Natural killer T cells are dispensable in the development of allergen-induced airway hyperresponsiveness, inflammation and remodelling in a mouse model of chronic asthma

Natural killer T (NK T) cells have been shown to play an essential role in the development of allergen-induced airway hyperresponsiveness (AHR) and/or airway inflammation in mouse models of acute asthma. Recently, NK T cells have been reported to be required for the development of AHR in a virus induced chronic asthma model. We investigated whether NK T cells were required for the development of allergen-induced AHR, airway inflammation and airway remodelling in a mouse model of chronic asthma. CD1d^−/− mice that lack NK T cells were used for the experiments. In the chronic model, AHR, eosinophilic inflammation, remodelling characteristics including mucus metaplasia, subepithelial fibrosis and increased mass of the airway smooth muscle, T helper type 2 (Th2) immune response and immunoglobulin (Ig)E production were equally increased in both CD1d^−/− mice and wild-type mice. However, in the acute model, AHR, eosinophilic inflammation, Th2 immune response and IgE production were significantly decreased in the CD1d^−/− mice compared to wild-type. CD1d-dependent NK T cells may not be required for the development of allergen-induced AHR, eosinophilic airway inflammation and airway remodelling in chronic asthma model, although they play a role in the development of AHR and eosinophilic inflammation in acute asthma model.     

Allergen-induced bronchial inflammation is associated with decreased levels of surfactant proteins A and D in a murine model of asthma

Background Increasing evidence suggests that pulmonary surfactant protein A (SP‐A) and D (SP‐D) participate in the lung defence against pathogens. However, the role of surfactant proteins in the pathogenesis of allergen‐induced airway inflammation has not been elucidated. In this study we examined the levels and distributions of SP‐A and SP‐D in a dust mite (Dermatophagoides pteronyssinus, Der p) allergen‐induced murine model of asthma. Methods The concentration of SP‐A and SP‐D in the bronchoalveolar lavage fluid (BALF) and the distribution of surfactant proteins in the lung were assayed by ELISA and immunohistochemistry methods, respectively. The effect of surfactant proteins on allergen‐induced pulmonary lymphocyte proliferation was also studied. Results We demonstrated that there were marked reductions of SP‐A and SP‐D levels in the BALF of Der p‐sensitized BALB/c mice at 48–72 h after allergen challenge (AC). Both purified SP‐A and SP‐D were able to suppress, in a dose dependent manner, Der p‐stimulated intrapulmonary lymphocyte proliferation of naïve mice with saline or allergen challenge, or of Der p‐sensitized mice with saline challenge. On the contrary, this suppressive effect was mild (< 9%) on lymphocytes from sensitized mice after AC. Conclusion These results indicated the involvement of pulmonary surfactant proteins in the allergic bronchial inflammation of sensitized mice.     

Interleukin-18-deficient mice exhibit diminished chronic inflammation and airway remodelling in ovalbumin-induced asthma model

Interleukin (IL)-18, which is produced by activated monocytes/macrophages and airway epithelial cells, is suggested to contribute to the pathophysiology of asthma by modulating airway inflammation. However, the involvement of IL-18 on modulating chronic airway inflammation and airway remodelling, which are characterized in a refractory asthma model exposed to long-term antigen, has not been investigated sufficiently. We examined the role of IL-18 in chronic airway inflammation and airway remodelling by long-term antigen exposure. IL-18-deficient and C57BL/6-wild-type mice were sensitized by ovalbumin (OVA) and were then exposed to aerosolized OVA twice a week for 12 weeks. We assessed airway inflammation by assessing the infiltration of cells into the airspace and lung tissues, and airway remodelling by airway mucus expression, peribronchial fibrosis and smooth muscle thickness. In IL-18-deficient mice, when exposed to OVA, the total cells and neutrophils of the bronchoalveolar lavage fluid (BALF) were diminished, as were the number of infiltrated cells in the lung tissues. IL-18-deficient mice exposed to OVA after 12 weeks showed significantly decreased levels of interferon (IFN)-gamma, IL-13 and transforming growth factor (TGF)-beta1 in the BALF. The airway hyperresponsiveness to acetyl-beta-methacholine chloride was inhibited in IL-18-deficient mice in comparison with wild-type mice. In addition, IL-18-deficient mice exposed to OVA had fewer significant features of airway remodelling. These findings suggest that IL-18 may enhance chronic airway inflammation and airway remodelling through the production of IFN-gamma, IL-13 and TGF-beta1 in the OVA-induced asthma mouse model.     

Progesterone increases airway eosinophilia and hyper-responsiveness in a murine model of allergic asthma

BACKGROUND: Sex hormones might affect the severity and evolution of bronchial asthma. From existing literature, there exists, however, no convincing evidence for either exacerbation or improvement of allergic symptoms by progesterone. OBJECTIVE: This study was aimed to explore the effect of exogenously administered progesterone in a mouse model of allergic asthma. METHODS: BALB/c mice were sensitized to ovalbumin (OVA) by intraperitoneal injections with OVA followed by chronic inhalation of nebulized OVA or physiologic saline (Sal). Medroxyprogesterone acetate or placebo was instilled daily into the oesophagus before and during the inhalatory OVA challenge phase. RESULTS: Progesterone worsened allergic airway inflammation in OVA-challenged mice, as evidenced by enhanced bronchial responsiveness to inhaled metacholine and increased bronchial eosinophilia. Elevated airway eosinophilia corresponded with higher bronchial and systemic IL-5 levels in the progesterone group. The ratio of IL-4/IFN-gamma levels in bronchoalveolar lavage fluid and numbers of eosinophil colony-forming units in the bone marrow were also elevated in the latter group. Progesterone, however, did not influence allergen-specific IgE production, nor did it affect bronchial responses in Sal-challenged mice. CONCLUSION: Our data show that exogenously administered progesterone aggravates the phenotype of eosinophilic airway inflammation in mice by enhancing systemic IL-5 production. Progesterone also increases bronchial hyper-reactivity.     

Therapeutic effects of DNA vaccine on allergen-induced allergic airway inflammation in mouse model

Vaccination with DNA encoding Dermatophagoides pteronyssinus group 2 （Der p 2） allergen previously showed its effects of immunologic protection on Der p 2 allergen-induced allergic airway inflammation in mice. In present study, we investigated whether DNA vaccine encoding Der p 2 could exert therapeutic role on allergen-induced allergic airway inflammation in mouse model and explored the mechanism of DNA vaccination in asthma specific-allergen immunotherapy. After sensitized and challenged by Der p 2, the BALB/c mice were immunized with DNA vaccine. The degrees of cellular infiltration were scored. IgE levels in serum and IL-4/IL-13 levels in BALF were determined by ELISA. The lung tissues were assessed by histological examinations. Expressions of STAT6 and NF-kB in lung were determined by immunohistochemistry staining. Vaccination of mice with DNA vaccine inhibited the development of airway inflammation and the production of mucin induced by allergen, and reduced the level of Der p 2-specific IgE level. Significant reductions of eosinophil infiltration and levels of IL-4 and IL-13 in BALF were observed after vaccination. Further more, DNA vaccination inhibited STAT6 and NF-kB expression in lung tissue in Der p 2-immunized mice. These results indicated that DNA vaccine encoding Der p 2 allergen could be used for therapy of allergen-induced allergic airway inflammation in our mouse model. Cellular ＆ Molecular Immunology.     

BCG对哮喘小鼠气道炎症反应的影响

目的：探讨卡介苗(BCG)对卯蛋白(OVA)致敏小鼠肺组织中T细胞的在体调节及其对气道炎症的作用。方法：将30只BALB／c小鼠分为3组，第1组吸入雾化的OVA(1次／(1．20min／次，连续10d)，建立致敏模型。第2组(对照)吸入雾化的生理盐水(时间和次数同第1组)；第3组(治疗组)于致敏前10d及14d，各皮内注射BCG 1次，致敏后6d吸入雾化的纯蛋白衍生物(PPD)。用SABC免疫组化法，检测肺组织中CD4^ 、CD8^ 及IFN—γ^ 细胞的变化，以及肺组织和支气管肺泡灌洗液(BALF)中炎性细咆的变化。结果：OVA致敏组小鼠肺组织中CD4^ T细胞增加，CD8^ T细胞无明显变化，主要表现为IFN—γ^-／CD4^ T细胞数的增加，IFN—γ^ ／CD4^ 细胞的比例降低。BCG治疗后，肺组织中CD4^ T细胞减少，CD8^ T细胞数大量增加；IFN—γ^ ／CD4^ 细胞的比例明显增大；BALF中炎性细胞数减少。结论：BCG可在体上调Th1细胞，减轻实验性哮喘动物气道的炎症反应。     

Influence of smoking on airway inflammation and remodelling in asthma

Background Although exposure to tobacco smoke has been associated with increased morbidity and mortality, cigarette smoking is still common in the asthmatic population. Induced sputum neutrophilia has been observed in asthmatic smokers, but the effects of regular smoking on their bronchial mucosa morphology remain to be defined. This study documents the inflammatory and remodelling features in bronchial biopsies of smoking compared with non‐smoking asthmatics. Methods We analysed bronchial biopsies from 24 steroid‐naïve young subjects with mild asthma: 12 non‐smoking and 12 currently smoking subjects. In addition to airway morphology assessment, inflammation and remodelling were analysed by immunohistochemistry using antibodies against CD3, CD68, major basic protein, neutrophil elastase, and tryptase. Expression of the cytokines IL‐4, IL‐5, IL‐8, IFN‐γ, transforming growth factor‐β, and TNF was determined by in situ hybridization. Results Compared with non‐smoking asthmatic subjects, smoking asthmatics' bronchial mucosa showed squamous cell metaplasia, in addition to increased expression of subepithelial neutrophil elastase, IFN‐γ, and intraepithelial IL‐8. Conclusions Smoking status modifies morphological and inflammatory processes in young subjects with mild asthma. The changes may possibly affect asthma treatment responses and clinical outcomes.     

The effects of nodakenin on airway inflammation,hyper-responsiveness and remodeling in a murine model of allergic asthma

Context: Nodakenin is a major coumarin glucoside in the root of Peucedanum decursivum Maxim, a commonly used traditional Chinese medicine for the treatment of asthma and chronic bronchitis for thousands of years.

Objective: In this work, the anti-asthma potential of nodakenin was studied by investigation of its effect to suppress airway inflammation, hyper-responsiveness and remodeling in a murine model of chronic asthma.

Materials and methods: BALB/c mice sensitized to ovalbumin (OVA) were challenged with aerosolized OVA for 8 weeks, orally administered with nodakenin at doses of 5, 10 and 20?mg/kg before each OVA challenge.

Results: Compared with the model group, nodakenin treatment markedly inhibited airway inflammation, hyper-responsiveness and remodeling, showing improvement in subepithelial fibrosis, smooth muscle hypertrophy, and goblet cell hyperplasia, and decreased levels of interleukin (IL)-4, IL-5, IL-13 and matrix metalloproteinase-2/-9 in bronchoalveolar lavage fluid, and the level of OVA-specific IgE in serum. In addition, the NF-κB DNA-binding activity in lung tissues was also reduced by nodakenin treatment.

Conclusions: These data indicated that nodakenin might mitigate the development of chronic experimental allergic asthma.     

Composition of the immunoglobulin classic antigen-binding site regulates allergic airway inflammation in a murine model of experimental asthma

Background When bound to mast cell FcɛRI, IgE serves as antigen receptor for allergic reactions, permitting specific identification of the allergen. Although the core of the classic antigen-binding site is heavy chain complementarity determining region 3 (CDR-H3), recent studies suggest that allergens might also bind IgE in a superantigen-like fashion outside the classic antigen-binding site.
Objective We sought to evaluate the contribution of the classic CDR-H3-centric antigen-binding site to the development of an allergic phenotype.
Methods Using a murine model of experimental asthma, we characterized a gene-targeted mouse strain expressing an altered range of CDR-H3s (ΔD-iD mice) in response to the hydrophobic allergen ovalbumin (OVA). Mutant and wild-type ( wt ) mice were sensitized intraperitoneally with OVA; non-sensitized mice served as controls.
Results We found the composition of the classic CDR-H3-centric antigen-binding site to be critical for the development of characteristic aspects of allergic asthma. (i) Compared with wt animals, ΔD-iD mice showed a significantly less pronounced OVA -induced rise in allergen-specific IgE levels and hence in total serum IgE levels. (ii) In addition, ΔD-iD mice demonstrated a significant reduction in eosinophilic airway inflammation, as well as in interleukin-4 (IL-4), IL-5 and IL-13 levels in BAL fluids.
Conclusion Allergic sensitization and airway inflammation depend on the composition of the predominant CDR-H3 repertoire, suggesting that the classic CDR-H3-centric antigen-binding site plays a crucial role in creating the immunological interface between allergen and IgE. Our results further emphasize a central role of IgE, not only in mediating but also in regulating the allergic immune response.     

Mannitol challenge for assessment of airway responsiveness,airway inflammation and inflammatory phenotype in asthma

Background Assessment of airway inflammation in asthma is becoming increasingly important, as the inflammatory phenotype underpins the treatment response. Objective This study aimed to evaluate mannitol as a tool for assessing airway responsiveness and airway inflammation in asthma, compared with hypertonic saline. Methods Fifty‐five subjects with stable asthma completed a hypertonic (4.5%) saline challenge and a mannitol challenge at two separate visits, performed 48–72 h apart, in random order. Results Induced sputum was obtained from 49 (89%) subjects during the saline challenge and 42 (76%) subjects during the mannitol challenge (P>0.05). There was a significant correlation between the greatest percentage fall in forced expiratory volume in 1 s (FEV₁) (r=0.6, P<0.0001), the dose–response slope (r=0.73), cumulative dose (r=0.55) and PD15 (r=0.46) for mannitol and hypertonic saline. The greatest percentage fall in FEV₁ to mannitol was less in non‐eosinophilic asthma. There was a lower total cell count in mannitol vs. hypertonic‐saline‐induced sputum. However, sputum eosinophils and neutrophils were not significantly different. Using mannitol, a higher proportion of subjects were classified as having eosinophilic asthma. There were no differences in IL‐8, neutrophil elastase or matrix‐metalloproteinase 9 concentrations in sputum samples induced with mannitol or hypertonic saline. Conclusion We conclude that mannitol can be used to induce good‐quality sputum, useful for analysis of inflammatory mediators and for predicting the inflammatory phenotype in asthma. Cite this as: L. G. Wood, H. Powell and P. G. Gibson, Clinical & Experimental Allergy, 2010 (40) 232–241.     

Airway inflammation and cough sensitivity in cough-variant asthma

BACKGROUND: Mechanisms underlying cough and bronchoconstriction in patients with cough-variant asthma (CVA) are not well established. Differences in location or degree of activation of eosinophils and allergic cytokines have been suggested as the likely causes. To address this issue, we have carried out a comparative study of airway inflammatory markers between patients with CVA and classic asthma (CA). The relationship between these markers with airway hyperresponsiveness (AHR) and cough sensitivity has also been studied. METHODS: Twenty-seven non-smokers and steroid-naive patients with CVA (12) and CA (15) were examined. Capsaicin challenge, histamine bronchoprovocation test, nitric oxide levels in exhaled air and sputum induction were performed in all of them. Differential cell sputum recount and supernatant concentrations of eosinophil granule-derived cationic proteins (ECP), interleukin (IL)5, IL8 and tumour necrosis factor (TNF)-alpha were also measured. RESULTS: There were no significant differences in either the inflammatory pattern of soluble markers or differential cell counts between CA and CVA. Histamine PC20 was correlated with IL-5 in CVA, whereas it was associated with sputum eosinophilia in CA. Cough sensitivity (log C5) and histamine PC20 were inversely related in CA. CONCLUSIONS: Although the pattern of inflammatory sputum markers in patients with asthma and cough-variant asthma is similar, its relation with bronchial hyperreactivity and cough sensitivity is different in each group.     

Eosinophilic airway inflammation and the prognosis of childhood asthma

BACKGROUND: Eosinophilic airway inflammation is a key pathophysiological feature of asthma that can predict treatment response. However, the prognostic value of sputum eosinophilia is not established. OBJECTIVE: The aim of this study was to determine the influence of induced sputum eosinophilia on the prognosis of childhood asthma. METHODS: A cohort of children with asthma was evaluated by induced sputum analysis at inception and classified as having either eosinophilic asthma (EA) (sputum eosinophils >2.5%) or non-eosinophilic asthma (NEA). After a mean follow-up period of 5 years, eligible subjects (n=83) were contacted and 69 subjects (33 EA, 36 NEA) evaluated. The children had a mean age of 15.9 years, and 61% were male. RESULTS: Children with EA reported more wheeze during the follow-up period (27% vs. 6% wheezed most years; P<0.0001), increased night waking during the past 12 months (28% vs. 3% reported weekly waking; P=0.01), and greater impairment of quality of life due to asthma (P=0.04). Subsequent beta2-agonist use was increased in children with EA (P=0.02), although there was no difference in corticosteroid use. In EA, subsequent forced expiratory volume in 1 s/forced vital capacity was lower (79% vs. 86%; P=0.01) and grass pollen allergy was more prevalent (77% vs. 27%; P=0.006). CONCLUSION: In children, eosinophilic airway inflammation is associated with deteriorating asthma over time. This is consistent with the hypothesis that airway inflammation has an adverse impact on the prognosis of childhood asthma, and suggests a role for monitoring inflammation in asthma management.     

Expression of tyrosine hydroxylase and neuropeptide tyrosine in mouse sympathetic airway-specific neurons under normal situation and allergic airway inflammation

BACKGROUND: The traditional neurotransmitter catecholamine and the neuropeptide tyrosine in sympathetic airway nerves have been proposed to be involved in the pathogenesis of airway diseases. OBJECTIVE: The aim of the present study was to investigate the effect of allergic airway inflammation on the expression of catecholamine enzyme tyrosine hydroxylase (TH), neuropeptide tyrosine (NPY) and tachykinins in mouse sympathetic airway ganglia. METHODS: Using neuronal tracing in combination with immunohistochemistry, the present study was designed to characterize TH, NPY and tachykinin profiles of superior cervical (SCG) and stellate ganglia after allergen challenge. RESULTS: The vast majority of fast blue-labelled SCG neurons (allergen: 97.5+/-1.22% (mean+/-SEM) vs. controls: 94.5+/-1.48%, P=0.18) and stellate neurons (allergen: 95.3+/-1.01% vs. controls: 93.6+/-1.33%, P=0.34) were immunoreactive for TH. Of the TH immunoreactive and fast blue-labelled SCG neurons, 52.0+/-1.01% allergen vs. 51.2+/-3.58% controls (P=0.83) and stellate neurons, 57.3%+/-0.97 allergen vs. 56.4+/-1.65% controls (P=0.64) were positive for TH only but not NPY, whereas 45.3+/-1.05% allergen vs. 43.3+/-1.18% controls (P=0.47) of fast blue-labelled SCG neurons and 37.9+/-0.86% allergen vs. 37.1+/-1.24% controls (P=0.62) of fast blue-labelled stellate neurons were immunoreactive for both TH and NPY immunoreactivities. There was a trend of an increase, but not significant one, in the percentage of TH-/NPY-immunoreactive and fast blue-labelled neurons in allergen-treated animals in comparison with the controls. Tachykinins, however, were not expressed by sympathetic neurons and were also not induced in sympathetic neurons after allergen challenge. CONCLUSION: The present study indicates that allergic airway inflammation does not alter the expression of noradrenalin and NPY in sympathetic ganglia and also shows that sympathetic neurons do not respond to allergic airway inflammation with tachykinins induction. However, a participation of catecholamine and NPY in the pathogenesis of allergic airway inflammation cannot be excluded in the present study as a higher neurotransmitter output per neuron following allergen challenge could be possible.     

Inter-relationships between airway inflammation, reticular basement membrane thickening and bronchial hyper-reactivity to methacholine in asthma; a systematic bronchoalveolar lavage and airway biopsy analysis

Background Asthma is accepted as a disease characterized by airway inflammation, with evidence that airway structural changes, or ‘remodelling’ occurs. There are few studies relating airway physiology, inflammation and remodelling, however. We have carried out a study of inter‐relationships between airway inflammation, airway remodelling, reticular basement membrane (RBM) thickening, and bronchial hyper‐reactivity (BHR), before and after high‐dose inhaled corticosteroid (fluticasone propionate 750 μg b.d.), in a group of relatively mild but symptomatic, steroid naïve asthma patients. Methods Double‐blind, randomized, placebo‐controlled, parallel group study of inhaled corticosteroid (ICS) in 35 asthmatics, with bronchoalveolar lavage (BAL) and airway endobronchial biopsy (EBB) for inflammatory cell profiles and EBB for airway remodelling carried out at baseline, 3 and 12 months. Results At baseline RBM thickening was related to BAL mast cells and EBB eosinophil counts. In turn baseline log EBB EG2 eosinophil count, log%BAL epithelial cells and log RBM thickness explained 55% of the variability in BHR. Conclusion We provide new information that airway inflammation, remodelling, and BHR in asthma are inter‐related and improved by ICS therapy. Our data potentially support the need for early and long‐term intervention with ICS even in relatively mild asthmatics, and the need to further assess the potential merit of longitudinal BHR testing in management of some patients, as this may reflect both airway inflammation and remodelling.     

Allergic airway inflammation induces tachykinin peptides expression in vagal sensory neurons innervating mouse airways

BACKGROUND: Allergic airway inflammation has been shown to induce pro-inflammatory neuropeptides such as tachykinin peptides substance P (SP) and neurokinin A (NKA) together with related peptide like calcitonin gene-related peptide (CGRP) in nodose sensory neurons innervating guinea-pig airways. OBJECTIVE: The present study was designed to examine the effects of allergen sensitization and challenge on the SP/NKA expression in the jugular-nodose ganglion neurons innervating the murine airways. METHODS: Using retrograde neuronal tracing technique in combination with double-labelling immunohistochemistry, the expression of SP/NKA was investigated in a murine model of allergic airway inflammation. RESULTS: Allergic airway inflammation was found to induce the expression of SP/NKA (13.2+/-1.43% vs. 5.8+/-0.37%, P<0.01) in large-diameter (>20 microm) vagal sensory neurons retrograde labelled with Fast blue dye from the main stem bronchi. CONCLUSION: Based on the induction of tachykinins in airway-specific large-sized jugular-nodose ganglia neurons by allergic airway inflammation, the present study suggests that allergen sensitization and challenge may lead to de novo induction of tachykinins in neurons. This may partly contribute to the pathogenesis of airways diseases such as allergic airway inflammation.