IL-13 fusion cytotoxin ameliorates chronic fungal-induced allergic airway disease in mice.

IL-13 has emerged as a major contributor to allergic and asthmatic responses, and as such it represents an attractive target in these diseases. In this study, IL-13-responsive cells in the lung were targeted via the intranasal administration of IL-13-PE38QQR (IL-13-PE), comprised of human IL-13 and a derivative of Pseudomonas exotoxin, to Aspergillus fumigatus-sensitized mice challenged with A. fumigatus spores, or conidia. Mice received 50, 100, or 200 ng of IL-13-PE or diluent alone (i.e., control group) on alternate days from day 14 to day 28 after the conidia challenge. The control group of mice exhibited significant airway hyperreactivity, goblet cell hyperplasia, and peribronchial fibrosis at day 28 after conidia. Although the two lower doses of IL-13-PE had limited therapeutic effects in mice with fungal-induced allergic airway disease, the highest dose of IL-13-PE tested significantly reduced all features of airway disease compared with the control group. Whole lung mRNA expression of IL-4Ralpha and IL-13Ralpha1 was markedly reduced, whereas bronchoalveolar lavage and whole lung levels of IFN-gamma were significantly elevated in mice treated with 200 ng of IL-13-PE compared with the control group. This study demonstrates that a therapy designed to target IL-13-responsive cells in the lung ameliorates established fungal-induced allergic airway disease in mice.     

Antifungal and airway remodeling roles for murine monocyte chemoattractant protein-1/CCL2 during pulmonary exposure to Asperigillus fumigatus conidia

Asperigillus fumigatus spores or conidia are quickly eliminated from the airways of nonsensitized individuals but persist in individuals with allergic pulmonary responsiveness to fungus. A. fumigatus-induced allergic airway disease is characterized by persistent airway hyperreactivity, inflammation, and fibrosis. The present study explored the role of CCR2 ligands in the murine airway response to A. fumigatus conidia. Nonsensitized and A. fumigatus-sensitized CBA/J mice received an intratracheal challenge of A. fumigatus conidia, and pulmonary changes were analyzed at various times after conidia. Whole lung levels of monocyte chemoattractant protein-1 (MCP-1/CCL2), but neither MCP-3/CCL7 nor MCP-5/CCL12, were significantly elevated at days 3 and 7 after conidia in nonsensitized mice. MCP-1/CCL2 was significantly increased in lung samples from A. fumigatus-sensitized mice at days 14 and 30 after a conidia challenge. Administration of anti-MCP-1/CCL2 antiserum to nonsensitized mice for14 days after the conidia challenge attenuated the clearance of conidia and significantly increased airway hyperreactivity, eosinophilia, and peribronchial fibrosis compared with nonsensitized mice that received conidia and normal serum. Adenovirus-directed overexpression of MCP-1/CCL2 in A. fumigatus-sensitized mice markedly reduced the number of conidia, airway inflammation, and airway hyperresponsiveness at day 7 after the conidia challenge in these mice. Immunoneutralization of MCP-1/CCL2 levels in A. fumigatus-sensitized mice during days14-30 after the conidia challenge did not affect the conidia burden but significantly reduced airway hyperreactivity, lung IL-4 levels, and lymphocyte recruitment into the airways compared with the control group. These data suggest that MCP-1/CCL2 participates in the pulmonary antifungal and allergic responses to A. fumigatus conidia.     

Airway remodeling is absent in CCR1-/- mice during chronic fungal allergic airway disease

Asthmatic-like reactions characterized by elevated IgE, Th2 cytokines, C-C chemokines, eosinophilic inflammation, and persistent airway hyperresponsiveness follow pulmonary exposure to the spores or conidia from Aspergillus fumigatus fungus in sensitized individuals. In addition to these features, subepithelial fibrosis and goblet cell hyperplasia characterizes fungal-induced allergic airway disease in mice. Because lung concentrations of macrophage inflammatory protein-1alpha and RANTES were significantly elevated after A. fumigatus-sensitized mice received an intrapulmonary challenge with A. fumigatus spores or conidia, the present study addressed the role of their receptor, C-C chemokine receptor 1 (CCR1), in this model. A. fumigatus-sensitized CCR1 wild-type (+/+) and CCR1 knockout (-/-) mice exhibited similar increases in serum IgE and polymorphonuclear leukocyte numbers in the bronchoalveolar lavage. Airway hyperresponsiveness was prominent in both groups of mice at 30 days after an intrapulmonary challenge with A. fumigatus spores or conidia. However, whole lung levels of IFN-gamma were significantly higher whereas IL-4, IL-13, and Th2-inducible chemokines such as C10, eotaxin, and macrophage-derived chemokine were significantly lower in whole lung samples from CCR1-/- mice compared with CCR1+/+ mice at 30 days after the conidia challenge. Likewise, significantly fewer goblet cells and less subepithelial fibrosis were observed around large airways in CCR1-/- mice at the same time after the conidia challenge. Thus, these findings demonstrate that CCR1 is a major contributor to the airway remodeling responses that arise from A. fumigatus-induced allergic airway disease.     

Blocking IL-15 prevents the induction of allergen-specific T cells and allergic inflammation in vivo

IL-15 has been shown to accelerate and boost allergic sensitization in mice. Using a murine model of allergic sensitization to OVA, we present evidence that blocking endogenous IL-15 during the sensitization phase using a soluble IL-15Ralpha (sIL-15Ralpha) suppresses the induction of Ag-specific, Th2-differentiated T cells. This significantly reduces the production of OVA-specific IgE and IgG and prevents the induction of a pulmonary inflammation. Release of proinflammatory TNF-alpha, IL-1beta, IL-6, and IL-12 as well as that of Th2 cytokines IL-4, IL-5, and IL-13 into the bronchi are significantly reduced, resulting in suppressed recruitment of eosinophils and lymphocytes after allergen challenge. It is of clinical relevance that the airway hyper-responsiveness, a major symptom of human asthma bronchiale, is significantly reduced by sIL-15Ralpha treatment. Ex vivo analysis of the draining lymph nodes revealed reduced numbers of CD8, but not CD4, memory cells and the inability of T cells of sIL-15Ralpha-treated mice to proliferate and to produce Th2 cytokines after in vitro OVA restimulation. This phenomenon is not mediated by enhanced numbers of CD4(+)/CD25(+) T cells. These results show that IL-15 is important for the induction of allergen-specific, Th2-differentiated T cells and induction of allergic inflammation in vivo.     

CXCR2 is necessary for the development and persistence of chronic fungal asthma in mice.

The role of CXCR during allergic airway and asthmatic diseases is yet to be fully characterized. Therefore, the present study addressed the role of CXCR2 during Aspergillus fumigatus-induced asthma. Mice deficient in CXCR2 (CXCR2-/-) and wild-type counterparts (CXCR2+/+) were sensitized to A. fumigatus Ags and challenged with A. fumigatus conidia, and the resulting allergic airway disease was monitored for up to 37 days. At days 3 and 7 after conidia, CXCR2-/- mice exhibited significantly greater methacholine-induced airway hyperreactivity than did CXCR2+/+ mice. In contrast, CXCR2-deficient mice exhibited significantly less airway hyperresponsiveness than the wild-type control groups at days 14 and 37 after conidia. At all times after conidia, whole lung levels of IL-4, IL-5, and eotaxin/CC chemokine ligand 11 were significantly lower in CXCR2-/- mice than in the wild-type controls. Eosinophil and T cell, but not neutrophil, recruitment into the airways of A. fumigatus-sensitized CXCR2-/- mice was significantly impaired compared with wild-type controls at all times after the conidia challenge. Whole lung levels of IFN-gamma, inflammatory protein-10/CXC ligand (CXCL) 10, and monokine induced by IFN-gamma (MIG)/CXCL9 were significantly increased in CXCR2-/- mice compared with CXCR2+/+ mice at various times after conidia. Interestingly, at day 3 after conidia, neutrophil recruitment and airway hyperresponsiveness in CXCR2-/- mice was mediated by inflammatory protein-10/CXCL10 and, to a lesser degree, MIG/CXCL9. Taken together, these data suggest that CXCR2 contributes to the persistence of asthmatic disease due to A. fumigatus.     

Decreased expression of membrane IL-5 receptor alpha on human eosinophils: I. Loss of membrane IL-5 receptor alpha on airway eosinophils and increased soluble IL-5 receptor alpha in the airway after allergen challenge

IL-5 is a key cytokine for eosinophil maturation, recruitment, activation, and possibly the development of inflammation in asthma. High concentrations of IL-5 are present in the airway after Ag challenge, but the responsiveness of airway eosinophils to IL-5 is not well characterized. The objectives of this study were to establish, following airway Ag challenge: 1) the expression of membrane (m)IL-5Ralpha on bronchoalveolar lavage (BAL) eosinophils; 2) the responsiveness of these cells to exogenous IL-5; and 3) the presence of soluble (s)IL-5Ralpha in BAL fluid. To accomplish these goals, blood and BAL eosinophils were obtained from atopic subjects 48 h after segmental bronchoprovocation with Ag. There was a striking reduction in mIL-5Ralpha on airway eosinophils compared with circulating cells. Furthermore, sIL-5Ralpha concentrations were elevated in BAL fluid, but steady state levels of sIL-5Ralpha mRNA were not increased in BAL compared with blood eosinophils. Finally, BAL eosinophils were refractory to IL-5 for ex vivo degranulation, suggesting that the reduction in mIL-5Ralpha on BAL eosinophils may regulate IL-5-mediated eosinophil functions. Together, the loss of mIL-5Ralpha, the presence of sIL-5Ralpha, and the blunted functional response (degranulation) of eosinophils to IL-5 suggest that when eosinophils are recruited to the airway, regulation of their functions becomes IL-5 independent. These observations provide a potential explanation for the inability of anti-IL-5 therapy to suppress airway hyperresponsiveness to inhaled Ag, despite a reduction in eosinophil recruitment.     

Complex role of the IL-4 receptor alpha in a murine model of airway inflammation: expression of the IL-4 receptor alpha on nonlymphoid cells of bone marrow origin contributes to severity of inflammation

Recent studies have suggested the IL-4Ralpha expressed on lung epithelium is necessary for TH2-mediated goblet cell differentiation and mucus hypersecretion in a murine model of allergic lung disease. However, the IL-4Ralpha is expressed on numerous cell types that could contribute to the overall pathology and severity of asthma. The relative role of the receptor on these cells has not yet been conclusively delineated. To dissect the contribution of IL-4Ralpha in the development of pulmonary allergic responses, we generated murine radiation bone marrow (BM) chimeras. BM from IL-4Ralpha(+) or IL-4Ralpha(-) mice was transferred into recipient mice that expressed or lacked IL-4Ralpha. In the absence of IL-4Ralpha in recipient mice, there was no goblet cell metaplasia or mucus hypersecretion in response to OVA, even in the presence of TH2 cells and substantial eosinophilic infiltration. More importantly, we found that expression of the IL-4Ralpha on a nonlymphoid, MHC class II(+), BM-derived cell type contributes to the severity of inflammation and mucus production. These results suggest that IL-4 and IL-13 contribute to the development of allergic inflammation by stimulating a complex interaction between IL-4Ralpha(+) cell types of both bone marrow and non-bone marrow origin.     

CpG oligodeoxynucleotides can reverse Th2-associated allergic airway responses and alter the B7.1/B7.2 expression in a murine model of asthma

CpG oligodeoxynucleotides (CpG-ODN) administered during Ag sensitization or before Ag challenge can inhibit allergic pulmonary inflammation and airway hyperreactivity in murine models of asthma. In this study, we investigated whether CpG-ODN can reverse an ongoing allergic pulmonary reaction in a mouse model of asthma. AKR mice were sensitized with conalbumin followed by two intratracheal challenges at weekly intervals. CpG-ODN was administered 24 h after the first Ag challenge. CpG-ODN administration reduced Ag-specific IgE levels, bronchoalveolar lavage fluid eosinophils, mucus production, and airway hyperreactivity. We found that postchallenge CpG-ODN treatment significantly increased IFN-gamma concentrations and decreased IL-13, IL-4, and IL-5 concentrations in bronchoalveolar lavage fluids and spleen cell culture supernatants. Postchallenge CpG-ODN treatment also increased B7.1 mRNA expression and decreased B7.2 mRNA expression in lung tissues. These results suggest that CpG-ODN may have potential for treatment of allergic asthma by suppressing Th2 responses during IgE-dependent allergic airway reactions. The down-regulation of Th2 responses by CPG-ODN may be associated with regulation of the costimulatory factors B7.1 and B7.2.     

The role of IL-13 in established allergic airway disease

The effectiveness of targeting IL-13 in models where airway hyperresponsiveness (AHR) and airway inflammation have already been established is not well-described. We investigated the effects of blocking IL-13 on the early and late phase airway responses and the development of AHR in previously sensitized and challenged mice. BALB/cByJ mice were sensitized (days 1 and 14) and challenged (days 28-30) with OVA. Six weeks later (day 72), previously sensitized/challenged mice were challenged with a single OVA aerosol and the early and late phase response and development of AHR were determined. Specific in vivo blockade of IL-13 was attained after i.p. injection of a soluble IL-13Ralpha2-IgG fusion protein (sIL-13Ralpha2Fc) on days 71-72 for the early and late responses and on days 71-73 for the development of AHR. sIL-13Ralpha2Fc administration inhibited the late, but not early, phase response and the OVA challenge-induced changes in lung resistance and dynamic compliance; as well, sIL-13Ralpha2Fc administration decreased bronchoalveolar lavage eosinophilia and mucus hypersecretion following the secondary challenge protocols. These results demonstrate that targeting IL-13 alone regulates airway responses when administrated to mice with established allergic airway disease. These data identify the importance of IL-13 in the development of allergen-induced altered airway responsiveness following airway challenge, even when administered before rechallenge of mice in which allergic disease had been previously established.     

Immunomodulatory role of C10 chemokine in a murine model of allergic bronchopulmonary aspergillosis

The immunomodulatory role of the chemokine C10 was explored in allergic airway responses during experimental allergic bronchopulmonary aspergillosis (ABPA). The intratracheal delivery of Asperigillus fumigatus Ag into A. fumigatus-sensitized mice resulted in significantly increased levels of C10 within the bronchoalveolar lavage, and these levels peaked at 48 h after A. fumigatus challenge. In addition, C10 levels in BAL samples were greater than 5-fold higher than levels of other chemokines such as monocyte-chemoattractant protein-1, eotaxin, and macrophage-inflammatory protein-1alpha. From in vitro studies, it was evident that major pulmonary sources of C10 may have included alveolar macrophages, lung fibroblasts, and vascular smooth muscle cells. Experimental ABPA was associated with severe peribronchial eosinophilia, bronchial hyperresponsiveness, and augmented IL-13 and IgE levels. The immunoneutralization of C10 with polyclonal anti-C10 antiserum 2 h before the intratracheal A. fumigatus challenge significantly reduced the airway inflammation and hyperresponsiveness in this model of ABPA, but had no effect on IL-10 nor IgE levels. Taken together, these data suggest that C10 has a unique role in the progression of experimental ABPA.     

Polymorphisms in IL-4R alpha correlate with airways hyperreactivity, eosinophilia, and Ym protein expression in allergic IL-13-/- mice

The development of airways hyperreactivity in allergic IL-13(-/-) mice is controversial and appears to correlate with the number of times that the original 129 x C57BL/6 founder strain has been crossed to the BALB/c background. In this investigation, we compared allergic responses in founder IL-13(-/-) mice crossed for either 5 (N5) or 10 (N10) generations to BALB/c mice. Whereas allergic N5 IL-13(-/-) mice developed airways hyperreactivity, tissue eosinophilia, elevated IgE, and pulmonary expression of Ym proteins, these processes were attenuated in N5 IL-13(-/-) mice treated with an IL-4-neutralizing Ab, and in N10 IL-13(-/-) mice. These data showed that IL-4 was more effective in regulating allergic responses in N5 IL-13(-/-) mice than in N10 IL-13(-/-) mice. To elucidate the mechanism associated with these observations, we show by restriction and sequence analysis that N5 IL-13(-/-) mice express the C57BL/6 form of IL-4Ralpha and N10 IL-13(-/-) mice express the BALB/c form. Despite the near identical predicted molecular mass of these isoforms, IL-4Ralpha from N5 IL-13(-/-) mice migrates with a slower electrophoretic mobility than IL-4Ralpha from N10 IL-13(-/-) mice, suggesting more extensive posttranslational modification of the N5 form. The Thre(49)Ile polymorphism in the extracellular domain of BALB/c IL-4Ralpha has been demonstrated to disrupt N-linked glycosylation of Asn(47) and increase the dissociation rate of the IL-4Ralpha/IL-4 interaction. Collectively, these data show that polymorphisms in IL-4Ralpha, which have been shown to affect the interaction with IL-4, correlate with the ability of IL-4 to regulate allergic responses in IL-13(-/-) mice.     

Comparative roles of IL-4, IL-13, and IL-4Ralpha in dendritic cell maturation and CD4+ Th2 cell function

IL-4 and IL-13 play key roles in Th2 immunity and asthma pathogenesis. Although the function of these cytokines is partially linked through their shared use of IL-4Ralpha for signaling, the interplay between these cytokines in the development of memory Th2 responses is not well delineated. In this investigation, we show that both IL-4 and IL-13 influence the maturation of dendritic cells (DC) in the lung and their ability to regulate secretion of IFN-gamma and Th2 cytokines by memory CD4(+) T cells. Cocultures of wild-type T cells with pulmonary DC from allergic, cytokine-deficient mice demonstrated that IL-4 enhanced the capacity of DC to stimulate T cell secretion of Th2 cytokines, whereas IL-13 enhanced the capacity of DC to suppress T cell secretion of IFN-gamma. Because IL-4Ralpha is critical for IL-4 and IL-13 signaling, we also determined how variants of IL-4Ralpha influenced immune cell function. T cells derived from allergic mice expressing a high-affinity IL-4Ralpha variant produced higher levels of IL-5 and IL-13 compared with T cells derived from allergic mice expressing a low-affinity IL-4Ralpha variant. Although DC expressing different IL-4Ralpha variants did not differ in their capacity to influence Th2 cytokine production, they varied in their capacity to inhibit IFN-gamma production by T cells. Thus, IL-4 and IL-13 differentially regulate DC function and the way these cells regulate T cells. The affinity of IL-4Ralpha also appears to be a determinant in the balance between Th2 and IFN-gamma responses and thus the severity of allergic disease.     

Therapeutic attenuation of pulmonary fibrosis via targeting of IL-4- and IL-13-responsive cells

Severe forms of idiopathic interstitial pneumonia (IIP), such as usual interstitial pneumonia, can be impervious to modern steroid and immunosuppressive treatment regimens, thereby emphasizing the need for novel effective therapies. Consequently, research attention has been directed toward understanding the cytokine networks that may affect fibroblast activation and, hence, the progression of certain IIPs. This led us to investigate whether the specific targeting of resident lung cells responsive to IL-4 and IL-13 exerted a therapeutic effect in an experimental model of IIP, namely the bleomycin-induced model of pulmonary fibrosis. IL-4, IL-13, and their corresponding receptor subunits, IL-4Ralpha, IL-13Ralpha1, and IL-13Ralpha2, were maximally expressed at the mRNA and protein levels in whole lung samples on day 21 or 28 after an intratracheal bleomycin challenge. The intranasal administration of an IL-13 immunotoxin chimeric molecule (IL13-PE) from days 21-28, but not for 1-wk periods at earlier times, after bleomycin challenge had a significant therapeutic effect on histological and biochemical parameters of bleomycin-induced pulmonary fibrosis compared with the control group. The intranasal IL13-PE therapy significantly reduced the numbers of IL-4 and IL-13 receptor-positive mononuclear cells and macrophages and the levels of profibrotic cytokine and chemokine in the lungs of bleomycin-challenged mice on day 28. Thus, this study demonstrates that IL-4- and/or IL-13-binding cells are required for the maintenance of pulmonary fibrosis induced by bleomycin and highlights the importance of further investigation of antifibrotic therapeutics that target these cells during pulmonary fibrosis.     

白细胞介素-33抗体治疗呼吸道合胞病毒诱导小鼠哮喘的效果观察

为探讨白细胞介素(interleukin,IL)-33在呼吸道合胞病毒(respiratory syncytial virus,RSV)感染导致哮喘急性加重过程中的作用,选取3周龄雌性BALB/c小鼠28只,随机分为5组,分别为正常对照组、单纯RSV感染组、哮喘组、RSV感染哮喘并对照抗体组、RSV感染哮喘并IL-33...     

Repeated intratracheal challenge with particulate antigen modulates murine lung cytokines

When lungs of experimental animals are repeatedly challenged with Ag, pulmonary inflammation wanes via unknown mechanisms. We hypothesized that changes in the balance of lung cytokines are responsible for immune down-regulation to repeated Ag challenge. We used intratracheal (IT) challenge of primed C57BL/6 mice with SRBC and on various days after single (1IT) or triple (3IT) challenge counted lung inflammatory cells and measured whole-lung cytokine mRNA and protein concentrations using RT-PCR and ELISA. We found that lung lymphocyte numbers and parenchymal lung inflammation decreased significantly at days 6 and 9 after final Ag challenge in 3IT mice compared with 1IT mice. Lungs of 3IT mice showed the following changes in relative mRNA expression: an earlier peak in IL-10, decreased IL-1beta, and a change from a Th2 response in 1IT mice to a Th1 response in 3IT mice (with pronounced increases in IL-12, IL-18, and IFN-gamma and decreased IL-4, IL-13, and IL-5). Similar types of changes were seen in whole-lung protein concentrations for TNF-alpha, IL-10, IL-12 p40, IFN-gamma, and IL-4. Additionally, mRNA expression of the endothelial selectins CD62E and CD62P decreased and lung lymphocyte apoptosis increased in the 3IT group. Thus, physiologic down-regulation of the pulmonary immune response to repeated Ag exposure is characterized by increased anti- and decreased proinflammatory cytokines that accompanies Th1 polarization. Similar mechanisms may act to minimize chronic lung inflammation in the majority of normal humans who do not develop progressive lung pathology when repeatedly exposed to inhaled or aspirated environmental Ags.     

IL-13 induces airways hyperreactivity independently of the IL-4R alpha chain in the allergic lung

The potent spasmogenic properties of IL-13 have identified this molecule as a potential regulator of airways hyperreactivity (AHR) in asthma. Although IL-13 is thought to primarily signal through the IL-13Ralpha1-IL-4Ralpha complex, the cellular and molecular components employed by this cytokine to induce AHR in the allergic lung have not been identified. By transferring OVA-specific CD4(+) T cells that were wild type (IL-13(+/+) T cells) or deficient in IL-13 (IL-13(-/-) T cells) to nonsensitized mice that were then challenged with OVA aerosol, we show that T cell-derived IL-13 plays a key role in regulating AHR, mucus hypersecretion, eotaxin production, and eosinophilia in the allergic lung. Moreover, IL-13(+/+) T cells induce these features (except mucus production) of allergic disease independently of the IL-4Ralpha chain. By contrast, IL-13(+/+) T cells did not induce disease in STAT6-deficient mice. This shows that IL-13 employs a novel component of the IL-13 receptor signaling system that involves STAT6, independently of the IL-4Ralpha chain, to modulate pathogenesis. We show that this novel pathway for IL-13 signaling is dependent on T cell activation in the lung and is critically linked to downstream effector pathways regulated by eotaxin and STAT6.     

Long-lived Th2 memory in experimental allergic asthma

Although life-long immunity against pathogens is beneficial, immunological memory responses directed against allergens are potentially harmful. Because there is a paucity of information about Th2 memory cells in allergic disease, we established a model of allergic asthma in BALB/c mice to explore the generation and maintenance of Th2 memory. We induced disease without the use of adjuvants, thus avoiding Ag depots, and found that unlike allergic asthma in mice immunized with adjuvant, immunizing with soluble and aerosol OVA resulted in pathological lung lesions resembling human disease. To test memory responses we allowed mice with acute disease to recover and then re-exposed them to aerosol OVA a second time. Over 400 days later these mice developed OVA-dependent eosinophilic lung inflammation, airway hyperresponsiveness, mucus hypersecretion, and IgE. Over 1 year after recuperating from acute disease, mice had persistent lymphocytic lung infiltrates, Ag-specific production of IL-4 and IL-5 from spleen and lung cells in vitro, and elevated IgG1. Moreover, when recuperated mice were briefly aerosol challenged, we detected early expression of Th2 cytokine RNA in lungs. Taken together, these data demonstrate the presence of long-lived Th2 memory cells in spleen and lungs involved in the generation of allergic asthma upon Ag re-exposure.     

Cyclooxygenase inhibition augments allergic inflammation through CD4-dependent, STAT6-independent mechanisms

Nonselective cyclooxygenase (COX) inhibition during the development of allergic disease in a murine model causes an increase in type 2 cytokines and lung eosinophilia; however, the mechanisms responsible for this augmented allergen-induced inflammation have not been examined. Ab depletion of CD4 and CD8 cells revealed that the heightened allergic inflammation caused by COX inhibition was CD4, but not CD8, dependent. Allergen sensitization and airway challenge alone led to undetectable levels of IL-5 and IL-13 in the lungs of IL-4, IL-4Ralpha, and STAT6 knockout (KO) mice, but COX inhibition during the development of allergic inflammation resulted in wild-type levels of IL-5 and IL-13 and heightened airway eosinophilia in each of the three KO mice. These results indicate that the effect of COX inhibition was independent of signaling through IL-4, IL-4Ralpha, and STAT6. However, whereas COX inhibition increased IgE levels in allergic wild-type mice, IgE levels were undetectable in IL-4, IL-4Ralpha, and STAT6 KO mice, suggesting that IL-13 alone is not a switch factor for IgE synthesis in this model. These results illustrate the central role played by products derived from the COX pathway in the regulation of allergic immune responses.     

Inhalation of toluene diisocyanate vapor induces allergic rhinitis in mice

Diisocyanates are the leading cause of occupational asthma, and epidemiological evidence suggests that occupational rhinitis is a comorbid and preceding condition in patients who develop asthma. The goal of the present studies was to develop and characterize a murine model of toluene diisocyanate (TDI)-induced rhinitis. Female C57BL/6 mice were exposed to workplace-relevant concentrations of TDI vapor via inhalation for 4 h/day for 12 days with or without a 2-wk rest period and TDI challenge. Mice exposed 12 consecutive weekdays to 50 parts per billion TDI vapor showed elevated total serum IgE and increased TDI-specific IgG titers. Breathing rates were decreased corresponding with increased inspiratory time. TDI exposure elevated IL-4, IL-5, IL-13, and IFN-gamma mRNA expression in the nasal mucosa, suggesting a mixed Th1/Th2 immune response. Expressions of mRNA for proinflammatory cytokines and adhesion molecules were also up-regulated. These cytokine changes corresponded with a marked influx of inflammatory cells into the nasal mucosa, eosinophils being the predominant cell type. Removal from exposure for 2 wk resulted in reduced Ab production, cytokine mRNA expression, and cellular inflammation. Subsequent challenge with 50 parts per billion TDI vapor resulted in robust up-regulation of Ab production, cytokine gene expression, as well as eosinophilic inflammation in the nasal mucosa. There were no associated changes in the lung. The present model shows that TDI inhalation induces immune-mediated allergic rhinitis, displaying the major features observed in human disease. Future studies will use this model to define disease mechanisms and examine the temporal/dose relationship between TDI-induced rhinitis and asthma.     

Reconstitution of a functional human type II IL-4/IL-13 receptor in mouse B cells: demonstration of species specificity

IL-13 is a Th2-derived pleiotropic cytokine that recently was shown to be a key mediator of allergic asthma. IL-13 mediates its effects via a complex receptor system, which includes the IL-4R alpha-chain, IL-4Ralpha, and at least two other cell surface proteins, IL-13Ralpha1 and IL-13Ralpha2, which specifically bind IL-13. IL-13 has been reported to have very limited effects on mouse B cells. It was unclear whether this was due to a lack of receptor expression, a disproportionate relative expression of the receptor components, or an additional subunit requirement in B cells. To determine the requirements for IL-13 signaling in murine B cells, we examined IL-13-dependent Stat6 activation and CD23 induction in the murine B cell line, A201.1. A201.1 cells responded to murine IL-4 via the type I IL-4R, but were unresponsive to IL-13, and did not express IL-13 receptor. B220(+) splenocytes also failed to signal in response to IL-13 and did not express IL-13 receptor. We transfected A201.1 cells with human IL-4Ralpha, IL-13Ralpha1, or both. Transfectants expressing either human IL-4Ralpha or human IL-13Ralpha1 alone were unable to respond or signal to IL-13. Thus, human IL-13Ralpha1 could not combine with the endogenous murine IL-4Ralpha to generate a functional IL-13R. However, cells transfected with both human IL-4Ralpha and IL-13Ralpha1 responded to IL-13. Thus, the relative lack of IL-13 responsiveness in murine B cells is due to a lack of receptor expression. Furthermore, the heterodimeric interaction between IL-4Ralpha and IL-13Ralpha1 is species specific.