Christine J. van Dalen

Eosinophil peroxidase produces hypobromous acid in the airways of stable asthmatics

Eosinophil peroxidase and myeloperoxidase use hydrogen peroxide to produce hypobromous acid and hypochlorous acid. These powerful oxidants may damage the lungs if they are produced as part of the inflammatory response in asthma. The aim of this study was to determine if peroxidases generate hypohalous acids in the airways of individuals with stable asthma, and if they affect lung function. Sputum was induced from patients with mild to moderate asthma and from healthy controls. Eosinophil peroxidase, myeloperoxidase, chlorinated and brominated tyrosyl residues, and protein carbonyls were measured in sputum supernatants. Eosinophil peroxidase protein was significantly elevated in asthmatic subjects whereas myeloperoxidase protein was not. There was significantly more 3-bromotyrosine (Br-Tyr) in proteins from the sputum of asthmatics compared to controls (0.79 vs. 0.23 mmol Br-Tyr/mol Tyr; medians p < .0001). Levels of 3-chlorotyrosine (0.23 vs. 0.14 mmol Cl-Tyr/mol Tyr; medians p = .11) and protein carbonyls (0.347 vs. 0.339 nmol/mg protein; medians p = .56) were not significantly increased in asthmatics. Levels of 3-bromotyrosine were strongly correlated with eosinophil peroxidase protein (r = 0.79, p < .0001). There were no significant correlations between the markers of oxidative stress and lung function. We conclude that eosinophil peroxidase produces substantial amounts of hypobromous acid in the airways of stable asthmatics. Although this highly reactive oxidant is a strong candidate for exacerbating inflammatory tissue damage in the lung, its role in asthma remains uncertain.

Suitability of Forced Expiratory Volume in 1 Second/Forced Vital Capacity vs Percentage of Predicted Forced Expiratory Volume in 1 Second for the Classification of Asthma Severity in Adolescents

OBJECTIVE To determine whether lung function alters asthma severity based on symptom history in asthmatic adolescents. DESIGN Data on asthma symptoms and lung function were collected from adolescents randomly selected from the general population. SETTING Five schools from the central Wellington, New Zealand, area during 2003 to 2005. PARTICIPANTS Two hundred twenty-four secondary school students aged 13 to 17 years (asthmatic, 118; nonasthmatic, 106). MAIN EXPOSURES Asthma questionnaire and lung function testing. MAIN OUTCOME MEASURES Distribution of asthmatic adolescents in each severity class based on symptoms, lung function, or a combination of both. RESULTS Median values for all spirometric parameters for asthmatic adolescents, apart from forced expiratory volume in the first second of expiration (FEV(1))/forced vital capacity (FVC), were in the normal range. Distribution of severity (based on symptoms and beta(2)-agonist use with adjustment for regular inhaled corticosteroid use) was 48.3%, mild; 28.8%, moderate; and 22.9%, severe asthma. For severity based on percentages of predicted FEV(1) and predicted forced expiratory flow, midexpiratory phase (FEF(25%-75%)) and FEV(1)/FVC, the percentages were 89.8%, 86.4%, and 63.5%, mild; 9.3%, 10.2%, and 18.6%, moderate; and 0.9%, 3.4%, and 17.8%, severe asthma, respectively. When percentages of predicted FEV(1) or predicted FEF(25%-75%) or FEV(1)/FVC were added to symptom severity, 6.8%, 5.1%, and 16.9% of asthmatic adolescents were reclassified into another severity group, respectively. CONCLUSIONS The majority of asthmatic adolescents have normal lung function despite experiencing significant asthma symptoms. Adding FEV(1)/FVC to symptom history changes the distribution of severity; however, both percentages of predicted FEV(1) and FEF(25%-75%) have little added effect in assessing asthma severity in adolescents.

Relationship between airway neutrophilia and ageing in asthmatics and non-asthmatics

Increased sputum neutrophilia has been observed in asthma, but also during normal ageing in asthmatics and non‐asthmatics. It remains unclear what constitutes ‘normal’ neutrophil levels in different age groups.

Importance of allergy in asthma: an epidemiologic perspective.

There has been a global epidemic of asthma during the past half-century. More recently, the prevalence has leveled off or declined in many Western countries, whereas the prevalence in less affluent nations is still increasing. The reasons for this and the different geographical patterns of asthma prevalence remain unclear. This paper provides an epidemiologic perspective on whether allergen exposure and allergies can explain these trends. In particular, the paper discusses 1) geographical and temporal trends in asthma and the role of allergens and allergy, 2) the importance of nonallergic mechanisms, 3) nonallergenic exposures that may modify the risk of allergies and asthma, and 4) new and emerging risk and protective factors. Although allergy and asthma are closely related, allergen exposure and allergy alone cannot explain current time trends and geographical patterns of asthma. Population-based studies focusing on recently identified risk and protective factors may provide further insight.

Measurement of Exhaled Nitric Oxide in a General Population Sample: A Comparison of the Medisoft HypAir FENO and Aerocrine NIOX Analyzers

Background and objective. Measuring the fraction of nitric oxide in exhaled breath (FENO) is increasingly utilized to assess airway inflammation in asthma. The primary aim of this study was to compare exhaled nitric oxide measurements obtained using two devices from different manufacturers, that is, the recently marketed portable and electrochemical-based Medisoft HypAir FENO and the well-established chemiluminescence-based Aerocrine NIOX analyzer, in an unselected population. Methods. FENO measurements were conducted in 106 subjects (86 healthy; 20 asthmatic; 56.6% atopic). Atopy and health status were assessed by skin prick tests and questionnaire, respectively. Results. The two instruments showed strong correlation over a wide range of FENO measurements (8–261.3 ppb with the HypAir, 5.6–156.8 ppb with the NIOX; r = 0.98; p < .0001). This correlation was observed in the population as a whole, as well as in healthy non-atopics, healthy atopics, and atopic asthmatics when considered separately. The measurements on the HypAir FENO were consistently 1.6 times (95% CI 1.11–2.05) higher than those obtained with the NIOX. Conclusions. FENO measurements obtained with the HypAir FENO correlated well with the NIOX, but were approximately 1.6 times higher. Therefore, a conversion factor is required if results are to be compared with the NIOX instrument.

Identifying leukocyte populations in fresh and cryopreserved sputum using flow cytometry

Airway inflammation is commonly assessed by sputum induction followed by a differential cell count (DCC) using light microscopy. This method is prone to intercounter variability and poor reproducibility. We aimed to develop a more objective method using flow cytometry (FCM).

Absence of airway inflammation in a large proportion of adolescents with asthma.

Neutrophilic inflammation has been implicated in non‐eosinophilic asthma (NEA) in adults, but little is known about NEA in children/adolescents. We assessed clinical and inflammatory characteristics of NEA in adolescent asthma.

Bromotyrosines in sputum proteins and treatment effects of terbutaline and budesonide in asthma

BACKGROUND Inhaled corticosteroids are widely used in the treatment of persistent asthma, usually combined with inhaled beta2-agonists. Previous research suggests that short-acting beta2-agonists (SABAs) may downregulate the anti-inflammatory effects of inhaled corticosteroids, thereby increasing asthma morbidity. OBJECTIVE To determine whether 3-bromotyrosine and 3,5-dibromotyrosine levels, specific markers of eosinophil activation, reflect treatment effects on airway inflammation of inhaled corticosteroids and SABAs and support previous conclusions. METHODS Levels of 3-bromotyrosine and 3,5-dibromotyrosine were measured in sputum supernatants using stable isotope dilution gas chromatography-mass spectrometry in a randomized, placebo-controlled, crossover study of treatment with terbutaline, budesonide, and their combination in patients with persistent asthma. Thirty-four individuals were randomized, and 28 completed the study. RESULTS Treatment with budesonide lowered median 3-bromotyrosine levels compared with treatment with placebo, terbutaline, and budesonide-terbutaline (0.24 vs 0.64, 0.62, and 0.43 3-bromotyosine/tyrosine [mmol/mol]; P < .05) and lowered median 3,5-dibromotyrosine levels compared with placebo and terbutaline treatments (0.04 vs 0.11 and 0.07 3,5-dibromotyrosine/ tyrosine [mmol/mol], P < .05). Unlike eosinophil numbers, 3-bromotyrosine and 3,5-dibromotyrosine levels did not increase with terbutaline treatment compared with placebo treatment but were significantly raised when terbutaline was added to budesonide treatment. 3-Bromotyrosine levels correlated significantly with eosinophil cationic protein levels in all groups. CONCLUSIONS 3-Bromotyrosine and 3,5-dibromotyrosine levels reflect treatment effects in asthma and support previous findings that SABAs impair the anti-inflammatory effects of inhaled corticosteroids. In addition to eosinophil numbers and eosinophil cationic protein levels, these modified tyrosine residues provide useful information about the inflammatory state of the airways.

Sputum basophils are increased in eosinophilic asthma compared with non-eosinophilic asthma phenotypes

Sputum basophil numbers are increased in allergic asthmatics, but it is unclear what role airway basophils play in “TH2‐low” asthma phenotypes. Using flow cytometry, we found that basophils were significantly increased in all asthmatics (n=26) compared with healthy controls (n=8) (P=0.007) with highest levels observed in eosinophilic asthma (EA); median 0.22%, IQR 0.11%‐0.47%; n=14) compared with non‐EA (NEA) (0.06%, 0.00%‐0.20%; n=12; P<0.05). In asthmatics, basophils were positively correlated with sputum eosinophils (r=0.54; P<0.005) and inversely with sputum neutrophils (r=−0.46: P<0.05), but not with FEV1 (% predicted), FEV1/FVC or bronchodilator reversibility. In a subgroup initially identified as inadequately controlled asthma (n=7), there was a trend (P=0.08) towards a reduction in sputum basophils following increased inhaled corticosteroid (ICS) treatment. Our findings suggest that basophils may be particularly important in eosinophilic asthma and that sputum basophil assessment could be a useful additional indicator of “TH2‐high” asthma.

Reactions of Myeloperoxidase and Production of Hypochlorous Acid in Neutrophil Phagosomes

It is generally accepted that the physiological function of myeloperoxidase is to produce hypochlorous acid. However, this green heme protein does not fit the classical definition of an enzyme. Usually an enzyme is specific in the reaction it catalyzes and in its choice of substrates. In contrast, myeloperoxidase has many and varied substrates, and catalyzes at least three different reactions. This indiscriminate catalytic power argues against a singular role for myeloperoxidase. Rather, it suggests that its function will be modulated by the local milieu. Therefore, we need to fully appreciate the complexity of its reaction mechanisms to understand how it acts within neutro-phil phagosomes during microbial killing and how it contributes to inflammation. In addition to hydrogen peroxide and chloride, potential physiological substrates for myeloperoxidase include superoxide, thiocyanate, tyrosine, tryptophan, urate, nitric oxide, and nitrite, and there are bound to be others. Their reactions also need to be considered when assessing the function of myeloperoxidase. In this chapter we outline the redox transformations that myeloperoxidase undergoes during turnover, and give examples of how the various alternative substrates influence its activity. We also take a kinetic modelling approach to predict how myeloperoxidase functions in the neutrophil phagosome and test what parameters are critical for efficient production of hypochlorous acid.