Phosphoinositide 3-kinase-delta inhibitor reduces vascular permeability in a murine model of asthma

BACKGROUND: Bronchial asthma is characterized by inflammation of the airways, which is usually accompanied by increased vascular permeability, resulting in plasma exudation. Vascular endothelial growth factor (VEGF) has been implicated in contributing to asthmatic tissue edema through its effect on vascular permeability. Many cellular responses of VEGF are regulated by the lipid products of phosphoinositide 3-kinase (PI3K). However, the effect of PI3K catalytic subunit p110delta on VEGF-mediated signaling is unknown. Recently, an isoform-specific small molecule inhibitor, IC87114, which is selective for p110delta catalytic activity, has been identified. OBJECTIVE: We have sought to investigate the role of PI3K-delta, more specifically in the increase of vascular permeability. METHODS: Female BALB/c mice were sensitized and challenged with ovalbumin. We have investigated the effect of IC87114 on airway inflammation, T(H)2 cytokines expression, airway hyperresponsiveness, plasma extravasation, hypoxia-inducible factor 1alpha expression, and VEGF expression in a murine model of asthma. RESULTS: Our current study has revealed that IC87114 reduces antigen-induced airway infiltration of inflammatory cells, secretion of T(H)2 cytokines in lungs, airway hyperresponsiveness, and vascular permeability. Moreover, we have found that inhibition of p110delta reduces ovalbumin-induced upregulation of VEGF level. CONCLUSION: These results suggest that PI3K-delta inhibitor attenuates antigen-induced airway inflammation and hyperresponsiveness by preventing vascular leakage in mice. CLINICAL IMPLICATIONS: These findings provide a crucial molecular mechanism for the potential role of PI3K-delta in asthma and other airway inflammatory disorders.     

Hydrogen peroxide induces vascular permeability via regulation of vascular endothelial growth factor

Oxidative stress plays critical roles in initiation and/or worsening of respiratory disease process. Although reactive oxygen species (ROS) are shown to cause vascular leakage, the mechanisms by which ROS induce an increase in vascular permeability are not clearly understood. In this study, we have used a murine model to evaluate the effect of hydrogen peroxide (H(2)O(2)) to examine roles of ROS and the molecular mechanism in vascular permeability. The results have revealed that ROS levels, vascular endothelial growth factor (VEGF) expression, hypoxia-inducible factor-1alpha protein level, airway hyperresponsiveness, and vascular permeability are increased after inhalation of H(2)O(2). Administration of antioxidants markedly reduced plasma extravasation and VEGF levels in lungs treated with H(2)O(2). These results indicate that ROS may modulate vascular permeability via upregulation of VEGF expression.     

Alveolar macrophage-derived vascular endothelial growth factor contributes to allergic airway inflammation in a mouse asthma model

Vascular endothelial growth factor (VEGF) is a potent proangiogenic factor that correlates with vascular permeability and remodelling in asthma. Recently, alveolar macrophages (AM) were shown to be an important source of VEGF during lung injury. Our previous studies demonstrated that AM are an important subset of macrophages in the initiation of asthmatic symptoms. Here, we further investigated whether AM-derived VEGF was required for allergic airway inflammation in asthma. In this study, we reported that the expression of VEGF in AM was significantly increased after allergen challenge. Depleting AM or neutralizing VEGF in alveolus prevented ovalbumin (OVA)-induced asthma-related inflammation by inhibiting the infiltration of inflammatory cells in the lung, reduced the level of the cytokines, IL-4, IL-5, and IL-13, in the bronchoalveolar lavage fluid (BALF) and decreased airway hyperresponsiveness (AHR). Moreover, the inhibition of miR-20b increased the protein level of VEGF in normal AM; conversely, increasing miR-20b in asthmatic AM resulted in decreased VEGF protein levels. These findings suggest that AM-derived VEGF is necessary for allergic airway inflammation in asthmatic mice and miR-20b negatively regulates this expression.     

Preclinical pharmacology of pranlukast

Cysteinyl leukotrienes (cysLTs) exert potent biological actions through their specific receptors. They induce pathophysiological responses such as potent bronchoconstriction, mucus secretion, mucosal oedema and eosinophil infiltration. The regulation of these responses may provide a novel therapeutic approach in the treatment of allergic and inflammatory diseases. We have developed a cysLT receptor antagonist, pranlukast as one of cysLT modifiers. Preclinical pharmacology studies demonstrate that pranlukast is a potent, selective and orally active antagonist of cysLT₁ receptor. Pranlukast competed with [³H]cysLTs for binding to cysLT receptors on guinea-pig lung and nasal mucosa membranes and inhibited LTD₄-induced calcium mobilization through human cysLT₁ receptors expressed on CHO cells. Oral administration of pranlukast inhibited cysLTs-induced symptomatology in guinea-pigs. In sensitized guinea-pig models of asthma, pranlukast inhibited antigen-induced early and late phase bronchoconstrictions, airway hyperresponsiveness, microvascular leakage and eosinophil infiltration into airways. Furthermore, in sensitized Brown Norway rats, pranlukast suppressed airway smooth muscle proliferation and collagen deposition characteristic of airway wall remodelling. In guinea-pig models of allergic rhinitis, pranlukast potently blocked a biphasic nasal obstruction resistant to antihistamines, sneezing and nasal secretion induced by nasal hyperresponsiveness, suggesting the prominent efficacy of pranlukast in allergic rhinitis. From these pharmacological properties of pranlukast, it is suggested that cysLT₁ receptor antagonists are useful and beneficial as a new class of antiallergic and anti-inflammatory agents in the treatment and management of asthma and allergic rhinitis.     

Analysis of vascular endothelial growth factor levels in induced sputum samples from patients with cough variant asthma.

BACKGROUND: We previously found that vascular endothelial growth factor (VEGF) levels in induced sputum samples are increased in patients with classic asthma and are associated with the degree of airflow obstruction and airway microvascular permeability. OBJECTIVE: To examine VEGF levels and the degree of airway microvascular permeability in patients with cough variant asthma (CVA). METHODS: Levels of VEGF in induced sputum samples and airway microvascular permeability were examined in 12 controls, 16 patients with CVA, and 16 patients with classic asthma. We also evaluated the relationship between VEGF level and the clinical features of these 2 disorders. RESULTS: Mean (SD) VEGF levels and airway vascular permeability index values were significantly higher in patients with CVA (VEGF: 2,520 [1,050] pg/mL; P < .001; vascular permeability index: 0.017 [0.006]; P = .003) and classic asthma (4,750 [1,260] pg/mL; P < .001; 0.028 [0.009]; P < .001) than in controls (1,420 [1,230] pg/mL; 0.009 [0.003]). Furthermore, these values were significantly higher in patients with classic asthma vs CVA. We also found significant correlations between VEGF level and airway vascular permeability index in patients with CVA (r = 0.60; P = .02) vs classic asthma (r = 0.83; P = .001). Furthermore, VEGF levels were inversely correlated with the degree of airflow obstruction and airway hyperresponsiveness to methacholine in patients with CVA and classic asthma. CONCLUSIONS: Airway microvascular hyperpermeability induced by elevated VEGF levels contributes to abnormal airway function in CVA and classic asthma, and differences in the clinical features of these 2 disorders may depend on airway VEGF levels.     

Leukotriene D4 upregulates eosinophil adhesion via the cysteinyl leukotriene 1 receptor

BACKGROUND: Eosinophils (EOS) are one of the cellular sources of cysteinyl leukotrienes (cysLTs) in allergic inflammation. There is evidence that cysLT(1)receptor antagonists possess anti-inflammatory properties in vivo in asthmatic airways. Although the exact mechanism of action remains unknown, cysLTs might regulate the cellular responses involved in allergic inflammation. OBJECTIVE: The present study was undertaken to examine whether LTD(4)modifies the adhesive property of EOS. METHODS: EOS were isolated from the blood of healthy subjects. Their adhesion to tissue culture plates or recombinant human (rh) adhesion proteins was then examined in the presence or absence of LTD(4). RESULTS: LTD(4)significantly augmented EOS adhesion to tissue culture plates (adhesion: 5.0% +/- 0.5% by medium control vs 9.1% +/- 1.2% by 1 micromol/L; P <.01; n = 10). The enhanced adhesion induced by LTD(4) was blocked by pranlukast, a cysLT(1) receptor antagonist, or an anti-beta(2) integrin antibody. Flow cytometry analysis revealed that LTD(4) significantly enhanced the expression of CD11b and CD18 on the EOS surface. Finally, LTD(4) augmented EOS adhesion to rh intercellular cell adhesion molecule 1 but not to rh vascular cell adhesion molecule 1 or fibronectin. CONCLUSIONS: The results suggest that LTD(4) directly upregulates the adhesive property of EOS via the cysLT(1) receptor and beta(2) integrin. LTD(4) generated from EOS or cells of some other type might contribute to the development of phenotypic change in airway EOS.     

Increased levels of vascular endothelial growth factor in induced sputum in asthmatic patients

BACKGROUND: Vascular endothelial growth factor (VEGF) is highly expressed in the airway of asthmatic patients. As VEGF increases airway vascular permeability, consequent thickening of the airway wall mucosa may lead to narrowing of the airway lumen. OBJECTIVE: We evaluated the relationship between VEGF levels in induced sputum and eosinophilic inflammatory profiles, and the degree of airway vascular permeability in asthmatic patients and we evaluated the effect of inhaled corticosteroids on VEGF levels in induced sputum. METHODS: Induced sputum specimens were obtained from 28 glucocorticosteroids free asthmatics and 11 healthy control subjects. We examined VEGF levels and airway vascular permeability index in induced sputum. After the initial sputum induction, 21 asthmatics received 8-week inhaled beclomethasone dipropionate (BDP, 800 micro g/day) therapy, then sputum induction was repeated. RESULTS: The VEGF levels in asthmatics were significantly higher than in healthy control subjects (P < 0.0001). The VEGF levels were negatively correlated with forced expiratory volume of 1 s (FEV1, % predicted, r = - 0.68, P < 0.001), the percentage of eosinophils (r = 0.51, P < 0.01) and ECP levels (r = 0.39, P < 0.05). Moreover, the VEGF levels were significantly correlated with airway vascular permeability index (r = 0.61, P < 0.001). After 8-week inhaled BDP therapy, the VEGF levels were significantly decreased compared to pretreatment levels (P < 0.0001) and the VEGF levels were significantly correlated with airway vascular permeability index even in post-treatment asthmatics (r = 0.62, P < 0.01). CONCLUSION: The VEGF levels in induced sputum were increased in asthmatics and its levels were associated with degree of airway narrowing and airway vascular permeability. These findings provide strong evidence that VEGF may play an important role in the pathogenesis of bronchial asthma.     

Expression of vascular endothelial growth factor, basic fibroblast growth factor, and angiogenin immunoreactivity in asthmatic airways and its relationship to angiogenesis

BACKGROUND: Angiogenesis is a prerequisite for airway remodeling in bronchial asthma. Several growth factors may play important roles in inflammation and angiogenesis through effects on inflammatory cell infiltration or neovascularization. OBJECTIVE: We sought to compare bronchial vascularity and expression of vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF), and angiogenin in bronchial biopsy specimens from asthmatic and healthy control subjects. METHODS: Bronchial biopsy specimens were obtained from 16 asthmatic subjects and 9 normal control subjects. The number of vessel profiles and the vascular area per unit area on a histologic section were estimated by using computerized image analysis after staining for type IV collagen in vessel walls. Numbers of VEGF+, bFGF+, and angiogenin+ cells were determined by means of immunoreactivity. RESULTS: The airways of asthmatic subjects had significantly more vessels (P < .05) and greater vascular area (P < .001) than that observed in control subjects. Asthmatic subjects exhibited higher VEGF and bFGF and angiogenin immunoreactivity in the submucosa than did control subjects (P < .001, respectively). Significant correlations were detected between the vascular area and the numbers of angiogenic factor-positive cells (VEGF: rs = 0.93, P < .001; bFGF: rs = 0.83, P < .001; angiogenin: rs = 0.88, P < .001) within the asthmatic airways. Furthermore, the degree of vascularity was inversely correlated with airway caliber and airway responsiveness. Colocalization analysis revealed that the angiogenic factor-positive cells were CD34+ cells, eosinophils, and macrophages. CONCLUSION: Our results suggest that increased vascularity of the bronchial mucosa in asthmatic subjects is closely related to the expression of angiogenic factors, which may then contribute to the pathogenesis of asthma.     

Effects of antiasthmatic agents on the functions of peripheral blood monocyte-derived dendritic cells from atopic patients

BACKGROUND: Dendritic cells (DCs), the antigen-presenting cells in the airway, play a critical role in asthma. Nevertheless, there is little information on the effects of antiasthmatic agents on DCs. OBJECTIVES: The purpose of the present study was to determine the effects of representative antiasthmatic agents, including cysteinyl leukotriene (cysLT) 1 receptor antagonists, corticosteroid, and tacrolimus, on DCs in inducing allergy. METHODS: Human peripheral blood monocyte-derived DCs (MoDCs) generated from atopic and healthy subjects were pulsed with Dermatophagoides farinae allergen in the presence of medium alone, pranlukast, montelukast, dexamethasone, or tacrolimus. The mRNA expressions of cysLT receptor, cysLTs producing enzymes, and various surface markers on MoDCs, as well as the concentrations of cysLTs, IL-10, and IL-12 in cultured supernatants, were determined. MoDCs were also cocultured in vitro with autologous CD4(+) T cells, and IL-5 and IFN-gamma production was measured. RESULTS: MoDCs of atopic patients expressed mRNAs of cysLT1 receptor and cysLT-producing enzymes, and allergen pulsing significantly increased cysLT production. MoDCs of atopic patients showed a T(H)2-favoring phenotype and induced T(H)2-skewed cytokine production from autologous CD4(+) T cells. Dexamethasone and tacrolimus inhibited allergen-pulsed MoDC-induced cytokine production by autologous CD4(+) T cells without and with IL-10 inhibition, respectively. CysLT1 receptor antagonists had no effect on MoDC functions. CONCLUSION: Our results indicate that MoDCs of atopic patients induce a T(H)2 reaction. Corticosteroid and tacrolimus, but not cysLT1 receptor antagonists, inhibit T(H)2 reactions, and this effect is probably mediated through different pathways.     

AMPK activation reduces vascular permeability and airway inflammation by regulating HIF/VEGFA pathway in a murine model of toluene diisocyanate-induced asthma

Background

Occupational asthma is characterized by airway inflammation and hyperresponsiveness associated with increased vascular permeability. AMP-activated protein kinase (AMPK) has been suggested to be a novel signaling molecule modulating inflammatory responses.

Objective

We sought to evaluate the involvement of AMPK in pathogenesis of occupational asthma and more specifically investigate the effect and molecular mechanisms of AMPK activation in regulating vascular permeability.

Methods

The mechanisms of action and therapeutic potential of an AMPK activator, 5-aminoimidazole-4-carboxamide-1-β-d-ribofuranoside (AICAR) were tested in a murine model of toluene diisocyanate (TDI)-induced asthma.

Results

AICAR attenuated airway inflammation and hyperresponsiveness increased by TDI inhalation. Moreover, TDI-induced increases in levels of hypoxia-inducible factor (HIF)-1α, HIF-2α, vascular endothelial growth factor A (VEGFA), and plasma exudation were substantially decreased by treatment with AICAR. Our results also showed that VEGFA expression was remarkably reduced by inhibition of HIF-1α and HIF-2α with 2-methoxyestradiol (2ME2) and that an inhibitor of VEGFA activity, CBO-P11 as well as 2ME2 significantly suppressed vascular permeability, airway infiltration of inflammatory cells, and airway hyperresponsiveness induced by TDI. In addition, AICAR reduced reactive oxygen species (ROS) generation and levels of malondialdehyde and T-helper type 2 cytokines (IL-4, IL-5, and IL-13), while this agent enhanced expression of an anti-inflammatory cytokine, IL-10.

Conclusions

These results suggest that AMPK activation ameliorates airway inflammatory responses by reducing vascular permeability via HIF/VEGFA pathway as well as by inhibiting ROS production and thus may be a possible therapeutic strategy for TDI-induced asthma and other airway inflammatory diseases.     

Increased vascular permeability precedes cellular inflammation as asthma control deteriorates

Background Airway microcirculation is abnormal in asthma but the role of vascular changes in asthma deteriorations remains poorly defined. We prospectively assessed the vascular changes accompanying worsening of asthma control by using an inhaled corticosteroid (ICS) dose-reduction model.
Objectives To evaluate airway vascularity, vascular permeability and expression of vascular endothelial growth factor (VEGF) in early asthma deterioration induced by ICS back-titration.
Methods Twenty mild-to-moderate persistent symptomatic asthmatics on low-to-moderate ICS were recruited and treated with 4 weeks of high-dose fluticasone propionate (1000 μg/day) to achieve symptom control. This was followed by dose reduction to half of the pre-study doses for 4–8 weeks until the symptoms began to return. Endobronchial biopsy and bronchoalveolar lavage (BAL) samples were obtained after both treatment periods.
Results Vascularity as measured by the number and size of blood vessels, as well as VEGF expression did not change following ICS reduction. Even on high-dose ICS, perivascular albumin staining and BAL microalbumin levels in asthmatic subjects, as markers of permeability, were elevated when compared with normal subjects and both further increased significantly after ICS reduction. There was a significant association between changes in vascular leakiness and clinical deterioration. Increases in airway albumin correlated with previously reported increases in airway wall infiltration with T lymphocytes.
Conclusions Our results suggest that airway vascular leakage is a major pathophysiologic feature of early asthma deterioration, occurring before recrudescence of cellular inflammation.     

Leukotriene D4 and eosinophil transendothelial migration, superoxide generation, and degranulation via beta2 integrin.

BACKGROUND: Evidence indicates that cysteinyl leukotriene (cysLT) 1 receptor antagonists possess anti-inflammatory properties in asthmatic patients in vivo. Although the exact mechanisms of these actions remain unknown, cysLTs regulate the locomotion and functions of eosinophils. We previously reported that leukotriene D4 augments the expression of eosinophil beta2 integrin and the adhesion of eosinophils to rh intercellular adhesion molecule 1 via beta2 integrin. OBJECTIVE: To examine whether leukotriene D4 modifies the transendothelial migration (TEM) and effector functions of eosinophils. METHODS: We evaluated the effects of leukotriene D4 on (1) eosinophil TEM across human umbilical vein endothelial cells, (2) superoxide anion (O2-) generation, and (3) eosinophil-derived neurotoxin release in eosinophils isolated from the blood of healthy individuals. RESULTS: Leukotriene D4 (0.1-1 microM) significantly induced eosinophil TEM, O2- generation, and eosinophil-derived neurotoxin release. Pranlukast, a cysLT1 receptor antagonist, significantly inhibited all of these parameters, although the inhibitory effect on O2- generation was partial. All of these responses were significantly inhibited by anti-beta2 integrin but not by anti-alpha4 integrin antibodies. CONCLUSIONS: Leukotriene D4 directly up-regulates the TEM and effector functions of eosinophils mainly via the cysLT1 receptor and beta2 integrin. These effects of leukotriene D4 probably contribute to the manifestation of eosinophil inflammation in asthmatic airways.     

A potent antiangiogenic factor, endostatin prevents the development of asthma in a murine model

BACKGROUND: Clinical studies suggest a role for angiogenesis in the development and persistence of chronic asthma, but whether angiogenic mediators contribute to acute asthma has not been fully studied. OBJECTIVE: The aim of this study was to investigate a role of vascular endothelial growth factor (VEGF), a major angiogenic and proinflammatory mediator, in allergen-induced acute asthma and to determine whether endostatin/Fc, a potent antiangiogenic factor can attenuate allergic airway responses. METHODS: We sensitized BALB/c mice with ovalbumin. We measured serum VEGF and examined immunoreactive VEGF around the airways 48 hours after the last challenge with either aerosolized PBS or ovalbumin once per day for 3 days. We also treated ovalbumin-sensitized mice with either endostatin/Fc or control fusion protein at the time of challenge with ovalbumin. We analyzed allergic airway responses 48 hours after the last ovalbumin challenge. RESULTS: Ovalbumin challenge induced immunolocalization of numerous VEGF-positive cells around airways and increased serum VEGF levels. Treatment with endostatin/Fc inhibited the airway hyperresponsiveness, pulmonary allergic inflammation, production of ovalbumin-specific IgE, and lung inflammatory mediators. Both VEGF-dependent and independent mechanisms are indicated by results using antibody blockade of VEGF receptors, which caused decreased allergic pulmonary inflammation but did not alter airway hyperresponsiveness or serum IgE levels. CONCLUSION: These data demonstrate for the first time that recombinant endostatin can prevent the development of asthma features in a mouse model and suggest that this class of agents merits further study as novel therapeutics for asthma.     

Imbalance between vascular endothelial growth factor and endostatin levels in induced sputum from asthmatic subjects

BACKGROUND: Angiogenesis has recently attracted considerable attention as a component of airway remodeling in bronchial asthma. Vascular endothelial growth factor (VEGF) is highly expressed in asthmatic airways, and its contribution to airway remodeling has been reported. Although angiogenesis is regulated by a balance of angiogenic and antiangiogenic factors, the relative levels of antiangiogenic factors in asthmatic airways have not been evaluated. OBJECTIVE: We sought to determine whether an imbalance between angiogenic and antiangiogenic factors exists in asthmatic airways. METHODS: We simultaneously measured VEGF and endostatin levels and evaluated their correlation and balance in induced sputum from 18 steroid-naive asthmatic subjects and 11 healthy control subjects. After initial sputum induction, asthmatic subjects underwent 8 weeks of inhaled beclomethasone dipropionate (BDP; 800 microg/d) therapy, and sputum induction was then repeated. RESULTS: VEGF and endostatin levels in induced sputum were significantly higher in asthmatic subjects than in control subjects (P <.001). There was a significant correlation between VEGF and endostatin levels in both control subjects (r = 0.995, P <.001) and asthmatic subjects (r = 0.923, P <.001). Moreover, the VEGF/endostatin level ratio in asthmatic subjects was significantly higher than that in control subjects (P <.0001). After 8 weeks of inhaled BDP therapy, the VEGF level in induced sputum in asthmatic subjects was significantly decreased (P <.001), whereas the endostatin level was not. A correlation between VEGF and endostatin levels existed even after BDP therapy (r = 0.861, P <.001). Moreover, the VEGF/endostatin level ratio was significantly decreased to the same level as in the control subjects after BDP therapy (P <.0001). CONCLUSION: There was an imbalance between VEGF and endostatin levels in induced sputum from asthmatic subjects. This imbalance might play an important role in the pathogenesis of bronchial asthma through its effects on angiogenesis.     

Pan-neurotrophin receptor p75 contributes to neuronal hyperreactivity and airway inflammation in a murine model of experimental asthma

Bronchial asthma represents a severe chronic inflammatory disease with increasing prevalence. The pathogenesis is characterized by complex neuroimmune dysregulation. Although the immunopathogenesis of the disease has been extensively studied, the nature of neuronal dysfunction still remains poorly understood. Recent data indicate that neurotrophins contribute to airway inflammation, broncho-obstruction and airway hyperresponsiveness. Using an established murine model of allergic bronchial asthma, the contribution of the pan-neurotrophin receptor p75(NTR) was defined. This receptor is expressed both in normal and asthmatic lungs and airways. Analysis of p75(NTR-/-) mice, as well as in vivo blocking of p75(NTR), revealed that airway inflammation is to a large extent dependent upon functional receptor expression. Furthermore, neuronal hyperreactivity depends entirely on this receptor. Based on these data, a novel molecular pathway in the neuroimmune pathogenesis of bronchial asthma could be defined.     

Blockade of cysteinyl leukotriene‐1 receptors suppresses airway remodelling in mice overexpressing GATA‐3

Background We demonstrated previously that GATA‐3 overexpression markedly enhanced allergen‐induced airway inflammation and airway remodelling, including subepithelial fibrosis, and smooth muscle cell hyperplasia, in transgenic mice. Objective Because cysteinyl leukotrienes (cysLTs) have been shown to be involved in such structural changes, the effects of a specific cysLT1 receptor antagonist, montelukast, were evaluated in a mouse model of chronic asthma. Methods GATA‐3‐overexpressing mice and wild‐type Balb/c mice were sensitized and repeatedly challenged by ovalbumin (OVA) or saline. The effects of montelukast on the development of airway remodelling were compared between the two mouse genotypes. Results CysLTs in the lung were increased after repeated allergen challenges, and significantly enhanced in GATA‐3‐overexpressing mice. The enhanced cysLT levels were accompanied by the development of eosinophilia, smooth muscle cell hyperplasia, and increased stromal cell‐derived factor‐1 gene expression with a small increase in pro‐collagen gene expression in OVA‐challenged GATA‐3‐overexpressing mice, but not in wild‐type mice. Montelukast significantly decreased lung cysLT levels and inhibited the GATA‐3‐overexpression‐related airway remodelling, potently preventing smooth muscle cell hyperplasia, but partially suppressed the increased pro‐collagen gene expression and eosinophilic inflammation. Increases in the levels of IL‐4, IL‐5, IL‐13, and eotaxin in bronchial lavage and TGF‐β gene expression in the lungs were induced by OVA in both mouse genotypes. Montelukast treatment also significantly reduced these levels to the levels seen after saline challenges in GATA‐3‐overexpressing mice. Conclusion Montelukast efficaciously prevented airway inflammation and remodelling in a GATA‐3‐overexpression antigen challenge mouse model by decreasing the cysLT‐driven Th2 cytokine cycle of amplification of airway pathologies. Cite this as: T. Kiwamoto, Y. Ishii, Y. Morishima, K. Yoh, N. Kikuchi, N. Haraguchi, H. Masuko, M. Kawaguchi, A. Nomura, T. Sakamoto, S. Takahashi and N. Hizawa, Clinical & Experimental Allergy, 2011 (41) 116–128.     

A case–control study of bronchial asthma associated with ulcerative colitis: role of airway microvascular permeability

BACKGROUND: Recent attention has been devoted to the respiratory manifestations that may be associated with diseases of distant organs. The most prevalent and distinctive pattern of respiratory involvement in ulcerative colitis (UC) is airway inflammation. OBJECTIVE: This study was designed to examine the contribution of airway microvascular permeability to the pathophysiological association of asthma with UC. METHODS: Sputum induction and methacholine provocation test were performed in 27 asthmatic patients (15 without UC and 12 with UC), nine patients with UC and 15 normal controls. Inflammatory indexes, vascular endothelial growth factor (VEGF) levels in induced sputum, airway vascular permeability index and exhaled nitric oxide (NO) levels were examined in all subjects. RESULTS: The percentage of eosinophils and concentration of eosinophil cationic protein in induced sputum were similar in all four groups. Though exhaled NO levels were significantly higher in asthmatics with or without UC than in normal controls or UC patients, these levels were comparable in asthmatics with and without UC. VEGF levels in induced sputum and airway vascular permeability index were significantly higher in asthmatics without UC (VEGF: 1920 (990) pg/mL; airway vascular permeability index: 0.018 (0.008)) and asthmatics with UC (6570 (1000) pg/mL; 0.040 (0.006)) than in normal controls (950 (700) pg/mL; 0.009 (0.003)), whose levels were comparable to those of UC patients (900 (600) pg/mL; 0.011 (0.003)). In particular, these parameters were markedly increased in asthmatics with UC than in asthmatics without UC. VEGF level was significantly correlated with airway vascular permeability index in asthmatics with UC. Moreover, VEGF level and airway vascular permeability index was inversely correlated with degree of airway obstruction and airway hyper-reactivity to methacholine in these asthmatics. CONCLUSION: Airway microvascular hyper-permeability induced by VEGF may have a profound effect on airway function and can explain the heightened airway hyper-responsiveness characteristic of asthma associated with UC.     

Gene expression of vascular endothelial growth factor and its receptors and angiogenesis in bronchial asthma

BACKGROUND: Angiogenesis is a feature of airway remodeling in bronchial asthma. The mechanism responsible for this angiogenesis is unknown. Vascular endothelial growth factor (VEGF) is a potent inducer of endothelial cells, which may contribute to chronic inflammation and angiogenesis. OBJECTIVE: We sought to investigate the molecular mechanisms underlying increased vascularity, and we examined the mRNA expression of VEGF and its receptors (flt-1 and flk-1) within bronchial biopsy specimens from asthmatic patients and normal control subjects. METHODS: Endobronchial biopsy specimens were examined immunocytochemically by staining with anti-type IV collagen mAb to evaluate vessel density by using computer-assisted image analysis. Specimens were also analyzed for the presence of the mRNAs of VEGF and its receptors with in situ hybridization. RESULTS: The extent of airway vascularity was increased in asthmatic subjects compared with that in control subjects (P <.01). Asthmatic subjects exhibited a greater expression of VEGF, flt-1, and flk-1 mRNA(+) cells in the airway mucosa compared with that in control subjects (P <.001 for each comparison). The degree of vascularity was associated with the number of VEGF, flt-1, and flk-1 mRNA(+) cells. Numbers of cells expressing VEGF mRNA inversely correlated with airway caliber (r = -0.83, P <.01) and airway hyperresponsiveness (r = -0.97, P <.001). Colocalization studies showed that macrophages, eosinophils, and CD34(+) cells were the major sources of VEGF; CD34(+) cells, macrophages, and T cells expressed both flt-1 and flk-1. CONCLUSION: These findings provide evidence that VEGF may play an important role in angiogenesis and subsequent airway remodeling in bronchial asthma.     

全氟辛酸暴露对哮喘小鼠炎症介质IL-4和IFN-γ及糖皮质激素受体的影响

目的:研究全氟辛酸(PFOA)对哮喘小鼠气道炎症、外周血炎症介质白细胞介素4(IL-4)和干扰素γ(IFN-γ)以及肺组织糖皮质激素受体(GR)表达的影响及其可能机制。方法:30只BALB/c小鼠随机分为正常对照(C)组、哮喘模型(A)组、哮喘+PFOA低剂量(AP10)组、哮喘+PFOA中剂量(AP50)组和哮喘+PFOA高剂量(AP100)组,根据不同分组分别制作哮喘模型及PFOA暴露模型。留取肺组织标本后进行HE染色,透射电镜标本制作,ELISA法检测外周血IL-4及IFN-γ水平,并以Western blot及免疫组织化学染色法检测肺组织GR的蛋白表达。结果:肺组织病理切片HE染色结果显示,与正常小鼠相比,哮喘小鼠气道及血管周围可见明显的炎症细胞浸润及黏液分泌,AP各组表现更为明显。透射电镜结果显示,哮喘小鼠肺组织超微结构破坏明显。ELISA检测血清炎症因子结果表明,与C组相比,A组及AP各组外周血炎症因子IL-4升高,IFN-γ明显降低(P0.05);与A组相比,AP10组及AP50组无明显差异,而AP100组外周血炎症因子IL-4升高,IFN-γ明显降低(P0.05)。Western blot结果显示哮喘小鼠肺组织GR表达较正常小鼠降低(P0.05),而A组与AP各组之间无显著差异。免疫组化结果提示GR蛋白主要表达于小鼠肺组织支气管柱状上皮细胞、气道平滑肌细胞及血管平滑肌细胞胞浆。结论:哮喘小鼠气道PFOA急性暴露可通过诱导Th2型免疫反应加剧肺部炎症,促进气道及血管周围的炎症细胞浸润并破坏肺组织超微结构,且与剂量相关。