S. Kierstein

Absence of bacterially induced RELMβ reduces injury in the dextran sodium sulfate model of colitis

Although inflammatory bowel disease (IBD) is the result of a dysregulated immune response to commensal gut bacteria in genetically predisposed individuals, the mechanism(s) by which bacteria lead to the development of IBD are unknown. Interestingly, deletion of intestinal goblet cells protects against intestinal injury, suggesting that this epithelial cell lineage may produce molecules that exacerbate IBD. We previously reported that resistin-like molecule beta (RELMbeta; also known as FIZZ2) is an intestinal goblet cell-specific protein that is induced upon bacterial colonization whereupon it is expressed in the ileum and colon, regions of the gut most often involved in IBD. Herein, we show that disruption of this gene reduces the severity of colitis in the dextran sodium sulfate (DSS) model of murine colonic injury. Although RELMbeta does not alter colonic epithelial proliferation or barrier function, we show that recombinant protein activates macrophages to produce TNF-alpha both in vitro and in vivo. RELMbeta expression is also strongly induced in the terminal ileum of the SAMP1/Fc model of IBD. These results suggest a model whereby the loss of epithelial barrier function by DSS results in the activation of the innate mucosal response by RELMbeta located in the lumen, supporting the hypothesis that this protein is a link among goblet cells, commensal bacteria, and the pathogenesis of IBD.

IL-4 and IL-13 Form a Negative Feedback Circuit with Surfactant Protein-D in the Allergic Airway Response

The innate immune molecule surfactant protein-D (SP-D) plays an important regulatory role in the allergic airway response. In this study, we demonstrate that mice sensitized and challenged with either Aspergillus fumigatus (Af) or OVA have increased SP-D levels in their lung. SP-D mRNA and protein levels in the lung also increased in response to either rIL-4 or rIL-13 treatment. Type II alveolar epithelial cell expression of IL-4Rs in mice sensitized and challenged with Af, and in vitro induction of SP-D mRNA and protein by IL-4 and IL-13, but not IFN-γ, suggested a direct role of IL-4R-mediated events. The regulatory function of IL-4 and IL-13 was further supported in STAT-6-deficient mice as well as in IL-4/IL-13 double knockout mice that failed to increase SP-D production upon allergen challenge. Interestingly, addition of rSP-D significantly inhibited Af-driven Th2 cell activation in vitro whereas mice lacking SP-D had increased numbers of CD4+ cells with elevated IL-13 and thymus- and activation-regulated chemokine levels in the lung and showed exaggerated production of IgE and IgG1 following allergic sensitization. We propose that allergen exposure induces elevation in SP-D protein levels in an IL-4/IL-13-dependent manner, which in turn, prevents further activation of sensitized T cells. This negative feedback regulatory circuit could be essential in protecting the airways from inflammatory damage after allergen inhalation.

Chronic obstructive pulmonary disease and inhaled steroids alter surfactant protein D (SP-D) levels: a cross-sectional study

BackgroundSurfactant protein D (SP-D), an innate immune molecule, plays an important protective role during airway inflammation. Deficiency of this molecule induces emphysematous changes in murine lungs, but its significance in human COPD remains unclear.MethodsWe collected bronchoalveolar lavage fluid from 20 subjects with varying degrees of COPD (8 former smokers and 12 current smokers) and 15 asymptomatic healthy control subjects (5 never smokers, 3 remote former smokers, and 7 current smokers). All subjects underwent a complete medical history and pulmonary function testing. SP-D was measured by Enzyme-Linked ImmunoSorbent Assay. Statistical analysis was performed using nonparametric methods and multivariable linear regression for control of confounding. The effect of corticosteroid treatment on SP-D synthesis was studied in vitro using an established model of isolated type II alveolar epithelial cell culture.ResultsAmong former smokers, those with COPD had significantly lower SP-D levels than healthy subjects (median 502 and 1067 ng/mL, respectively, p = 0.01). In a multivariable linear regression model controlling for age, sex, race, and pack-years of tobacco, COPD was independently associated with lower SP-D levels (model coefficient -539, p = 0.04) and inhaled corticosteroid use was independently associated with higher SP-D levels (398, p = 0.046). To support the hypothesis that corticosteroids increase SP-D production we used type II alveolar epithelial cells isolated from adult rat lungs. These cells responded to dexamethasone treatment by a significant increase of SP-D mRNA (p = 0.041) and protein (p = 0.037) production after 4 days of culture.ConclusionAmong former smokers, COPD is associated with lower levels of SP-D and inhaled corticosteroid use is associated with higher levels of SP-D in the lung. Dexamethasone induced SP-D mRNA and protein expression in isolated epithelial cells in vitro. Given the importance of this molecule as a modulator of innate immunity and inflammation in the lung, low levels may play a role in the pathogenesis and/or progression of COPD. Further, we speculate that inhaled steroids may induce SP-D expression and that this mechanism may contribute to their beneficial effects in COPD. Larger, prospective studies are warranted to further elucidate the role of surfactant protein D in modulating pulmonary inflammation and COPD pathogenesis.

Social Stress Enhances Allergen-Induced Airway Inflammation in Mice and Inhibits Corticosteroid Responsiveness of Cytokine Production

Chronic psychosocial stress exacerbates asthma, but the underlying mechanisms remain poorly understood. We hypothesized that psychosocial stress aggravates allergic airway inflammation by altering innate immune cell function. The effects of stress on airway inflammation, lung function, and glucocorticoid responsiveness were studied in a novel in vivo murine model of combined social disruption stress and allergic sensitization. The effects of corticosterone were assessed on cytokine profile and glucocorticoid receptor activation in LPS-stimulated spleen cell cultures in vitro. Airway inflammation resolved 48 h after a single allergen provocation in sensitized control mice, but not in animals that were repeatedly exposed to stress before allergen challenge. The enhanced eosinophilic airway inflammation 48 h after allergen challenge in these mice was associated with increased levels of IL-5, GM-CSF, IgG1, thymus-activated and regulatory chemokine, TNF-α, and IL-6 in the airways and a diminished inhibition of these mediators by corticosterone in LPS-stimulated splenocyte cultures in vitro. Stress-induced reduction of the corticosteroid effects paralleled increased p65 expression and a decreased DNA-binding capability of the glucocorticoid receptor in vitro. Furthermore, glucocorticoid receptor mRNA and protein expression in the lungs of mice exposed to both stress and allergen was markedly reduced in comparison with that in either condition alone or in naive mice. Thus, exposure to repeated social stress before allergen inhalation enhances and prolongs airway inflammation and alters corticosterone responsiveness. We speculate that these effects were mediated at least in part by impaired glucocorticoid receptor expression and function.

Ozone inhalation induces exacerbation of eosinophilic airway inflammation and hyperresponsiveness in allergen-sensitized mice

Background:  Ozone (O3) exposure evokes asthma exacerbations by mechanisms that are poorly understood. We used a murine model to characterize the effects of O3 on allergic airway inflammation and hyperresponsiveness and to identify factors that might contribute to the O3‐induced exacerbation of asthma.

Surfactant protein D inhibits TNF-α production by macrophages and dendritic cells in mice

BACKGROUND Surfactant protein (SP) D shares target cells with the proinflammatory cytokine TNF-alpha, an important autocrine stimulator of dendritic cells and macrophages in the airways. OBJECTIVE We sought to study the mechanisms by which TNF-alpha and SP-D can affect cellular components of the pulmonary innate immune system. METHODS Cytokine and SP-D protein and mRNA expression was assessed by means of ELISA, Western blotting, and real-time PCR, respectively, by using in vivo models of allergic airway sensitization. Macrophage and dendritic cell phenotypes were analyzed by means of FACS analysis. Maturation of bone marrow-derived dendritic cells was investigated in vitro. RESULTS TNF-alpha, elicited either by allergen exposure or pulmonary overexpression, induced SP-D, IL-13, and mononuclear cell influx in the lung. Recombinant IL-13 by itself was also capable of enhancing SP-D in vivo and in vitro, and the SP-D response to allergen challenge was impaired in IL-13-deficient mice. Allergen-induced increase of SP-D in the airways coincided with resolution of TNF-alpha release and cell influx. SP-D-deficient mice had constitutively high numbers of alveolar mononuclear cells expressing TNF-alpha, MHC class II, CD86, and CD11b, characteristics of proinflammatory, myeloid dendritic cells. Recombinant SP-D significantly suppressed all of these molecules in bone marrow-derived dendritic cell cultures. CONCLUSIONS TNF-alpha can contribute to enhanced SP-D production in the lung indirectly through inducing IL-13. SP-D, on the other hand, can antagonize the proinflammatory effects of TNF-alpha on macrophages and dendritic cells, at least partly, by inhibiting production of this cytokine.

Essential role of IFNβ and CD38 in TNFα-induced airway smooth muscle hyper-responsiveness

We recently identified autocrine interferon (IFN)beta as a novel mechanism mediating tumor necrosis factor (TNF)alpha-induced expression of inflammatory genes in airway smooth muscle (ASM) cells, including CD38, known to regulate calcium signaling. Here, we investigated the putative involvement of IFNbeta in regulating TNFalpha-induced airway hyper-responsiveness (AHR), a defining feature of asthma. Using our pharmacodynamic model to assess ex vivo AHR isolated murine tracheal rings, we found that TNFalpha-induced enhanced contractile responses to carbachol and bradykinin was abrogated by neutralizing anti-IFNbeta antibody or in tracheal rings deficient in CD38. In cultured human ASM cells, where CD38 has been involved in TNFalpha-induced enhanced calcium signals to carbachol and bradykinin, we found that neutralizing anti-IFNbeta prevented TNFalpha enhancing action only on carbachol responses but not to that induced by bradykinin. In a well-characterized model of allergic asthma (mice sensitized and challenged with Aspergillus fumigatus (Af)), we found heightened expression of both IFNbeta and CD38 in the airways. Furthermore, allergen-associated AHR to methacholine, assessed by lung resistance and dynamic compliance, was completely suppressed in CD38-deficient mice, despite the preservation of airway inflammation. These data provide the first evidence that ASM-derived IFNbeta and CD38 may play a significant role in the development of TNFalpha-associated AHR.

Surfactant Protein-A inhibits Aspergillus fumigatus -induced allergic T-cell responses

BackgroundThe pulmonary surfactant protein (SP)-A has potent immunomodulatory activities but its role and regulation during allergic airway inflammation is unknown.MethodsWe studied changes in SP-A expression in the bronchoalveolar lavage (BAL) using a murine model of single Aspergillus fumigatus (Af) challenge of sensitized animals.ResultsSP-A protein levels in the BAL fluid showed a rapid, transient decline that reached the lowest values (25% of controls) 12 h after intranasal Af provocation of sensitized mice. Decrease of SP-A was associated with influx of inflammatory cells and increase of IL-4 and IL-5 mRNA and protein levels. Since levels of SP-A showed a significant negative correlation with these BAL cytokines (but not with IFN-γ), we hypothesized that SP-A exerts an inhibitory effect on Th2-type immune responses. To study this hypothesis, we used an in vitro Af-rechallenge model. Af-induced lymphocyte proliferation of cells isolated from sensitized mice was inhibited in a dose-dependent manner by addition of purified human SP-A (0.1–10 μg/ml). Flow cytometric studies on Af-stimulated lymphocytes indicated that the numbers of CD4+ (but not CD8+) T cells were significantly increased in the parental population and decreased in the third and fourth generation in the presence of SP-A. Further, addition of SP-A to the tissue culture inhibited Af-induced IL-4 and IL-5 production suggesting that SP-A directly suppressed allergen-stimulated CD4+ T cell function.ConclusionWe speculate that a transient lack of this lung collectin following allergen exposure of the airways may significantly contribute to the development of a T-cell dependent allergic immune response.

Surfactant protein D and asthma

Because of its structural uniqueness, specific localization and functional versatility, the lung collectin surfactant protein (SP)-D is an important major player of pulmonary immune processes. Recent publications indicated that collectins are potent modulators of the innate immune response in the lung and reports on human asthma and animal models suggested that they may be involved in allergen-induced airway changes. As this molecule is capable of inhibiting T cell activation and allergic inflammatory events, it is suggested that it may function as a local regulator of a T-helper type 2 (Th2) inflammation. Two reports in this issue of Clinical and Experimental Allergy provide further evidence for the importance of this molecule in allergic asthma. The first one by Koopmans et al. [1] shows that SP-D may be a valuable biomarker that increases in serum of patients following allergen exposure. The second paper by Schaub et al. [2] investigates the effects of the absence of this molecule in SP-D deficient mice during allergic airway inflammation.

Systemic FasL Neutralization Increases Eosinophilic Inflammation in a Mouse Model of Asthma

Eosinophils and lymphocytes are pathogenically important in allergic inflammation and sensitive to Fas‐mediated apoptosis. Fas ligand (FasL) activity therefore should play a role in regulating the allergic immune response. We aimed to characterize the role of FasL expression in airway eosinophilia in Aspergillus fumigatus (Af)‐induced sensitization and to determine whether FasL neutralization alters the inflammatory response.