Mandy Pui

MicroRNA expression in response to controlled exposure to diesel exhaust: attenuation by the antioxidant N-acetylcysteine in a randomized crossover study.

Background: Adverse health effects associated with diesel exhaust (DE) are thought to be mediated in part by oxidative stress, but the detailed mechanisms are largely unknown. MicroRNAs (miRNAs) regulate gene expression post-transcriptionally and may respond to exposures such as DE. Objectives: We profiled peripheral blood cellular miRNAs in participants with mild asthma who were exposed to controlled DE with and without antioxidant supplementation. Methods: Thirteen participants with asthma underwent controlled inhalation of filtered air and DE in a double-blinded, randomized crossover study of three conditions: a) DE plus placebo (DEP), b) filtered air plus placebo (FAP), or c) DE with N-acetylcysteine supplementation (DEN). Total cellular RNA was extracted from blood drawn before exposure and 6 hr after exposure for miRNA profiling by the NanoString nCounter assay. MiRNAs significantly associated with DEP exposure and a predicted target [nuclear factor (erythroid-derived 2)-like 2 (NRF2)] as well as antioxidant enzyme genes were assessed by reverse transcription–quantitative polymerase chain reaction (RT-qPCR) for validation, and we also assessed the ability of N-acetylcysteine supplementation to block the effect of DE on these specific miRNAs. 8-hydroxy-2´-deoxyguanosine (8-OHdG) was measured in plasma as a systemic oxidative stress marker. Results: Expression of miR-21, miR-30e, miR-215, and miR-144 was significantly associated with DEP. The change in miR-144 was validated by RT-qPCR. NRF2 and its downstream antioxidant genes [glutamate cysteine ligase catalytic subunit (GCLC) and NAD(P)H:quinone oxidoreductase 1 (NQO1)] were negatively associated with miR-144 levels. Increases in miR-144 and miR-21 were associated with plasma 8-hydroxydeoxyguanosine 8-OHdG level and were blunted by antioxidant (i.e, DEN). Conclusions: Systemic miRNAs with plausible biological function are altered by acute moderate-dose DE exposure. Oxidative stress appears to mediate DE-associated changes in miR-144.

Diesel exhaust augments allergen-induced lower airway inflammation in allergic individuals: a controlled human exposure study

Rationale Traffic-related air pollution has been shown to augment allergy and airway disease. However, the enhancement of allergenic effects by diesel exhaust in particular is unproven in vivo in the human lung, and underlying details of this apparent synergy are poorly understood. Objective To test the hypothesis that a 2 h inhalation of diesel exhaust augments lower airway inflammation and immune cell activation following segmental allergen challenge in atopic subjects. Methods 18 blinded atopic volunteers were exposed to filtered air or 300 µg PM2.5/m3 of diesel exhaust in random fashion. 1 h post-exposure, diluent-controlled segmental allergen challenge was performed; 2 days later, samples from the challenged segments were obtained by bronchoscopic lavage. Samples were analysed for markers and modifiers of allergic inflammation (eosinophils, Th2 cytokines) and adaptive immune cell activation. Mixed effects models with ordinal contrasts compared effects of single and combined exposures on these end points. Results Diesel exhaust augmented the allergen-induced increase in airway eosinophils, interleukin 5 (IL-5) and eosinophil cationic protein (ECP) and the GSTT1 null genotype was significantly associated with the augmented IL-5 response. Diesel exhaust alone also augmented markers of non-allergic inflammation and monocyte chemotactic protein (MCP)-1 and suppressed activity of macrophages and myeloid dendritic cells. Conclusion Inhalation of diesel exhaust at environmentally relevant concentrations augments allergen-induced allergic inflammation in the lower airways of atopic individuals and the GSTT1 genotype enhances this response. Allergic individuals are a susceptible population to the deleterious airway effects of diesel exhaust. Trial registration number NCT01792232.

Anti-Oxidant N-Acetylcysteine Diminishes Diesel Exhaust-Induced Increased Airway Responsiveness in Person with Airway Hyper-Reactivity

BACKGROUND Inhalation of diesel exhaust (DE) at moderate concentrations causes increased airway responsiveness in asthmatics and increased airway resistance in both healthy and asthmatic subjects, but the effect of baseline airway responsiveness and anti-oxidant supplementation on this dynamic is unknown. OBJECTIVES We aimed to determine if changes in airway responsiveness due to DE are attenuated by thiol anti-oxidant supplementation, particularly in those with underlying airway hyper-responsiveness. METHODS Participants took N-acetylcysteine (600 mg) or placebo capsules three times daily for 6 days. On the last of these 6 days, participants were exposed for 2 h to either filtered air (FA) or DE (300 μg/m(3) of particulate matter smaller than 2.5 microns). Twenty-six non-smokers were studied under each of three experimental conditions (filtered air with placebo, diesel exhaust with placebo, and diesel exhaust with N-acetylcysteine) using a randomized, double-blind, crossover design, with a 2-week washout between conditions. Methacholine challenge was performed pre-exposure (baseline airway responsiveness) and post-exposure (effect of exposure). RESULTS Anti-oxidant supplementation reduced baseline airway responsiveness in hyper-responsive individuals by 20% (p = 0.001). In hyper-responsive individuals, airway responsiveness increased 42% following DE compared with FA (p = 0.03) and this increase was abrogated with anti-oxidant supplementation (diesel exhaust with N-acetylcysteine vs. filtered air with placebo, p = 0.85). CONCLUSIONS Anti-oxidant (N-acetylcysteine) supplementation protects against increased airway responsiveness associated with DE inhalation and reduces need for supplement bronchodilators in those with baseline airway hyper-responsiveness. Individuals with variants in genes of oxidative stress metabolism when exposed to DE are protected from increases in airway responsiveness if taking anti-oxidant supplementation.

Genes, the environment and personalized medicine

The brave new world envisioned by proponents of personalized medicine has attracted considerable interest and investment during the past decade or so. The excitement is understandable because personalized medicine promises to drastically improve individual health and make more efficient use of existing resources, changing both health care and public health for the better. Improved use of resources is becoming particularly important, as many national healthcare schemes are straining to maintain affordable health care of acceptable quality under the combined pressures of rising costs, an aging population and the increasing prevalence of many chronic and common diseases. Although not a panacea for all these problems, personalized medicine could theoretically reduce healthcare costs, as an individual’s genetic or other biological information could be used to make better or earlier diagnoses of disease, apply cheaper, preventive measures to decrease disease risk, and make more efficient use of therapeutic options. However, there remains a considerable gap between theory and reality, and we think that the prevailing focus on an individual’s genes and biology insufficiently incorporates the important role of environmental factors in disease etiology and health. Including these factors in our approach to personalized medicine and population health should bring that theory closer to reality. However, it will require a fundamental change to the current research agenda and public health policies to emphasize the role of the social and physical environments and related epigenetic changes.

PM10-stimulated airway epithelial cells activate primary human dendritic cells independent of uric acid: application of an in vitro model system exposing dendritic cells to airway epithelial cell-conditioned media.

Airway epithelial cells represent the first line of defence against inhaled insults, including air pollution. Air pollution can activate innate immune signalling in airway epithelial cells leading to the production of soluble mediators that can influence downstream inflammatory cells. Our objective was to develop and validate a model of dendritic cell exposure to airway epithelial cell‐conditioned media. After establishing the model, we explored how soluble mediators released from airway epithelial cells in response to air pollution influenced the phenotype of dendritic cells.

Respiratory Impairment and Systemic Inflammation in Cedar Asthmatics Removed from Exposure

Background Prior research has shown that removing occupational asthmatics from exposure does not routinely lead to significant improvements in respiratory impairment. These studies were of limited duration and factors determining recovery remain obscure. Our objective was to evaluate residual respiratory impairment and associated sputum and blood biomarkers in subjects with Western red cedar asthma after exposure cessation. Methods Subjects previously diagnosed with cedar asthma, and removed from exposure to cedar dust for at least one year, were recruited. Subjects completed a questionnaire and spirometry. PC20 (methacholine concentration that produces 20% fall in FEV1 (forced expiratory volume at 1 second)) sputum cellularity and select Th1/Th2 (T helper cells 1 and 2) cytokine concentrations in peripheral blood were determined. The asthma impairment class was determined and multivariate analyses were performed to determine its relationship with sputum cell counts and serum cytokines. Results 40 non-smoking males (mean age 62) were examined at a mean interval of 25 years from cedar asthma diagnosis and 17 years from last cedar exposure. 40% were in impairment class 2/3. On average, the PC20 had increased by 2.0 mg/ml; the FEV1 decreased by 1.5 L, with greater decrease in those with greater impairment. Higher impairment was associated with serum interferon-gamma (mean = 1.3 pg/ml in class 2/3 versus 0.62 pg/ml in class 0/1, p = 0.04), mainly due to the FEV1 component (correlation with interferon-gamma = −0.46, p = 0.005). Conclusion Years after exposure cessation, patients with Western red cedar asthma have persistent airflow obstruction and respiratory impairment, associated with systemic inflammation.

Flow cytometry to identify leukocyte sub-populations in blood and induced sputum in asthmatic and healthy volunteers exposed to diesel exhaust

Methods Mild asthmatics and normal controls were recruited as study subjects. This crossover study was double-blinded, randomized and counter-balanced to the order of three conditions: diesel exhaust with anti-oxidant, diesel exhaust with placebo, or filtered air with placebo. The subjects were exposed to either filtered air or diesel exhaust (300 ug PM2.5/m ) in a state-of-the-art diesel exhaust exposure facility. An anti-oxidant, N-acetylcysteine (600 mg), or a placebo was taken orally for five days preceding, and on the day of the exposure. Each subject was exposed to each of the three conditions. Peripheral blood samples were taken pre-exposure, and also at 2, 6, and 30 hours after the beginning of exposure. Sputum induction was performed by inhalation of hypertonic saline according to ATS guidelines pre-exposure, and also at 6, and 30 hours after the beginning of exposure. FACSCanto II (BD Biosciences) was used for flow cytometry. A 5-colour, 12-marker (CD3/CD9/ CD14/CD16/CD19/CD20/CD45/CD56/CD83/CD206/ CD326/HLA-DR) combination was used to identify dendritic cells, macrophages, monocytes, neutrophils, eosinophils, and bronchial epithelial cells. Direct immunolabelling was performed on whole peripheral blood. After incubation, red blood cells were lysed. Remaining cells were washed and resuspended in PBS with 0.5% paraformaldehyde. Sputum plugs were homogenized with 0.1% DTT, filtered, and then centrifuged to remove supernatant. Sputum cells were resuspended in PBS at 1 million per mL. Direct immunolabelling was performed. After incubation, cells were washed and resuspended in PBS with 0.5% paraformaldehyde. Spectral compensation for flow cytometry was performed using an automatic calibration technique (BD CompBeads). Cellular debris was eliminated on the SSC/FSC scattergram. A gating strategy was designed to identify the leukocyte sub-populations and bronchial epithelial cells. Surface markers were chosen based on differential cell-specific expression according to existing literature.

Sputum adiponectin as a marker for western red cedar asthma

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A qualitative study of the knowledge, attitudes, and behaviors of people exposed to diesel exhaust at the workplace in British Columbia, Canada

Purpose To identify exposure-related knowledge, attitudes and behaviors of individuals occupationally exposed to diesel exhaust (DE); to reveal strengths, knowledge gaps and misperceptions therein. Methods A Mental Models approach was used to gather information about current scientific understanding of DE exposure hazards and the ways in which exposure can be reduced. Thirty individuals in British Columbia who were regularly exposed to occupational DE were interviewed. The audio was recorded and transcribed. Data was grouped together and examined to draw out themes around DE awareness, hazard assessment and risk reduction behaviors. These themes were then compared and contrasted with existing grey and research literature in order to reveal strengths, gaps and misperceptions regarding DE exposure. Results Study participants were aware and concerned about their exposure to DE but had incomplete and sometimes incorrect understanding of exposure pathways, health effects, and effective strategies to reduce their exposures. The perceived likelihood of exposure to DE was significantly greater compared to that of other work hazards (p<0.01), whereas the difference for their perceived severity of consequences was not significant. There was no universally perceived main source of information regarding DE, and participants generally distrusted sources of information based on their past experience with the source. Most of the actions that were taken to address DE exposure fell into the area of administrative controls such as being aware of sources of DE and avoiding these sources. Conclusions This study of the knowledge, attitude, and behavior of those occupationally exposed to DE found, most notably, that more education and training and the creation of a health effects inventory regarding DE exposure were desired.