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J Agbetile,
A Fairs,
D Desai,
B Hargadon,
M Bourne,
K Mutalithas,
R Edwards,
J P Morley,
W R Monteiro,
N S Kulkarni,
R H Green,
I D Pavord, P Bradding,
C E Brightling,
A J Wardlaw,
C H Pashley
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ABSTRACT: Fungal sensitization is common in severe asthma, but the clinical relevance of this and the relationship with airway colonization by fungi remain unclear. The range of fungi that may colonize the airways in asthma is unknown.
To provide a comprehensive analysis on the range of filamentous fungi isolated in sputum from people with asthma and report the relationship with their clinico-immunological features of their disease.
We recruited 126 subjects with a diagnosis of asthma, 94% with moderate-severe disease, and 18 healthy volunteers. At a single stable visit, subjects underwent spirometry; sputum fungal culture and a sputum cell differential count; skin prick testing to both common aeroallergens and an extended fungal panel; specific IgE to Aspergillus fumigatus. Fungi were identified by morphology and species identity was confirmed by sequencing. Four patients had allergic bronchopulmonary aspergillosis.
Forty-eight percent of asthma subjects were IgE-sensitized to one fungal allergen and 22% to ≥ 2. Twenty-seven different taxa of filamentous fungi were isolated from 54% of their sputa, more than one species being detected in 17%. This compared with 3 (17%) healthy controls culturing any fungus (P < 0.01). Aspergillus species were most frequently cultured in isolation followed by Penicillium species. Post-bronchodilator FEV (1) (% predicted) in the subjects with asthma was 71(± 25) in those with a positive fungal culture vs. 83 (± 25) in those culture-negative, (P < 0.01).
Numerous thermotolerant fungi other than A. fumigatus can be cultured from sputum of people with moderate-to-severe asthma; a positive culture is associated with an impaired post-bronchodilator FEV (1) , which might be partly responsible for the development of fixed airflow obstruction in asthma. Sensitization to these fungi is also common.
Clinical & Experimental Allergy 05/2012; 42(5):782-91. · 5.03 Impact Factor
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ABSTRACT: Mast cells, traditionally regarded as effector cells of the immune system, have more recently been demonstrated to be key figures in initiating, developing and sustaining complex pathophysiological processes underlying asthma and other allergic diseases. Asthma is characterised by airway inflammation alongside a disturbance to airway physiology manifesting as variable airflow obstruction and airway hyper-responsiveness (AHR). Evidence has emerged that mast cells influence airway function by forming close intercellular relationships with different structural components of the airway wall. In asthma, mast cells are seen to localise to the airway epithelium, to mucous glands and to the airway smooth muscle (ASM). It is mast cell-ASM interaction that is most fundamental to the asthma phenotype and many mast cell mediators have been demonstrated to have important effects on ASM function. In asthma, alongside the inflammatory and physiological changes, structural changes occur to the airway wall in the form of denudation of the epithelium, goblet cell and mucous gland hyperplasia, subepithelial fibrosis, abnormal extracellular matrix (ECM) deposition, vascular proliferation and increased ASM mass. There are many ways in which mast cells can contribute to these structural changes through direct cell to cell communication and more indirectly through mediator release. Mast cells exhibit an array of diverse functions and roles and are fundamental to our current understanding of asthma pathogenesis including severe asthma. Novel targeting of mast cells and their mediators therefore should offer significant therapeutic potential in the treatment of asthma.
Current pharmaceutical design 03/2011; 17(7):685-98. · 4.41 Impact Factor
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D Desai,
C J Newby,
P Haldar,
S Shah,
S Gupta,
M Bafadhel,
A Singapuri,
S Siddiqui,
J Woods,
A Herath,
I K Anderson, P Bradding,
R H Green,
A J Wardlaw,
I D Pavord,
R D May,
C E Brightling
Thorax 01/2011; 66(supplement 4):A56. · 6.84 Impact Factor
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R D May,
P Haldar,
S Shah,
S Gupta,
M Bafadhel,
J Woods,
A Herath,
I K Anderson,
R Green,
A Wardlaw,
D Desai, P Bradding,
I D Pavord,
C E Brightling
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ABSTRACT: Corresponding author's email: dhananjaydesai@hotmail.com Understanding the phenotypic heterogeneity of asthma is likely to shed light upon its immunopathogenesis. Introduction: To examine patterns of cytokine expression in different clinical phenotypes of severe asthma. Rationale: We performed k-means clustering on demographic, lung function, allergen testing and sputum induction data of 164 patients Methods: attending our Difficult Asthma Clinic. Sputum supernatant was analysed for 23 mediators using Meso Scale Discovery platform. The data obtained was reduced using principal component analysis. Factorial patterns of mediator expression were compared across clinical phenotypes. Using unbiased data reduction tools we were able to identify severe asthma clinical sub-phenotypes. Cluster analysis identified 4 Results: clinical phenotypes: A. Discordant high symptoms/non-eosinophilic, obese, normal FEV ‚; B. Late onset, concordant 1 symptoms/eosinophilia, normal FEV ‚; C. Early onset, discordant low symptoms/eosinophilia, low FEV ; D. Early onset, concordant 1 1 symptoms/eosinophilia, normal FEV ‚. The individual mediators did not differ significantly across clusters. Four cytokine factors were 1 identified 1. IL6R, IL8, TNFRI; 2. CXCL11, IL6, CCL5; 3. CCL26, IL13, IL5 and; 4. IL10, IL17, IL4. The factors loaded onto the clusters as shown in the figure below where data are shown as the mean factor loading ± SEM. Graph showing cytokine factor loading across clinical clusters Clinical phenotypes of severe asthma differ in their patterns of mediator expression. The biological basis of clinical disease Conclusion: expression requires further study. Clinical clustering is proving a useful tool at identifying underlying biology. This abstract is funded by: None
American Journal of Respiratory and Critical Care Medicine 01/2011; 183:A3179. · 11.08 Impact Factor
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Thorax 04/2010; 65(4):370. · 6.84 Impact Factor
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ABSTRACT: Airway smooth muscle (ASM) hyperplasia and mast cell localization within the ASM bundle are important features of asthma. The cause of this increased ASM mass is uncertain and whether it is a consequence of ASM-mast cell interactions is unknown.
We sought to investigate ASM proliferation and survival in asthma and the effects of co-culture with mast cells.
Primary ASM cultures were derived from 11 subjects with asthma and 12 non-asthmatic controls. ASM cells were cultured for up to 10 days in the presence or absence of serum either alone or in co-culture with the human mast cell line-1, unstimulated human lung mast cells (HLMC) or IgE/anti-IgE-activated HLMC. Proliferation was assessed by cell counts, CFSE assay and thymidine incorporation. Apoptosis and necrosis were analysed by Annexin V/propidium iodide staining using flow cytometry and by assessment of nuclear morphology using immunofluorescence. Mast cell activation was confirmed by the measurement of histamine release.
Using a number of techniques, we found that ASM proliferation and survival was not significantly different between cells derived from subjects with or without asthma. Co-culture with mast cells did not affect the rate of proliferation or survival of ASM cells.
Our findings do not support a role for increased airway smooth proliferation and survival as the major mechanism driving ASM hyperplasia in asthma.
Clinical & Experimental Allergy 02/2010; 40(2):279-88. · 5.03 Impact Factor
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ABSTRACT: Chemokine receptors play an important role in cell migration and wound repair. In asthma, CCR3 and 7 are expressed by airway smooth muscle (ASM) and CCR7 has been implicated in the development of ASM hyperplasia. The expression profile of other chemokine receptors by ASM and their function needs to be further explored.
We sought to investigate ASM chemokine receptor expression and function in asthma.
ASM cells were derived from 17 subjects with asthma and 36 non-asthmatic controls. ASM chemokine receptor expression was assessed by flow cytometry and immunofluorescence. The function of chemokine receptors expressed by more than 10% of ASM cells was investigated by intracellular calcium measurements, chemotaxis, wound healing, proliferation and survival assays.
In addition to CCR3 and 7, CXCR1, 3 and 4 were highly expressed by ASM. These CXC chemokine receptors were functional with an increase in intracellular calcium following ligand activation and promotion of wound healing [CXCL10 (100 ng/mL) 34 +/- 2 cells/high-powered field (hpf) vs. control 29 +/- 1; P=0.03; n=8]. Spontaneous wound healing was inhibited by CXCR3 neutralizing antibody (mean difference 7 +/- 3 cells/hpf; P=0.03; n=3). CXC chemokine receptor activation did not modulate ASM chemotaxis, proliferation or survival. No differences in chemokine receptor expression or function were observed between ASM cells derived from asthmatic or non-asthmatic donors.
Our findings suggest that the chemokine receptors CXCR1, 3 and 4 modulate some aspects of ASM function but their importance in asthma is uncertain.
Clinical & Experimental Allergy 10/2009; 39(11):1684-92. · 5.03 Impact Factor
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ABSTRACT: Airway smooth muscle hyperplasia is a feature of asthma, and increases with disease severity. CCR3-mediated recruitment of airway smooth muscle progenitors towards the airway smooth muscle bundle has been proposed as one possible mechanism involved in airway smooth muscle hyperplasia. Mast cells are microlocalized to the airway smooth muscle bundle and whether mast cells influence CCR3-mediated migration is uncertain.
We examined the expression of CCR3 by primary cultures of airway smooth muscle cells from asthmatics and nonasthmatics. CCR3 function was examined using intracellular calcium measurements, chemotaxis, wound healing, cell proliferation and survival assays. We investigated the recovery and function of both recombinant and airway smooth muscle-derived CCL11 (eotaxin) after co-culture with beta-tryptase and human lung mast cells.
Airway smooth muscle expressed CCR3. Airway smooth muscle CCR3 activation by CCL11 mediated intracellular calcium elevation, concentration-dependent migration and wound healing, but had no effect on proliferation or survival. Co-culture with beta-tryptase or mast cells degraded recombinant and airway smooth muscle-derived CCL11, and beta-tryptase inhibited CCL11-mediated airway smooth muscle migration.
CCL11 mediates airway smooth muscle migration. However co-culture with beta-tryptase or mast cells degraded recombinant and airway smooth muscle-derived CCL11 and inhibited CCL11-mediated airway smooth muscle migration. Therefore these findings cast doubt on the importance of the CCL11/CCR3 axis in the development of airway smooth muscle hyperplasia in asthma.
Allergy 10/2008; 63(9):1148-55. · 6.27 Impact Factor
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ABSTRACT: The Th2 cytokine interleukin-13 (IL-13) has been implicated in the pathogenesis of chronic obstructive pulmonary disease (COPD). We sought to examine IL-13 expression in COPD subjects in induced sputum and bronchus specimens. We hypothesized that inflammatory cells expressing IL-13 localize to the airway smooth muscle bundle and bronchial glands.
Interleukin-13 was measured in sputum samples from subjects with COPD (n = 34) across a range of severity (Global initiative for chronic Obstructive Lung Disease 2-4) and controls (n = 14) using ELISA. IL-13+ cells and inflammatory cells were enumerated within surgically resected proximal airway using immunohistochemical techniques from subjects with COPD (n = 10), smoking (n = 10) and nonsmoking controls (n = 8).
Sputum IL-13 was measurable in only 6/34 subjects with COPD and was not found in the smoking or nonsmoking control subjects. In subjects with COPD and controls there was a paucity of IL-13+ cells. The distribution of inflammatory cells within different airway compartments was similar in COPD and controls except for an increase in CD3(+) lymphocytes within bronchial glands in COPD (P = 0.04).
Our findings do not support a role for IL-13 in COPD. However, the tissue localization of inflammatory cells to airway compartments, particularly the increase of T cells in glands in COPD may be important in disease.
Allergy 10/2008; 63(9):1239-43. · 6.27 Impact Factor
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ABSTRACT: Asthma is a chronic and sometimes fatal disease, which affects people of all ages throughout the world. Important hallmarks of asthma are airway inflammation and remodelling, with associated bronchial hyperresponsiveness and variable airflow obstruction. These features are orchestrated by cells of both the innate (eosinophils, neutrophils and mast cells) and the adaptive (T(H)2 T cells) immune system, in concert with structural airway cells. Chemokines are important for the recruitment of both immune and structural cells to the lung, and also for their microlocalisation within the lung tissue. Specific blockade of the responses elicited by chemokines and chemokine receptors responsible for the pathological migration of airway cells could therefore be of great therapeutic interest for the treatment of asthma.
British Journal of Pharmacology 08/2007; 151(6):725-36. · 4.41 Impact Factor
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ABSTRACT: Mast cell recruitment and activation are critical for the initiation and progression of inflammation and fibrosis. Mast cells infiltrate specific structures in many diseased tissues such as the airway smooth muscle (ASM) in asthma. This microlocalisation of mast cells is likely to be key to disease pathogenesis. Human lung mast cells (HLMC) express the Ca2+ activated K+ channel K(Ca)3.1 which modulates mediator release, and is proposed to facilitate the retraction of the cell body during migration of several cell types. A study was undertaken to test the hypothesis that blockade of K(Ca)3.1 would attenuate HLMC proliferation and migration.
HLMC were isolated and purified from lung material resected for bronchial carcinoma. HLMC proliferation was assessed by cell counts at various time points following drug exposure. HLMC chemotaxis was assayed using standard Transwell chambers (8 microm pore size). Ion currents were measured using the single cell patch clamp technique.
K(Ca)3.1 blockade with triarylmethane-34 (TRAM-34) did not inhibit HLMC proliferation and clotrimazole had cytotoxic effects. In contrast, HLMC migration towards the chemokine CXCL10, the chemoattractant stem cell factor, and the supernatants from tumour necrosis factor alpha stimulated asthmatic ASM was markedly inhibited with both the non-selective K(Ca)3.1 blocker charybdotoxin and the highly specific K(Ca)3.1 blocker TRAM-34 in a dose dependent manner. Although K(Ca)3.1 blockade inhibits HLMC migration, K(Ca)3.1 is not opened by the chemotactic stimulus, suggesting that it must be involved downstream of the initial receptor-ligand interactions.
Since modulation of K(Ca)3.1 can inhibit HLMC chemotaxis to diverse chemoattractants, the use of K(Ca)3.1 blockers such as TRAM-34 could provide new therapeutic strategies for mast cell mediated diseases such as asthma.
Thorax 11/2006; 61(10):880-5. · 6.84 Impact Factor
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ABSTRACT: The Th2 cytokine interleukin (IL)-13 is implicated in the development of various allergic diseases including asthma. The IL-13 receptor, IL-13Ralpha1, is expressed on most leukocytes, except T-cells. Evidence to support IL-13Ralpha1 expression on mast cells is limited.
We investigated: (i) IL-13Ralpha1 expression by human lung mast cells (HLMC); (ii) the number of IL-13Ralpha1+ bronchial submucosal mast cells in subjects with asthma and normal controls and (iii) the effect of IL-13 priming on HLMC expression of high-affinity IgE receptor (FcepsilonRI), stem cell factor receptor (CD117), histamine release, proliferation, and survival.
Human lung mast cell expressed IL-13Ralpha1 mRNA. IL-13Ralpha1 was highly expressed on the surface HLMC (82+/-9%). Bronchial submucosal mast cell IL-13Ralpha1 expression was higher in asthmatics (86+/-2%) than normal controls (78+/-2%; P=0.015). IL-13 priming for 30 min did not increase HLMC histamine release, in the presence or absence of SCF or in response to IgE/anti-IgE activation. IL-13 priming for 5 days upregulated HLMC FcepsilonRI expression (22% increase in fluorescent intensity; P=0.003), increased histamine release following IgE/anti-IgE activation by 56% (P=0.03) and increased proliferation by 50% (P=0.003) without affecting cell survival or CD117 expression. The IL-13 specific neutralizing antibody CAT-354 inhibited all IL-13 mediated effects.
Human lung mast cell express IL-13Ralpha1 and activation by IL-13 for 5 days increased FcepsilonRI expression and proliferation. Histamine release was not affected by short-term priming with IL-13, but was upregulated by priming for 5 days suggesting that this effect was mediated by the increased FcepsilonRI expression. These data support the view that targeting IL-13 may be beneficial in the treatment of asthma.
Allergy 10/2006; 61(9):1047-53. · 6.27 Impact Factor
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ABSTRACT: Mast cell microlocalisation within the airway smooth muscle (ASM) bundle is an important determinant of the asthmatic phenotype. We hypothesised that mast cells migrate towards ASM in response to ASM derived chemokines.
Primary ASM cultures from subjects with and without asthma were stimulated with interleukin (IL)-1beta, IL-4, and IL-13 alone and in combination. Mast cell chemotaxis towards these ASM supernatants was investigated, and the chemotaxins mediating migration by using specific blocking antibodies for stem cell factor (SCF) and the chemokine receptors CCR3, CXCR1, 3 and 4 as well as the Gi inhibitor pertussis toxin and the tyrosine kinase inhibitor genistein were defined. The concentrations of CCL11, CXCL8, CXCL10, TGF-beta, and SCF in the supernatants were measured and the effect of non-asthmatic ASM supernatants on the mast cell chemotactic activity of asthmatic ASM was examined.
Human lung mast cells and HMC-1 cells migrated towards Th2 stimulated ASM from asthmatics but not non-asthmatics. Mast cell migration was mediated through the combined activation of CCR3 and CXCR1. CCL11 and CXCL8 expression by ASM increased markedly after stimulation, but was similar in those with and without asthma. ASM supernatants from non-asthmatics inhibited mast cell migration towards the asthmatic ASM supernatant.
Th2 stimulated ASM from asthmatics is chemotactic for mast cells. Non-asthmatic ASM releases a mediator or mediators that inhibit mast cell migration towards stimulated asthmatic ASM. Specifically targeting mast cell migration into the ASM bundle may provide a novel treatment for asthma.
Thorax 09/2006; 61(8):657-62. · 6.84 Impact Factor
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ABSTRACT: Background: The Th2 cytokine interleukin (IL)-13 is implicated in the development of various allergic diseases including asthma. The IL-13 receptor, IL-13R1, is expressed on most leukocytes, except T-cells. Evidence to support IL-13R1 expression on mast cells is limited.Methods: We investigated: (i) IL-13R1 expression by human lung mast cells (HLMC); (ii) the number of IL-13R1+ bronchial submucosal mast cells in subjects with asthma and normal controls and (iii) the effect of IL-13 priming on HLMC expression of high-affinity IgE receptor (FcɛRI), stem cell factor receptor (CD117), histamine release, proliferation, and survival.Results: Human lung mast cell expressed IL-13R1 mRNA. IL-13R1 was highly expressed on the surface HLMC (82 ± 9%). Bronchial submucosal mast cell IL-13R1 expression was higher in asthmatics (86 ± 2%) than normal controls (78 ± 2%; P = 0.015). IL-13 priming for 30 min did not increase HLMC histamine release, in the presence or absence of SCF or in response to IgE/anti-IgE activation. IL-13 priming for 5 days upregulated HLMC FcɛRI expression (22% increase in fluorescent intensity; P = 0.003), increased histamine release following IgE/anti-IgE activation by 56% (P = 0.03) and increased proliferation by 50% (P = 0.003) without affecting cell survival or CD117 expression. The IL-13 specific neutralizing antibody CAT-354 inhibited all IL-13 mediated effects.Conclusion: Human lung mast cell express IL-13R1 and activation by IL-13 for 5 days increased FcɛRI expression and proliferation. Histamine release was not affected by short-term priming with IL-13, but was upregulated by priming for 5 days suggesting that this effect was mediated by the increased FcɛRI expression. These data support the view that targeting IL-13 may be beneficial in the treatment of asthma.
Allergy 08/2006; 61(9):1047 - 1053. · 6.27 Impact Factor
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ABSTRACT: Human lung mast cells (HLMC) lie in close proximity to the bronchial epithelium in asthma and adhere with high affinity to bronchial epithelial monolayers in vitro. We investigated the consequences of this adhesive interaction on HLMC activation in response to Fc epsilon RI cross-linking.
Human lung mast cells were cultured with the bronchial epithelial cell line BEAS-2B or plastic control for either 30 min or 16 h and then activated with anti-IgE. Histamine was measured by radioenzymatic assay.
After co-culture for 30 min, IgE-dependent histamine release from HLMC was identical on both BEAS-2B and plastic. After 16 h of co-culture, there was a marked decrease in constitutive and IgE-dependent histamine release from HLMC cultured on BEAS-2B compared with those cultured on plastic or fibronectin. In contrast, the Ca(2+)/ATPase inhibitor thapsigargin produced concentration-dependent histamine release that was significantly increased on BEAS-2B compared with plastic. IgE-dependent degranulation was not significantly affected by BEAS-2B-conditioned medium.
BEAS-2B bronchial epithelial cells attenuate IgE-dependent but not thapsigargin-induced histamine release from HLMC. The differential effect with anti-IgE compared with thapsigargin suggests that the mechanism includes interference with the proximal Fc epsilon RI signalling pathway.
Allergy 06/2006; 61(5):569-75. · 6.27 Impact Factor
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ABSTRACT: The mechanism of chronic mast cell activation in asthma is unclear. Monomeric immunoglobulin (Ig)E in the absence of allergen induces mediator release from rodent mast cells, indicating a possible role for IgE in the continued activation of mast cells within the asthmatic bronchial mucosa. In this study it was investigated whether monomeric IgE induces Ca2+ influx and mediator release from human lung mast cells (HLMC). Purified HLMC were cultured for 4 weeks and then exposed to monomeric human myeloma IgE. Ratiometric Ca2+ imaging was performed on single fura-2-loaded cells. Histamine release was measured by radioenzymatic assay; leukotriene C4 (LTC4) and interleukin (IL)-8 were measured by ELISA. At concentrations experienced in vivo, monomeric IgE induced dose-dependent histamine release, LTC4 production and IL-8 synthesis. This was associated with a rise in cytosolic free Ca2+. Enhanced histamine release was still evident 1 week after initial exposure to IgE suggesting that continued exposure maintains enhanced secretion. Monomeric immunoglobulin E alone activates cultured human lung mast cells initiating Ca2+ influx, degranulation, arachidonic acid metabolism and cytokine synthesis. These findings support the hypothesis that immunoglobulin E loading of mast cells within the asthmatic airway contributes to the disordered airway physiology of this disease.
European Respiratory Journal 06/2005; 25(5):858-63. · 5.89 Impact Factor
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C E Brightling,
S McKenna,
B Hargadon,
S Birring,
R Green,
R Siva,
M Berry,
D Parker,
W Monteiro,
I D Pavord, P Bradding
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ABSTRACT: An association between the sputum eosinophil count and the response to a 2 week course of prednisolone has previously been reported in patients with chronic obstructive pulmonary disease (COPD). Whether the response to inhaled corticosteroids is related to the presence of eosinophilic inflammation is unclear.
A randomised, double blind, crossover trial of placebo and mometasone furoate (800 microg/day), each given for 6 weeks with a 4 week washout period, was performed in subjects with COPD treated with bronchodilator therapy only. Spirometric tests, symptom scores, chronic respiratory disease questionnaire (CRQ), and induced sputum were performed before and after each treatment phase.
Ninety five patients were recruited of which 60 were randomised. Overall there were no treatment associated changes in forced expiratory volume in 1 second (FEV(1)), total CRQ, or sputum characteristics. After stratification into tertiles by baseline eosinophil count, the net improvement in post-bronchodilator FEV(1) increased with mometasone compared with placebo progressively from the least to the most eosinophilic tertile. The mean change in post-bronchodilator FEV(1) with mometasone compared with placebo in the highest tertile was 0.11 l (95% CI 0.03 to 0.19). This improvement was not associated with a fall in the sputum eosinophil count.
An increased sputum eosinophil count is related to an improvement in post-bronchodilator FEV(1) following treatment with inhaled mometasone in COPD, but the improvement is not associated with a reduction in the sputum eosinophil count.
Thorax 04/2005; 60(3):193-8. · 6.84 Impact Factor
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ABSTRACT: Cough is a common symptom in idiopathic pulmonary fibrosis that is difficult to treat and has a major impact on quality of life. We tested the hypothesis that the cough and increased cough reflex sensitivity seen in patients with idiopathic pulmonary fibrosis may be due to airway inflammation in a prospective, cross-sectional study.
We measured the induced sputum inflammatory cell profile and cell-free supernatant inflammatory mediator concentrations in 15 patients with idiopathic pulmonary fibrosis, 17 healthy controls and 15 patients with chronic obstructive pulmonary disease.
Both the geometric mean sputum differential eosinophil cell count and median eosinophilic-cationic-protein concentration were significantly higher in patients with idiopathic pulmonary fibrosis than controls (2.1% vs 0.3%; p <0.001 and 1.1 mg/ml versus 0.2 mg/ml; p=0.03 respectively). There were no significant differences in sputum eosinophil counts and eosinophilic-cationic-protein concentrations between patients with idiopathic pulmonary fibrosis and chronic obstructive pulmonary disease. Sputum leukotriene-B4 concentrations were significantly lower in patients with idiopathic pulmonary fibrosis (p=0.03) and chronic obstructive pulmonary disease (p=0.008) compared to controls.
Idiopathic pulmonary fibrosis is characterised by the presence of active eosinophilic airway inflammation raising the possibility that airway inflammation may contribute to symptoms such as cough.
Inflammation Research 03/2005; 54(2):51-6. · 2.11 Impact Factor
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ABSTRACT: Mast cells contribute to the pathogenesis of asthma and allergy through the release of a plethora of pro-inflammatory mediators and cytokines. Their study is hampered by the difficult access to human tissue samples. Human mast cells have been cultured from CD34+ progenitors in the bone marrow of normal volunteers following iliac crest bone marrow biopsy but this is invasive. Hip bone marrow could provide a more convenient less invasive source of mast cell progenitors.
To characterize mast cells cultured from human bone marrow obtained at routine hip surgery.
Mononuclear cells were isolated from the bone marrow reamings of patients undergoing routine hip replacement surgery and were cultured with recombinant stem cell factor (SCF), IL-6 and IL-10. Cell surface markers were examined using flow cytometry, protease expression monitored using immunohistochemistry, histamine measured by radioenzymic assay, Ca2+ responses analysed using ratiometric Ca2+ imaging, and ion currents recorded via the patch-clamp technique.
Mast cells were absent at baseline, but accounted for 65 +/- 7% of cells after 8-12 weeks of culture, equating to a mean 0.6 +/- 0.14 x 10(6) mast cells per culture. Fifty-three percent of tryptase+ cells also contained chymase. The remaining cells comprised a population of large CD1a+ HLA-DR+ and Fc epsilon RI alpha+ cells, most likely dendritic cells. All mast cells expressed CD117 and the high-affinity IgE receptor alpha-chain (Fc epsilon RI alpha) constitutively, and developed a Ca2+ response following IgE-dependent activation. These cells exhibited 7.8 +/- 2.9% net IgE-dependent histamine release, and demonstrated a similar ion channel profile to human lung mast cells. In particular, the intermediate conductance Ca(2+)-activated K+ channel opened following IgE-dependent activation.
Mast cells grown from human hip marrow provide a rich non-invasive source of functionally mature mast cells. In addition, this culture system may be useful for the generation of Fc epsilon RI alpha+ dendritic cells.
Clinical & Experimental Allergy 03/2005; 35(2):226-33. · 5.03 Impact Factor
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ABSTRACT: Background Mast cells contribute to the pathogenesis of asthma and allergy through the release of a plethora of pro-inflammatory mediators and cytokines. Their study is hampered by the difficult access to human tissue samples. Human mast cells have been cultured from CD34+ progenitors in the bone marrow of normal volunteers following iliac crest bone marrow biopsy but this is invasive. Hip bone marrow could provide a more convenient less invasive source of mast cell progenitors.Objective To characterize mast cells cultured from human bone marrow obtained at routine hip surgery.Methods Mononuclear cells were isolated from the bone marrow reamings of patients undergoing routine hip replacement surgery and were cultured with recombinant stem cell factor (SCF), IL-6 and IL-10. Cell surface markers were examined using flow cytometry, protease expression monitored using immunohistochemistry, histamine measured by radioenzymic assay, Ca2+ responses analysed using ratiometric Ca2+ imaging, and ion currents recorded via the patch-clamp technique.Results Mast cells were absent at baseline, but accounted for 65±7% of cells after 8–12 weeks of culture, equating to a mean 0.6±0.14 × 106 mast cells per culture. Fifty-three percent of tryptase+ cells also contained chymase. The remaining cells comprised a population of large CD1a+ HLA-DR+ and FcɛRIα+ cells, most likely dendritic cells. All mast cells expressed CD117 and the high-affinity IgE receptor α-chain (FcɛRIα) constitutively, and developed a Ca2+ response following IgE-dependent activation. These cells exhibited 7.8±2.9% net IgE-dependent histamine release, and demonstrated a similar ion channel profile to human lung mast cells. In particular, the intermediate conductance Ca2+-activated K+ channel opened following IgE-dependent activation.Conclusions Mast cells grown from human hip marrow provide a rich non-invasive source of functionally mature mast cells. In addition, this culture system may be useful for the generation of FcɛRIα+ dendritic cells.
Clinical & Experimental Allergy 02/2005; 35(2):226 - 233. · 5.03 Impact Factor
