FcγRIIb Inhibits Allergic Lung Inflammation in a Murine Model of Allergic Asthma

Allergic asthma is characterized by airway eosinophilia, increased mucin production and allergen-specific IgE. Fc gamma receptor IIb (FcγRIIb), an inhibitory IgG receptor, has recently emerged as a negative regulator of allergic diseases like anaphylaxis and allergic rhinitis. However, no studies to date have evaluated its role in allergic asthma. Our main objective was to study the role of FcγRIIb in allergic lung inflammation. We used a murine model of allergic airway inflammation. Inflammation was quantified by BAL inflammatory cells and airway mucin production. FcγRIIb expression was measured by qPCR and flow cytometry and the cytokines were quantified by ELISA. Compared to wild type animals, FcγRIIb deficient mice mount a vigorous allergic lung inflammation characterized by increased bronchoalveolar lavage fluid cellularity, eosinophilia and mucin content upon ragweed extract (RWE) challenge. RWE challenge in sensitized mice upregulated FcγRIIb in the lungs. Disruption of IFN-γ gene abrogated this upregulation. Treatment of naïve mice with the Th1-inducing agent CpG DNA increased FcγRIIb expression in the lungs. Furthermore, treatment of sensitized mice with CpG DNA prior to RWE challenge induced greater upregulation of FcγRIIb than RWE challenge alone. These observations indicated that RWE challenge upregulated FcγRIIb in the lungs by IFN-γ- and Th1-dependent mechanisms. RWE challenge upregulated FcγRIIb on pulmonary CD14+/MHC II+ mononuclear cells and CD11c+ cells. FcγRIIb deficient mice also exhibited an exaggerated RWE-specific IgE response upon sensitization when compared to wild type mice. We propose that FcγRIIb physiologically regulates allergic airway inflammation by two mechanisms: 1) allergen challenge mediates upregulation of FcγRIIb on pulmonary CD14+/MHC II+ mononuclear cells and CD11c+ cells by an IFN-γ dependent mechanism; and 2) by attenuating the allergen specific IgE response during sensitization. Thus, stimulating FcγRIIb may be a therapeutic strategy in allergic airway disorders.     

Multiplexed Digital mRNA Profiling of the Inflammatory Response in the West Nile Swiss Webster Mouse Model

Background and purpose

The ability to track changes in gene expression following viral infection is paramount to understanding viral pathogenesis. This study was undertaken to evaluate the nCounter, a high throughput digital gene expression system, as a means to better understand West Nile virus (WNV) dissemination and the inflammatory response against WNV in the outbred Swiss Webster (SW) mouse model over the course of infection.

Methodology

The nCounter Mouse Inflammation gene expression kit containing 179 inflammation related genes was used to analyze gene expression changes in multiple tissues over a nine day course of infection in SW mice following intraperitoneal injection with WNV. Protein expression levels for a subset of these cytokine/chemokine genes were determined using a multiplex protein detection system (BioPlex) and comparisons of protein/RNA expression levels made.

Results

Expression analysis of spleen, lung, liver, kidney and brain of SW mice infected with WNV revealed that Cxcl10 and Il12b are differentially expressed in all tissues tested except kidney. Data stratification of positively confirmed infected (WNV (+)) versus non-infected (WNV (−) tissues allowed differentiation of the systemic inflammatory gene response from tissue-specific responses arising from WNV infection. Significant (p<0.05) decrease in C3ar1 was found in WNV (−) spleen. Il23a was significantly upregulated, while Il10rb was down-regulated in WNV (−) lung. Il3 and Mbl2 were down-regulated in WNV (−) liver. In WNV (+) livers, Stat1, Tlr2, chemokines Cxcl1, Cxcl3, Cxcl9, Cxcl10, cytokines Il6, Il18, cytokine-related gene Il1r and cytokine agonist Ilrn were significantly upregulated. In WNV (−) brain tissues, Csf2 and Cxcl10 were significantly upregulated. Similar gene and protein expression kinetics were found for Ccl2, Ccl3, Ccl4 and Ccl5 and correlated with the presence of infectious virus. In summary, the utility of the nCounter platform for rapid identification of gene expression changes in SW mice associated with WNV infection was demonstrated.     

CC chemokine ligand 1 promotes recruitment of eosinophils but not Th2 cells during the development of allergic airways disease

One of the characteristic features of allergic asthma is recruitment of large numbers of inflammatory cells including eosinophils and Th2 lymphocytes to the lung. This influx of inflammatory cells is thought to be a controlled and coordinated process mediated by chemokines and their receptors. It is thought that distinct, differential expression of chemokine receptors allows selective migration of T cell subtypes in response to the chemokines that bind these receptors. Th2 cells preferentially express CCR8 and migrate selectively to its ligand, CC chemokine ligand (CCL)1. We studied the role of the CCR8 ligand, CCL1, in the specific recruitment of Th2 cells and eosinophils to the lung in a murine model of allergic airway disease. We have demonstrated for the first time that CCL1 is up-regulated in the lung following allergen challenge. Moreover, a neutralizing Ab to CCL1 reduced eosinophil migration to the lung, but had no effect on recruitment of Th2 cells following allergen challenge. In addition, there was no change in airway hyperresponsiveness or levels of Th2 cytokines. In a Th2 cell transfer system of pulmonary inflammation, anti-CCL1 also failed to affect recruitment of Th2 cells to the lung following allergen challenge. Significantly, intratracheal instillation of rCCL1 increased recruitment of eosinophils but not Th2 cells to the lung in allergen-sensitized and -challenged mice. In summary, our results indicate that CCL1 is important for the pulmonary recruitment of eosinophils, rather than allergen-specific Th2 cells, following allergen challenge.     

CCR8 is not essential for the development of inflammation in a mouse model of allergic airway disease

Chemokine receptors play an important role in the trafficking of various immune cell types to sites of inflammation. Several chemokine receptors are differentially expressed in Th1 and Th2 effector populations. Th2 cells selectively express CCR3, CCR4, and CCR8, which could direct their trafficking to sites of allergic inflammation. Additionally, increased expression of the CCR8 ligand, TCA-3, has been detected in affected lungs in a mouse model of asthma. In this study, CCR8-deficient mice were generated to address the biological role of CCR8 in a model of allergic airway disease. Using two different protocols of allergen challenge, we demonstrate that absence of CCR8 does not affect the development of pulmonary eosinophilia and Th2 cytokine responses. In addition, administration of anti-TCA-3-neutralizing Ab during allergen sensitization and rechallenge failed to inhibit airway allergic inflammation. These results suggest that CCR8 does not play an essential role in the pathogenesis of inflammation in this mouse model of allergic airway disease.     

Administration of pentoxifylline during allergen sensitization dissociates pulmonary allergic inflammation from airway hyperresponsiveness

Asthma, a chronic inflammatory disease characterized by intermittent, reversible airflow obstruction and airway hyperresponsiveness (AHR), is classically characterized by an excess of Th2 cytokines (IL-13, IL-4) and depletion of Th1 cytokines (IFN-gamma, IL-12). Recent studies indicating an important role for Th1 immunity in the development of AHR with allergic inflammation suggest that Th1/Th2 balance may be important in determining the association of AHR with allergic inflammation. We hypothesized that administration of pentoxifylline (PTX), a phosphodiesterase inhibitor known to inhibit Th1 cytokine production, during allergen (OVA) sensitization and challenge would lead to attenuation of AHR in a murine model of allergic pulmonary inflammation. We found that PTX treatment led to attenuation of AHR when administered at the time of allergen sensitization without affecting other hallmarks of pulmonary allergic inflammation. Attenuation of AHR with PTX treatment was found in the presence of elevated bronchoalveolar lavage fluid levels of the Th2 cytokine IL-13 and decreased levels of the Th1 cytokine IFN-gamma. PTX treatment during allergen sensitization leads to a divergence of AHR and pulmonary inflammation following allergen challenge.     

Regulatory T cells more effectively suppress Th1-induced airway inflammation compared with Th2

Asthma is a syndrome with different inflammatory phenotypes. Animal models have shown that, after sensitization and allergen challenge, Th2 and Th1 cells contribute to the development of allergic airway disease. We have previously demonstrated that naturally occurring regulatory T cells (nTregs) can only marginally suppress Th2-induced airway inflammation and airway hyperresponsiveness. In this study, we investigated nTreg-mediated suppression of Th2-induced and Th1-induced acute allergic airway disease. We demonstrate in vivo that nTregs exert their suppressive potency via cAMP transfer on Th2- and Th1-induced airway disease. A comparison of both phenotypes revealed that, despite similar cAMP transfers, Th1-driven airway hyperresponsiveness and inflammation are more susceptible to nTreg-dependent suppression, suggesting that potential nTreg-based therapeutic strategies might be more effective in patients with predominantly neutrophilic airway inflammation based on deregulated Th1 response.     

An essential role for DNA methyltransferase 3a in melanoma tumorigenesis

Abnormal DNA methylation and associated silencing of tumor suppressor genes are common to many types of cancers. Among the three coordinate DNA methyltransferases (Dnmts), Dnmt1 and Dnmt3b were both shown to be important for cancer cell survival and tumorigenesis. However, the relationship between Dnmt3a and tumorigenesis is still largely unknown. Here, we show that inhibition of Dnmt3a expression, by stable transfection of a Dnmt3a-RNA interference (RNAi) construct dramatically inhibited melanoma growth and metastasis in mouse melanoma models. Microarray analysis revealed that genes critical for the tumor immune response, were implicated in the inhibition of melanoma growth. Expression of a cluster of class I and class II MHC genes, class II transactivator (Ciita), as well as a subset of 5 chemokines (Cxcl9, Cxcl16, Ccl12, Ccl4, and Ccl2) were up-regulated. Furthermore, we determined that the promoter IV of Ciita was significantly demethylated in Dnmt3a-depleted tumors. In addition, several known tumor-related genes, which are critical for developmental processes and cell cycle, were confirmed to be misregulated, including TgfB1, Socs1, Socs2, E2F6, Ccne1, and Cyr61. The results presented in this report strongly suggest that Dnmt3a plays an essential role in melanoma tumorigenesis, and that the underlying mechanisms include the modulation of the tumor immune response, as well as other processes.     

Adoptive transfer of induced-Treg cells effectively attenuates murine airway allergic inflammation

Both nature and induced regulatory T (Treg) lymphocytes are potent regulators of autoimmune and allergic disorders. Defects in endogenous Treg cells have been reported in patients with allergic asthma, suggesting that disrupted Treg cell-mediated immunological regulation may play an important role in airway allergic inflammation. In order to determine whether adoptive transfer of induced Treg cells generated in vitro can be used as an effective therapeutic approach to suppress airway allergic inflammation, exogenously induced Treg cells were infused into ovalbumin-sensitized mice prior to or during intranasal ovalbumin challenge. The results showed that adoptive transfer of induced Treg cells prior to allergen challenge markedly reduced airway hyperresponsiveness, eosinophil recruitment, mucus hyper-production, airway remodeling, and IgE levels. This effect was associated with increase of Treg cells (CD4(+)FoxP3(+)) and decrease of dendritic cells in the draining lymph nodes, and with reduction of Th1, Th2, and Th17 cell response as compared to the controls. Moreover, adoptive transfer of induced Treg cells during allergen challenge also effectively attenuate airway inflammation and improve airway function, which are comparable to those by natural Treg cell infusion. Therefore, adoptive transfer of in vitro induced Treg cells may be a promising therapeutic approach to prevent and treat severe asthma.     

Regulation of cockroach antigen-induced allergic airway hyperreactivity by the CXCR3 ligand CXCL9

Allergic airway disease is characterized by a robust lymphocytic infiltrate, elaboration of Th2-type inflammatory mediators, pulmonary eosinophil accumulation, and airway hyperreactivity. The CXCR3 ligands, CXCL9 (monokine induced by IFN-gamma) and CXCL10 (IFN-inducible protein, 10 kDa), are IFN-gamma-inducible, Th1-type chemokines. As CXCL10 has been previously shown to participate in the modulation of allergic inflammation, we were interested in investigating the possible role that CXCL9 may play in this inflammatory response. Expression of CXCL9 was primarily identified in airway epithelial cells by immunohistochemical staining. Airway neutralization of CXCL9 at the time of allergen challenge significantly increased airway hyperreactivity, airway eosinophil accumulation, and IL-4 levels in the bronchoalveolar lavage while significantly decreasing airway levels of IL-12. In contrast, introduction of exogenous CXCL9 into the airway at the time of allergen challenge dramatically reduced airway hyper-reactivity and eosinophil accumulation. Moreover, pulmonary levels of IL-4 were significantly reduced, whereas levels of IL-12 were significantly increased, with exogenous CXCL9 treatment. In lymphocytes restimulated with CXCL9 and allergen in vitro, CXCL9 down-regulated IL-4 expression and up-regulated IFN-gamma expression, suggesting that CXCL9 is able to direct activated lymphocytes toward a Th1-type phenotype. Additionally, CXCL9 was shown to inhibit CC chemokine ligand 11-induced eosinophil chemotaxis in in vitro assays. Taken together, our results demonstrate that the CXCR3 ligand CXCL9 is involved in regulation of the allergic response in the lung by regulation of lymphocyte activation and eosinophil recruitment.     

Early-life viral infection and allergen exposure interact to induce an asthmatic phenotype in mice

Background

Early-life respiratory viral infections, notably with respiratory syncytial virus (RSV), increase the risk of subsequent development of childhood asthma. The purpose of this study was to assess whether early-life infection with a species-specific model of RSV and subsequent allergen exposure predisposed to the development of features of asthma.

Methods

We employed a unique combination of animal models in which BALB/c mice were neonatally infected with pneumonia virus of mice (PVM, which replicates severe RSV disease in human infants) and following recovery, were intranasally sensitised with ovalbumin. Animals received low-level challenge with aerosolised antigen for 4 weeks to elicit changes of chronic asthma, followed by a single moderate-level challenge to induce an exacerbation of inflammation. We then assessed airway inflammation, epithelial changes characteristic of remodelling, airway hyperresponsiveness (AHR) and host immunological responses.

Results

Allergic airway inflammation, including recruitment of eosinophils, was prominent only in animals that had recovered from neonatal infection with PVM and then been sensitised and chronically challenged with antigen. Furthermore, only these mice exhibited an augmented Th2-biased immune response, including elevated serum levels of anti-ovalbumin IgE and IgG₁ as well as increased relative expression of Th2-associated cytokines IL-4, IL-5 and IL-13. By comparison, development of AHR and mucous cell change were associated with recovery from PVM infection, regardless of subsequent allergen challenge. Increased expression of IL-25, which could contribute to induction of a Th2 response, was demonstrable in the lung following PVM infection. Signalling via the IL-4 receptor α chain was crucial to the development of allergic inflammation, mucous cell change and AHR, because all of these were absent in receptor-deficient mice. In contrast, changes of remodelling were evident in mice that received chronic allergen challenge, regardless of neonatal PVM infection, and were not dependent on signalling via the IL-4 receptor.

Conclusion

In this mouse model, interaction between early-life viral infection and allergen sensitisation/challenge is essential for development of the characteristic features of childhood asthma, including allergic inflammation and a Th2-biased immune response.     

Airway inflammation and IgE production induced by dust mite allergen-specific memory/effector Th2 cell line can be effectively attenuated by IL-35

CD4(+) memory/effector T cells play a central role in orchestrating the rapid and robust immune responses upon re-encounter with specific Ags. However, the immunologic mechanism(s) underlying these responses are still not fully understood. To investigate this, we generated an allergen (major house dust mite allergen, Blo t 5)-specific murine Th2 cell line that secreted IL-4, IL-5, IL-10, and IL-13, but not IL-9 or TNF-α, upon activation by the cognate Ag. These cells also exhibited CD44(high)CD62L(-) and CD127(+) (IL-7Rα(+)) phenotypes, which are characteristics of memory/effector T cells. Experiments involving adoptive transfer of this Th2 cell line in mice, followed by three intranasal challenges with Blo t 5, induced a dexamethasone-sensitive eosinophilic airway inflammation. This was accompanied by elevation of Th2 cytokines and CC- and CXC-motif chemokines, as well as recruitment of lymphocytes and polymorphic mononuclear cells into the lungs. Moreover, Blo t 5-specific IgE was detected 4 d after the last intranasal challenge, whereas elevation of Blo t 5-specific IgG1 was found at week two. Finally, pulmonary delivery of the pVAX-IL-35 DNA construct effectively downregulated Blo t 5-specific allergic airway inflammation, and i.m. injection of pVAX-IL-35 led to long-lasting suppression of circulating Blo t 5-specific and total IgE. This model provides a robust research tool to elucidate the immunopathogenic role of memory/effector Th2 cells in allergic airway inflammation. Our results suggested that IL-35 could be a potential therapeutic target for allergic asthma through its attenuating effects on allergen-specific CD4(+) memory/effector Th2 cell-mediated airway inflammation.     

Potential role of senescent macrophages in radiation-induced pulmonary fibrosis

Radiation-induced pulmonary fibrosis (RIPF) is a late toxicity of therapeutic radiation in clinic with poor prognosis and limited therapeutic options. Previous results have shown that senescent cells, such as fibroblast and type II airway epithelial cell, are strongly implicated in pathology of RIPF. However, the role of senescent macrophages in the development RIPF is still unknown. In this study, we report that ionizing radiation (IR) increase cellular senescence with higher expression of senescence-associated β-galactosidase (SA-β-Gal) and senescence-specific genes (p16, p21, Bcl-2, and Bcl-xl) in irradiated bone marrow-derived monocytes/macrophages (BMMs). Besides, there’s a significant increase in the expression of pro-fibrogenic factors (TGF-β1 and Arg-1), senescence-associated secretory phenotype (SASP) proinflammatory factors (Il-1α, Il-6, and Tnf-α), SASP chemokines (Ccl2, Cxcl10, and Ccl17), and SASP matrix metalloproteinases (Mmp2, Mmp9 and Mmp12) in BMMs exposed to 10 Gy IR. In addition, the percentages of SA-β-Gal⁺ senescent macrophages are significantly increased in the macrophages of murine irradiated lung tissue. Moreover, robustly elevated expression of p16, SASP chemokines (Ccl2, Cxcl10, and Ccl17) and SASP matrix metalloproteinases (Mmp2, Mmp9, and Mmp12) is observed in the macrophages of irradiated lung, which might stimulate a fibrotic phenotype in pulmonary fibroblasts. In summary, irradiation can induce macrophage senescence, and increase the secretion of SASP in senescent macrophages. Our findings provide important evidence that senescent macrophages might be the target for prevention and treatment of RIPF.Subject terms: Senescence, Chemokines     

IL-12 contributes to allergen-induced airway inflammation in experimental asthma

Lack of sufficient IL-12 production has been suggested to be one of the basic underlying mechanisms in atopy, but a potential role of IL-12 in established allergic airway disease remains unclear. We took advantage of a mouse model of experimental asthma to study the role of IL-12 during the development of bronchial inflammation. Administration of anti-IL-12p35 or anti-IL-12p40 mAb to previously OVA-sensitized BALB/c mice concomitantly with exposure to nebulized OVA, abolished both the development of bronchial hyperresponsiveness to metacholine as well as the eosinophilia in bronchoalveolar lavage fluid and peripheral blood. Anti-IL-12 treatment reduced CD4(+) T cell numbers and IL-4, IL-5, and IL-13 levels in the bronchoalveolar lavage fluid and the mRNA expression of IL-10, eotaxin, RANTES, MCP-1, and VCAM-1 in the lung. Anti-IL-12p35 treatment failed to show these effects in IFN-gamma knockout mice pointing to the essential role of IFN-gamma in IL-12-induced effects. Neutralization of IL-12 during the sensitization process aggravated the subsequent development of allergic airway inflammation. These data together with recent information on the role of dendritic cells in both the sensitization and effector phase of allergic respiratory diseases demonstrate a dual role of IL-12. Whereas IL-12 counteracts Th2 sensitization, it contributes to full-blown allergic airway disease upon airway allergen exposure in the postsensitization phase, with enhanced recruitment of CD4(+) T cells and eosinophils and with up-regulation of Th2 cytokines, chemokines, and VCAM-1. IFN-gamma-producing cells or cells dependent on IFN-gamma activity, play a major role in this unexpected proinflammatory effect of IL-12 in allergic airway disease.     

Establishment of tolerance to commensal bacteria requires a complex microbiota and is accompanied by decreased intestinal chemokine expression

Intricate regulation of tolerance to the intestinal commensal microbiota acquired at birth is critical. We hypothesized that epithelial cell tolerance toward early gram-positive and gram-negative colonizing bacteria is established immediately after birth, as has previously been shown for endotoxin. Gene expression in the intestine of mouse pups born to dams that were either colonized with a conventional microbiota or monocolonized (Lactobacillus acidophilus or Eschericia coli) or germ free was examined on day 1 and day 6 after birth. Intestinal epithelial cells from all groups of pups were stimulated ex vivo with L. acidophilus and E. coli to assess tolerance establishment. Intestine from pups exposed to a conventional microbiota displayed lower expression of Ccl2, Ccl3, Cxcl1, Cxcl2, and Tslp than germ-free mice, whereas genes encoding proteins in Toll-like receptor signaling pathways and cytokines were upregulated. When comparing pups on day 1 and day 6 after birth, a specific change in gene expression pattern was evident in all groups of mice. Tolerance to ex vivo stimulation with E. coli was only established in conventional animals. Colonization of the intestine was reflected in the spleen displaying downregulation of Cxcl2 compared with germ-free animals on day 1 after birth. Colonization reduced the expression of genes involved in antigen presentation in the intestine-draining mesenteric lymph nodes, but not in the popliteal lymph nodes, as evidenced by gene expression on day 23 after birth. We propose that microbial detection systems in the intestine are upregulated by colonization with a diverse microbiota, whereas expression of proinflammatory chemokines is reduced to avoid excess recruitment of immune cells to the maturing intestine.     

Dendritic cell co-transfected with FasL and allergen genes induces T cell tolerance and decreases airway inflammation in allergen induced murine model

Dendritic cell (DC) displays tremendous functional plasticity in response to antigens and plays important roles in inducing immune tolerance. In this study, we investigated the effects of immature DC (imDC) co-transfected with FasL and allergen Der p2 genes (FasL-Der p2-DC) on inducing immune tolerance and modulating airway inflammation of Der p2-induced allergic mice. Moreover, we compared the effects of FasL-Der p2-DC with FasL-transfected imDC (FasL-DC) and Der p2-transfected imDC (Der p2-DC) respectively. Results showed that FasL-Der p2-DC and Der p2-pulsed FasL-DC induced T cell unresponsiveness to Der p2 via apoptosis. Der p2-DC could induce T cell hyporesponsiveness to Der p2. FasL-Der p2-DC, FasL-DC and Der p2-DC could inhibit Th2 response and reduce allergic airway inflammation. FasL-Der p2-DC was more effective than FasL-DC and Der p2-DC, respectively. These results demonstrate that FasL and allergen genes co-expressing DC might be a promising approach to allergy therapy.     

Local nitric oxide levels reflect the degree of allergic airway inflammation after segmental allergen challenge in asthmatics.

Nitric oxide (NO) levels are increased in the exhaled air of asthmatics. As NO levels correlate with allergic airway inflammation, NO measurement has been suggested for disease monitoring. In patients with asthma, we previously demonstrated that intrabronchial treatment with a natural porcine surfactant enhanced airway inflammation after segmental allergen provocation. We studied whether local levels of NO reflect the degree of allergic airway inflammation following segmental allergen challenge with or without surfactant pretreatment. Segmental NO, as well as nitrite and nitrate in bronchoalveolar lavage (BAL) fluid, was measured before and after segmental challenge with either saline, saline plus allergen, or surfactant plus allergen in 16 patients with asthma and five healthy subjects. The data were compared with inflammatory BAL cells. Segmental NO levels were increased after instillation of saline (p < 0.05), or surfactant plus allergen in asthmatics (p < 0.05), and values were higher after surfactant plus allergen compared to saline challenge. Nitrate BAL levels were not altered after saline challenge but increased after allergen challenge (p < 0.05) and further raised by surfactant (p < 0.05), whereas nitrite levels were not altered by any treatment. Segmental NO and nitrate levels correlated with the degree of eosinophilic airway inflammation, and nitrate levels also correlated with neutrophil and lymphocyte numbers in BAL. In healthy subjects, NO, nitrite, and nitrate were unaffected. Thus, segmental NO and nitrate levels reflect the degree of allergic airway inflammation in patients with asthma. Measurement of both markers can be useful in studies using segmental allergen provocation, to assess local effects of potential immunomodulators.     

IL-10- and IL-12-independent down-regulation of allergic sensitization by stimulation of CD40 signaling

Interaction between CD154 (CD40 ligand) on activated T lymphocytes and its receptor CD40 has been shown to be critically involved in the generation of cell-mediated as well as humoral immunity. CD40 triggering activates dendritic cells (DC), enhances their cytokine production, up-regulates the expression of costimulatory molecules, and induces their maturation. It is unknown how stimulation of CD40 during sensitization to an airborne allergen may affect the outcome of allergic airway inflammation. We took advantage of a mouse model of allergic asthma and a stimulatory mAb to CD40 (FGK45) to study the effects of CD40-mediated DC activation on sensitization to OVA and subsequent development of OVA-induced airway inflammation. Agonistic anti-CD40 mAb (FGK45) injected during sensitization with OVA abrogated the development of allergic airway inflammation upon repeated airway challenges with OVA. Inhibition of bronchial eosinophilia corresponded with reduced Th2 cytokine production and was independent of IL-12, as evidenced by a similar down-regulatory effect of anti-CD40 mAb in IL-12 p40-deficient mice. In addition, FGK45 equally down-regulated allergic airway inflammation in IL-10-deficient mice, indicating an IL-10-independent mechanism of action of FGK45. In conclusion, our results show that CD40 signaling during sensitization shifts the immune response away from Th2 cytokine production and suppresses allergic airway inflammation in an IL-12- and IL-10-independent way, presumably resulting from enhanced DC activation during sensitization.     

Inhibition of Aldose Reductase Prevents Experimental Allergic Airway Inflammation in Mice

Background

The bronchial asthma, a clinical complication of persistent inflammation of the airway and subsequent airway hyper-responsiveness, is a leading cause of morbidity and mortality in critically ill patients. Several studies have shown that oxidative stress plays a key role in initiation as well as amplification of inflammation in airways. However, still there are no good anti-oxidant strategies available for therapeutic intervention in asthma pathogenesis. Most recent studies suggest that polyol pathway enzyme, aldose reductase (AR), contributes to the pathogenesis of oxidative stress–induced inflammation by affecting the NF-κB-dependent expression of cytokines and chemokines and therefore inhibitors of AR could be anti-inflammatory. Since inhibitors of AR have already gone through phase-III clinical studies for diabetic complications and found to be safe, our hypothesis is that AR inhibitors could be novel therapeutic drugs for the prevention and treatment of asthma. Hence, we investigated the efficacy of AR inhibition in the prevention of allergic responses to a common natural airborne allergen, ragweed pollen that leads to airway inflammation and hyper-responsiveness in a murine model of asthma.

Methods and Findings

Primary Human Small Airway Epithelial Cells (SAEC) were used to investigate the in vitro effects of AR inhibition on ragweed pollen extract (RWE)-induced cytotoxic and inflammatory signals. Our results indicate that inhibition of AR prevents RWE -induced apoptotic cell death as measured by annexin-v staining, increase in the activation of NF-κB and expression of inflammatory markers such as inducible nitric oxide synthase (iNOS), cycloxygenase (COX)-2, Prostaglandin (PG) E₂, IL-6 and IL-8. Further, BALB/c mice were sensitized with endotoxin-free RWE in the absence and presence of AR inhibitor and followed by evaluation of perivascular and peribronchial inflammation, mucin production, eosinophils infiltration and airway hyperresponsiveness. Our results indicate that inhibition of AR prevents airway inflammation and production of inflammatory cytokines, accumulation of eosinophils in airways and sub-epithelial regions, mucin production in the bronchoalveolar lavage fluid and airway hyperresponsiveness in mice.

Conclusions

These results suggest that airway inflammation due to allergic response to RWE, which subsequently activates oxidative stress-induced expression of inflammatory cytokines via NF-κB-dependent mechanism, could be prevented by AR inhibitors. Therefore, inhibition of AR could have clinical implications, especially for the treatment of airway inflammation, a major cause of asthma pathogenesis.