, tumor necrosis factor-alpha, KC, granulocyte-macrophage colony-stimulating factor, and macrophage inflammatory protein-1alpha, and production of virus-specific antibodies, were largely unaffected. Increased mortality of mice exposed to neonatal hyperoxia occurred by Day 14 of infection, and was associated with persistent inflammation and fibrosis. CONCLUSIONS These data suggest that the disruptive effect of hyperoxia on neonatal lung development also reprograms key innate immunoregulatory pathways in the lung, which may contribute to exacerbated pathology and poorer resistance to respiratory viral infections typically seen in people who had BPD.

The Effects of Maternal Exposure to Bisphenol A on Allergic Lung Inflammation into Adulthood

Bisphenol A (BPA) is a high-production volume chemical classified as an environmental estrogen and used primarily in the plastics industry. BPAs increased usage correlates with rising BPA levels in people and a corresponding increase in the incidence of asthma. Due to limited studies, the contribution of maternal BPA exposure to allergic asthma pathogenesis is unclear. Using two established mouse models of allergic asthma, we examined whether developmental exposure to BPA alters hallmarks of allergic lung inflammation in adult offspring. Pregnant C57BL/6 dams were gavaged with 0, 0.5, 5, 50, or 500 μg BPA/kg/day from gestational day 6 until postnatal day 21. To induce allergic inflammation, adult offspring were mucosally sensitized with inhaled ovalbumin containing low-dose lipopolysaccharide or ip sensitized using ovalbumin with alum followed by ovalbumin aerosol challenge. In the mucosal sensitization model, female offspring that were maternally exposed to ≥ 50 μg BPA/kg/day displayed enhanced airway lymphocytic and lung inflammation, compared with offspring of control dams. Peritoneally sensitized, female offspring exposed to ≤ 50 μg BPA/kg/day presented dampened lung eosinophilia, compared with vehicle controls. Male offspring did not exhibit these differences in either sensitization model. Our data demonstrate that maternal exposure to BPA has subtle and qualitatively different effects on allergic inflammation, which are critically dependent upon route of allergen sensitization and sex. However, these subtle, yet persistent changes due to developmental exposure to BPA did not lead to significant differences in overall airway responsiveness, suggesting that early life exposure to BPA does not exacerbate allergic inflammation into adulthood.

Aryl Hydrocarbon Receptor Activation during Influenza Virus Infection Unveils a Novel Pathway of IFN-γ 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-γ levels. We report here that the majority of IFN-γ-producing cells in the lung are neutrophils and macrophages not lymphocytes, and elevated IFN-γ 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-γ production by B cells, γδ 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-γ production. We also show that AhR-mediated increases in IFN-γ 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-γ 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.

Environmental toxicants and the developing immune system: A missing link in the global battle against infectious disease?

There is now compelling evidence that developmental exposure to chemicals from our environment contributes to disease later in life, with animal models supporting this concept in reproductive, metabolic, and neurodegenerative diseases. In contrast, data regarding how developmental exposures impact the susceptibility of the immune system to functional alterations later in life are surprisingly scant. Given that the immune system forms an integrated network that detects and destroys invading pathogens and cancer cells, it provides the bodys first line of defense. Thus, the consequences of early life exposures that reduce immune function are profound. This review summarizes available data for pollutants such as cigarette smoke and dioxin-like compounds, which consistently support the idea that developmental exposures critically impact the immune system. These findings suggest that exposure to common chemicals from our daily environment represent overlooked contributors to the fact that infectious diseases remain among the top five causes of death worldwide.

Examining the relationship between impaired host resistance and altered immune function in mice treated with TCDD.

Exposure to TCDD suppresses the immune response to numerous antigens, including bacterial and viral pathogens. Although we administer a non-lethal infection with influenza A virus, we often observe significant mortality in TCDD-treated animals. With the goal of identifying which TCDD-induced defects impair host resistance, we conducted a dose response study to examine whether alteration of particular immunological endpoints could be correlated with mortality. C57Bl/6 mice were treated with vehicle control, or 1, 2.5, 5, 7.5 or 10 microg/kg TCDD 1 day prior to intranasal (i.n.) infection with influenza virus. Survival was monitored for 9 days, when remaining mice were sacrificed and multiple endpoints evaluated. Lymphocyte migration to the lung and the production of virus-specific IgG2a, IgG1, and IgG2b antibodies were significantly diminished, even at the lower doses. IgA was enhanced in all groups treated with TCDD. In contrast, T cell expansion in the lymph node, and the production of IFNgamma and IL-12 were relatively resistant to suppression. Treatment with TCDD also enhanced pulmonary neutrophilia in infected mice. These results suggest that decreased antibody production and hyperinflammation may contribute to the death of TCDD-treated mice, and underscore the importance of evaluating numerous endpoints before concluding that a chemical is or is not immunotoxic.

The aryl hydrocarbon receptor is a modulator of anti-viral immunity ☆

Although immune modulation by AhR ligands has been studied for many years, the impact of AhR activation on host defenses against viral infection has not, until recently, garnered much attention. The development of novel reagents and model systems, new information regarding anti-viral immunity, and a growing appreciation for the global health threat posed by viruses have invigorated interest in understanding how environmental signals affect susceptibility to and pathological consequences of viral infection. Using influenza A virus as a model of respiratory viral infection, recent studies show that AhR activation cues signaling events in both leukocytes and non-immune cells. Functional alterations include suppressed lymphocyte responses and increased inflammation in the infected lung. AhR-mediated events within and extrinsic to hematopoietic cells has been investigated using bone marrow chimeras, which show that AhR alters different elements of the immune response by affecting different tissue targets. In particular, suppressed CD8(+) T cell responses are due to deregulated events within leukocytes themselves, whereas increased neutrophil recruitment to and IFN-gamma levels in the lung result from AhR-regulated events extrinsic to bone marrow-derived cells. This latter discovery suggests that epithelial and endothelial cells are overlooked targets of AhR-mediated changes in immune function. Further support that AhR influences host cell responses to viral infection are provided by several studies demonstrating that AhR interacts directly with viral proteins and affects viral latency. While AhR clearly modulates host responses to viral infection, we still have much to understand about the complex interactions between immune cells, viruses, and the host environment.

Aryl Hydrocarbon Receptor Activation Impairs the Priming but Not the Recall of Influenza Virus-Specific CD8+ T Cells in the Lung

The response of CD8+ T cells to influenza virus is very sensitive to modulation by aryl hydrocarbon receptor (AhR) agonists; however, the mechanism underlying AhR-mediated alterations in CD8+ T cell function remains unclear. Moreover, very little is known regarding how AhR activation affects anamnestic CD8+ T cell responses. In this study, we analyzed how AhR activation by the pollutant 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) alters the in vivo distribution and frequency of CD8+ T cells specific for three different influenza A virus epitopes during and after the resolution of a primary infection. We then determined the effects of TCDD on the expansion of virus-specific memory CD8+ T cells during recall challenge. Adoptive transfer of AhR-null CD8+ T cells into congenic AhR+/+ recipients, and the generation of CD45.2AhR−/−→CD45.1AhR+/+ chimeric mice demonstrate that AhR-regulated events within hemopoietic cells, but not directly within CD8+ T cells, underlie suppressed expansion of virus-specific CD8+ T cells during primary infection. Using a dual-adoptive transfer approach, we directly compared the responsiveness of virus-specific memory CD8+ T cells created in the presence or absence of TCDD, which revealed that despite profound suppression of the primary response to influenza virus, the recall response of virus-specific CD8+ T cells that form in the presence of TCDD is only mildly impaired. Thus, the delayed kinetics of the recall response in TCDD-treated mice reflects the fact that there are fewer memory cells at the time of reinfection rather than an inherent defect in the responsive capacity of virus-specific memory CD8+ cells.

Aryl Hydrocarbon Receptor Activation Reduces Dendritic Cell Function during Influenza Virus Infection

It has long been known that activation of the aryl hydrocarbon receptor (AhR) by ligands such as 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) suppresses T cell-dependent immune responses; however, the underlying cellular targets and mechanism remain unclear. We have previously shown that AhR activation by TCDD reduces the proliferation and differentiation of influenza virus-specific CD8(+) T cells through an indirect mechanism; suggesting that accessory cells are critical AhR targets during infection. Respiratory dendritic cells (DCs) capture antigen, migrate to lymph nodes, and play a key role in activating naive CD8(+) T cells during respiratory virus infection. Herein, we report an examination of how AhR activation alters DCs in the lung and affects their trafficking to and function in the mediastinal lymph nodes (MLN) during infection with influenza virus. We show that AhR activation impairs lung DC migration and reduces the ability of DCs isolated from the MLN to activate naive CD8(+) T cells. Using novel AhR mutant mice, in which the AhR protein lacks its DNA-binding domain, we show that the suppressive effects of TCDD require that the activated AhR complex binds to DNA. These new findings suggest that AhR activation by chemicals from our environment impacts DC function to stimulate naive CD8(+) T cells and that immunoregulatory genes within DCs are critical targets of AhR. Moreover, our results reinforce the idea that environmental signals and AhR ligands may contribute to differential susceptibilities and responses to respiratory infection.

Developmental exposure to bisphenol A modulates innate but not adaptive immune responses to influenza A virus infection.

Bisphenol A (BPA) is used in numerous products, such as plastic bottles and food containers, from which it frequently leaches out and is consumed by humans. There is a growing public concern that BPA exposure may pose a significant threat to human health. Moreover, due to the widespread and constant nature of BPA exposure, not only adults but fetuses and neonates are also exposed to BPA. There is mounting evidence that developmental exposures to chemicals from our environment, including BPA, contribute to diseases late in life; yet, studies of how early life exposures specifically alter the immune system are limited. Herein we report an examination of how maternal exposure to a low, environmentally relevant dose of BPA affects the immune response to infection with influenza A virus. We exposed female mice during pregnancy and through lactation to the oral reference dose for BPA listed by the US Environmental Protection Agency, and comprehensively examined immune parameters directly linked to disease outcomes in adult offspring following infection with influenza A virus. We found that developmental exposure to BPA did not compromise disease-specific adaptive immunity against virus infection, or reduce the host’s ability to clear the virus from the infected lung. However, maternal exposure to BPA transiently reduced the extent of infection-associated pulmonary inflammation and anti-viral gene expression in lung tissue. From these observations, we conclude that maternal exposure to BPA slightly modulates innate immunity in adult offspring, but does not impair the anti-viral adaptive immune response, which is critical for virus clearance and survival following influenza virus infection.