rie Lau

Double-stranded RNA induces disproportionate expression of thymic stromal lymphopoietin versus interferon-β in bronchial epithelial cells from donors with asthma

Background Thymic stromal lymphopoietin (TSLP) is an epithelial cell-derived cytokine that strongly activates dendritic cells and can initiate allergic inflammation. Since exposure to rhinovirus or double-stranded (ds) RNA (a surrogate of viral infection) induces TSLP expression in bronchial epithelial cells (BECs), this cytokine may link innate antiviral responses and the type 2 adaptive immune response. Objective As BECs from donors with asthma have a deficient interferon (IFN) response to rhinovirus infection, a study was undertaken to test the hypothesis that their antiviral response shows a bias towards TSLP production. Methods Primary BECs were grown from subjects with asthma and healthy volunteers. After exposure to dsRNA, interleukin (IL)-8, IFNβ and TSLP mRNA and protein expression were measured by RT-qPCR and ELISA, respectively. Results dsRNA dose-dependently increased IL-8 expression in BECs with no significant difference between the groups. However, BECs from subjects with asthma expressed less IFNβ and more TSLP mRNA and protein in response to dsRNA than BECs from those without asthma (median (IQR) 57 (38–82) pg/ml vs 106 (57–214) pg/ml for IFNβ (p<0.05) and 114 (86–143) pg/ml vs 65 (32–119) pg/ml for TSLP (p<0.05) in response to 10 μg/ml dsRNA for 24 h). Induction of TSLP mRNA by dsRNA was blocked by Toll-like receptor 3 or protein kinase inhibitors or by preventing de novo protein synthesis, but not by neutralisation of type I IFN receptors. Conclusion BECs from subjects with asthma are biased towards higher TSLP and lower IFNβ production in response to dsRNA, suggesting that viral infection in asthma may lead to an altered mediator profile that biases towards a Th2 immune response.

Potentially pathogenic airway bacteria and neutrophilic inflammation in treatment resistant severe asthma.

Background Molecular microbiological analysis of airway samples in asthma has demonstrated an altered microbiome in comparison to healthy controls. Such changes may have relevance to treatment-resistant severe asthma, particularly those with neutrophilic airway inflammation, as bacteria might be anticipated to activate the innate immune response, a process that is poorly steroid responsive. An understanding of the relationship between airway bacterial presence and dominance in severe asthma may help direct alternative treatment approaches. Objective We aimed to use a culture independent analysis strategy to describe the presence, dominance and abundance of bacterial taxa in induced sputum from treatment resistant severe asthmatics and correlate findings with clinical characteristics and airway inflammatory markers. Methods Induced sputum was obtained from 28 stable treatment-resistant severe asthmatics. The samples were divided for supernatant IL-8 measurement, cytospin preparation for differential cell count and Terminal Restriction Fragment Length Polymorphism (T-RFLP) profiling for bacterial community analysis. Results In 17/28 patients, the dominant species within the airway bacterial community was Moraxella catarrhalis or a member of the Haemophilus or Streptococcus genera. Colonisation with these species was associated with longer asthma disease duration (mean (SD) 31.8 years (16.7) vs 15.6 years (8.0), p = 0.008), worse post-bronchodilator percent predicted FEV1 (68.0% (24.0) vs 85.5% (19.7), p = 0.025) and higher sputum neutrophil differential cell counts (median (IQR) 80% (67–83) vs 43% (29–67), p = 0.001). Total abundance of these organisms significantly and positively correlated with sputum IL-8 concentration and neutrophil count. Conclusions Airway colonisation with potentially pathogenic micro-organisms in asthma is associated with more severe airways obstruction and neutrophilic airway inflammation. This altered colonisation may have a role in the development of an asthma phenotype that responds less well to current asthma therapies.

Inflammatory mediators in naturally occurring rhinitis

The mediators released during the allergic inflammatory reaction induce the clinical symptoms of the allergic disease and although there have been numerous studies investigating mediator release in allergen challenge models of allergic rhinitis very few have extended this approach to the study of natural disease.

Nebulised salbutamol administered during sputum induction improves bronchoprotection in patients with asthma

Background: Inhalation of hypertonic or even isotonic saline during sputum induction may cause bronchospasm in susceptible patients with asthma, despite premedication with 400 µg inhaled salbutamol delivered by pressurised metered dose inhaler (pMDI). The bronchoprotection afforded by additional inhaled salbutamol administered through the ultrasonic nebuliser during sputum induction was investigated. Methods: Twenty patients with moderate to severe asthma underwent sputum induction by inhaling saline 4.5% (or 0.9% if post-bronchodilation forced expiratory volume in 1 second (FEV1) <65% predicted) for 10 minutes according to two protocols given 1 week apart in random order. At visit A the patients received 400 µg salbutamol administered through a pMDI + spacer 20 minutes before induction while at visit B the premedication was supplemented by 1500 µg nebulised salbutamol inhaled throughout the induction procedure. Both the investigator and the patients were blind to the nebulised solution used. FEV1 was recorded during sputum induction at 1, 3, 5, and 10 minutes. Sputum cell counts and histamine, tryptase and albumin levels in the supernatants were determined. Results: The mean (SE) maximal reduction in FEV1 over the 10 minute period of sputum induction was 11.7 (2.8)% at visit A, which was significantly greater than at visit B (2.6 (1.2)%; mean difference 9% (95% CI 2.7 to 15.4), p<0.01). Total and differential sputum cell counts as well as albumin, tryptase, and histamine levels did not differ between the two visits. Conclusion: The addition of inhaled salbutamol through an ultrasonic nebuliser markedly improves bronchoprotection against saline induced bronchoconstriction in patients with moderate to severe asthma undergoing sputum induction without affecting cell counts and inflammatory markers.

Evidence of mast-cell activation in a subset of patients with eosinophilic chronic obstructive pulmonary disease

Although asthma has been viewed mainly as an eosinophilic disease, and chronic obstructive pulmonary disease (COPD) as a neutrophilic disease, recent studies have shown increased neutrophil counts in severe asthma and sputum eosinophilia in some COPD patients. In an attempt to further characterise these two syndromes according to pathology, the current authors have conducted a study of induced sputum in 15 subjects with COPD, 17 asthmatics, and 17 nonatopic healthy individuals. Sputum was analysed for cytology and levels of eosinophil cationic protein (ECP), albumin, tryptase and soluble intercellular adhesion molecule‐1. The COPD subjects differed from the asthmatics as they had higher sputum neutrophil and lower columnar epithelial cell counts, but there were no differences in any soluble marker studied. When compared to control subjects, both the asthmatic and COPD subjects had raised eosinophil counts and ECP levels. In a subset of COPD subjects with sputum eosinophilia (>3% of total cells), significantly increased levels of tryptase were detected. In conclusion, although chronic obstructive pulmonary disease is a more neutrophilic disease than asthma, the two diseases are difficult to distinguish on the basis of sputum levels of the soluble markers traditionally associated with asthma. However, a subset of patients with chronic obstructive pulmonary disease with airway eosinophilia and mast-cell activation might represent a distinct pathological phenotype.

Bronchial Eosinophilic Infiltration in Crohn's Disease in the Absence of Pulmonary Disease

Immunological and functional bronchopulmonary abnormalities may be present in up to two‐thirds of patients with Crohns disease. Having recently described a mild increase in methacholine airways responsiveness in these patients, we investigated whether this physiological abnormality is associated with bronchial inflammation since it has previously been described in asthma.

The novel use of the human nasal epithelial cell line RPMI 2650 as an in vitro model to study the influence of allergens and cytokines on transforming growth factor-beta gene expression and protein release.

Background The epithelial accumulation of mast cells is a feature of allergic rhinitis and this has been linked to the expression of the known mast cell chemoattractant transforming growth factor‐β (TGF‐β) at this site. Little is known concerning the regulation of TGF‐β gene expression or protein release by nasal epithelial cells. To address this we have utilized the RPMI 2650 human nasal epithelial cell line, which has some features that closely resemble normal nasal epithelium and has been reported to secrete a TGF‐β‐like molecule.

Comparative nasal effects of bradykinin and histamine: influence on nasal airways resistance and plasma protein exudation.

BACKGROUND--Bradykinin may contribute to the pathogenesis of allergic rhinitis. Like histamine, nasal challenge with bradykinin induces rhinorrhoea, nasal blockage, and plasma protein leakage. Their comparative nasal potencies have not, however, been fully elucidated. METHODS--Three double blind, randomised, placebo controlled and cross-over studies were undertaken to compare objectively the nasal effects of bradykinin, histamine, and vehicle. RESULTS--Both bradykinin and histamine produced dose dependent increases in nasal airways resistance (NAR). There was no significant difference in the effects of bradykinin and histamine on NAR at any dose level. On a molar basis, however, bradykinin was 6.98 times more potent than histamine in inducing a 50% increase in NAR. Nasal challenge with bradykinin and histamine also induced significant rhinorrhoea compared with vehicle. The amount of rhinorrhoea induced by histamine was significantly greater than that induced by bradykinin at any dose level. Bradykinin and histamine induced dose dependent nasal pain and nasal itch respectively. When administered as single doses both bradykinin (1.9 mumol) and histamine (1.9 mumol) induced significant rhinorrhoea compared with the vehicle. The volume of rhinorrhoea secretions induced by histamine was 29% greater than that induced by bradykinin. In contrast, although NAR was increased significantly more by histamine than by the vehicle, the effect of bradykinin on NAR was significantly greater than histamine and vehicle in both magnitude and duration of effect. The incremental effect of bradykinin on lavage albumin levels was also significantly greater than both histamine and vehicle. CONCLUSIONS--This study shows that the nasal vascular effects of histamine are less prominent than its actions on rhinorrhoea, and that the greater obstructive effect of bradykinin than histamine on NAR may contribute to the relative lack of efficacy of H1 antihistamines on nasal blockage in clinical disease.

Enhancement of neutrophil function by the bronchial epithelium stimulated by epidermal growth factor.

The bronchial epithelium is an important physical barrier that regulates physiological processes including leukocyte trafficking. The aim of the present study was to elucidate the mechanisms whereby the bronchial epithelium, stimulated by epidermal growth factor (EGF) as part of a response to acute or chronic injury, could activate and chemoattract human neutrophils. Subconfluent human bronchial epithelial (16HBE) cells were stimulated with EGF to mimic the in vivo events after injury. The effect of the resulting EGF-conditioned media (CM) was compared with that of basal-CM with respect to neutrophil activation and chemotaxis. Such findings were then confirmed using primary bronchial epithelial cells (PBECs) from healthy volunteers. EGF-CM from 16HBE cells caused increased expression of CD11b/CD66b and CD62L loss on neutrophils when compared with basal-CM. EGF-CM contained significant neutrophil chemotactic activity involving granulocyte-macrophage colony-stimulating factor and interleukin-8 that was potentiated by leukotriene B4. This was dependent on neutrophil phosphatidylinositol-3-kinase activation and Akt phosphorylation, with partial regulation by phospholipase D, but not mammalian target of rapamycin. Consistent with these observations, EGF-CM derived from PBECs displayed increased chemotactic activity. The present results suggest that the enhanced chemotactic activity of the epidermal growth factor-conditioned epithelium can enhance neutrophil-mediated immunity during acute injury, while during continued injury and repair, as in chronic asthma, this could contribute to persistent neutrophilic inflammation.

The allergen specificity of the late asthmatic reaction

To cite this article: Hatzivlassiou M, Grainge C, Kehagia V, Lau L, Howarth PH. The allergen specificity of the late asthmatic reaction. Allergy 2010; 65: 355–358.