Assessment of biological responses of EpiAirway 3-D cell constructs versus A549 cells for determining toxicity of ambient air pollution

EpiAirway™ 3-D constructs are human-derived cell cultures of differentiated airway epithelial cells that may represent a more biologically relevant model of the human lung. However, limited information is available of its utility for exposures to air pollutants at the air-liquid interface (ALI)

Effects of 1,3-Butadiene, Isoprene, and Their Photochemical Degradation Products on Human Lung Cells

Because of potential exposure both in the workplace and from ambient air, the known carcinogen 1,3-butadiene (BD) is considered a priority hazardous air pollutant. BD and its 2-methyl analog, isoprene (ISO), are chemically similar but have very different toxicities, with ISO showing no significant carcinogenesis. Once released into the atmosphere, reactions with species induced by sunlight and nitrogen oxides convert BD and ISO into several photochemical reaction products. In this study, we determined the relative toxicity and inflammatory gene expression induced by exposure of A549 cells to BD, ISO, and their photochemical degradation products in the presence of nitric oxide. Gas chromatography and mass spectrometry analyses indicate the initial and major photochemical products produced during these experiments for BD are acrolein, acetaldehyde, and formaldehyde, and products for ISO are methacrolein, methyl vinyl ketone, and formaldehyde; both formed < 200 ppb of ozone. After exposure the cells were examined for cytotoxicity and interleukin-8 (IL-8) gene expression, as a marker for inflammation. These results indicate that although BD and ISO alone caused similar cytotoxicity and IL-8 responses compared with the air control, their photochemical products significantly enhanced cytotoxicity and IL-8 gene expression. This suggests that once ISO and BD are released into the environment, reactions occurring in the atmosphere transform these hydrocarbons into products that induce potentially greater adverse health effects than the emitted hydrocarbons by themselves. In addition, the data suggest that based on the carbon concentration or per carbon basis, biogenic ISO transforms into products with proinflammatory potential similar to that of BD products

A Toxicogenomic Comparison of Primary and Photochemically Altered Air Pollutant Mixtures

Background: Air pollution contributes significantly to global increases in mortality, particularly within urban environments. Limited knowledge exists on the mechanisms underlying health effects resulting from exposure to pollutant mixtures similar to those occurring in ambient air. In order to clarify the mechanisms underlying exposure effects, toxicogenomic analyses are used to evaluate genomewide transcript responses and map these responses to molecular networks

In Vitro Exposures in Diesel Exhaust Atmospheres: Resuspension of PM from Filters versus Direct Deposition of PM from Air

One of the most widely used in vitro particulate matter (PM) exposures methods is the collection of PM on filters, followed by resuspension in a liquid medium, with subsequent addition onto a cell culture. To avoid disruption of equilibria between gases and PM, we have developed a direct in vitro sampling and exposure method (DSEM) capable of PM-only exposures. We hypothesize that the separation of phases and post-treatment of filter-collected PM significantly modifies the toxicity of the PM compared to direct deposition, resulting in a distorted view of the potential PM health effects

Epigenetic Changes Induced by Air Toxics: Formaldehyde Exposure Alters miRNA Expression Profiles in Human Lung Cells

Bac k g r o u n d: Exposure to formaldehyde, a known air toxic, is associated with cancer and lung disease. Despite the adverse health effects of formaldehyde, the mechanisms underlying formaldehydeinduced disease remain largely unknown. Research has uncovered microRNAs (miRNAs) as key posttranscriptional regulators of gene expression that may influence cellular disease state. Although studies have compared different miRNA expression patterns between diseased and healthy tissue, this is the first study to examine perturbations in global miRNA levels resulting from formaldehyde exposure. Objectives: We investigated whether cellular miRNA expression profiles are modified by formaldehyde exposure to test the hypothesis that formaldehyde exposure disrupts miRNA expression levels within lung cells, representing a novel epigenetic mechanism through which formaldehyde may induce disease. Me t h o d s: Human lung epithelial cells were grown at air–liquid interface and exposed to gaseous formaldehyde at 1 ppm for 4 hr. Small RNAs and protein were collected and analyzed for miRNA expression using microarray analysis and for interleukin (IL-8) protein levels by enzyme-linked immunosorbent assay (ELISA). Res u l t s: Gaseous formaldehyde exposure altered the miRNA expression profiles in human lun

In Vitro Exposure to Isoprene-Derived Secondary Organic Aerosol by Direct Deposition and its Effects on COX-2 and IL-8 Gene Expression

Atmospheric oxidation of isoprene, the most abundant non-methane hydrocarbon emitted into Earth's atmosphere primarily from terrestrial vegetation, is now recognized as a major contributor to the global secondary organic aerosol (SOA) burden. Anthropogenic pollutants significantly enhance isoprene SOA formation through acid-catalyzed heterogeneous chemistry of epoxide products. Since isoprene SOA formation as a source of fine aerosol is a relatively recent discovery, research is lacking on evaluating its potential adverse effects on human health. The objective of this study was to examine the effect of isoprene-derived SOA on inflammation-associated gene expression in human lung cells using a direct deposition exposure method. We assessed altered expression of inflammation-related genes in human bronchial epithelial cells (BEAS-2B) exposed to isoprene-derived SOA generated in an outdoor chamber facility. Measurements of gene expression of known inflammatory biomarkers interleukin 8 (IL-8) and cyclooxygenase 2 (COX-2) in exposed cells, together with complementary chemical measurements, showed that a dose of 0.067 µg cm−2 of SOA from isoprene photooxidation leads to statistically significant increases in IL-8 and COX-2 mRNA levels. Resuspension exposures using aerosol filter extracts corroborated these findings, supporting the conclusion that isoprene-derived SOA constituents induce the observed changes in mRNA levels. The present study is an attempt to examine the early biological responses of isoprene SOA exposure in human lung cells

The Gillings Sampler – An electrostatic air sampler as an alternative method for aerosol in vitro exposure studies

There is growing interest in studying the toxicity and health risk of exposure to multi-pollutant mixtures found in ambient air, and the U.S. Environmental Protection Agency (EPA) is moving towards setting standards for these types of mixtures. Additionally, the Health Effects Institute's strategic plan aims to develop and apply next-generation multi-pollutant approaches to understanding the health effects of air pollutants. There's increasing concern that conventional in vitro exposure methods are not adequate to meet EPA's strategic plan to demonstrate a direct link between air pollution and health effects. To meet the demand for new in vitro technology that better represents direct air-to-cell inhalation exposures, a new system that exposes cells at the air-liquid interface was developed. This new system, named the Gillings Sampler, is a modified two-stage electrostatic precipitator that provides a viable environment for cultured cells. Polystyrene latex spheres were used to determine deposition efficiencies (38-45%), while microscopy and imaging techniques were used to confirm uniform particle deposition. Negative control A549 cell exposures indicated the sampler can be operated for up to 4 hours without inducing any significant toxic effects on cells, as measured by lactate dehydrogenase (LDH) and interleukin-8 (IL-8). A novel positive aerosol control exposure method, consisting of a p-tolualdehyde (TOLALD) impregnated mineral oil aerosol (MOA), was developed to test this system. Exposures to the toxic MOA at a 1 ng/cm2 dose of TOLALD yielded a reproducible 1.4 and 2 fold increase in LDH and IL-8 mRNA levels over controls. This new system is intended to be used as an alternative research tool for aerosol in vitro exposure studies. While further testing and optimization is still required to produce a “commercially ready” system, it serves as a stepping-stone in the development of cost-effective in vitro technology that can be made accessible to researchers in the near future

Epigenetic Events Determine Tissue-Specific Toxicity of Inhalational Exposure to the Genotoxic Chemical 1,3-Butadiene in Male C57BL/6J Mice

1,3-Butadiene (BD), a widely used industrial chemical and a ubiquitous environmental pollutant, is a known human carcinogen. Although genotoxicity is an established mechanism of the tumorigenicity of BD, epigenetic effects have also been observed in livers of mice exposed to the chemical. To better characterize the diverse molecular mechanisms of BD tumorigenicity, we evaluated genotoxic and epigenotoxic effects of BD exposure in mouse tissues that are target (lung and liver) and non-target (kidney) for BD-induced tumors. We hypothesized that epigenetic alterations may explain, at least in part, the tissue-specific differences in BD tumorigenicity in mice. We evaluated the level of N-7-(2,3,4-trihydroxybut-1-yl)guanine adducts and 1,4-bis-(guan-7-yl)-2,3-butanediol crosslinks, DNA methylation, and histone modifications in male C57BL/6 mice exposed to filtered air or 425 ppm of BD by inhalation (6 h/day, 5 days/week) for 2 weeks. Although DNA damage was observed in all three tissues of BD-exposed mice, variation in epigenetic effects clearly existed between the kidneys, liver, and lungs. Epigenetic alterations indicative of genomic instability, including demethylation of repetitive DNA sequences and alterations in histone-lysine acetylation, were evident in the liver and lung tissues of BD-exposed mice. Changes in DNA methylation were insignificant in the kidneys of treated mice, whereas marks of condensed heterochromatin and transcriptional silencing (histone-lysine trimethylation) were increased. These modifications may represent a potential mechanistic explanation for the lack of tumorigenesis in the kidney. Our results indicate that differential tissue susceptibility to chemical-induced tumorigenesis may be attributed to tissue-specific epigenetic alterations

Delays in Appropriate Antibiotic Therapy for Gram-Negative Bloodstream Infections: A Multicenter, Community Hospital Study

BackgroundGram-negative bacterial bloodstream infection (BSI) is a serious condition with estimated 30% mortality. Clinical outcomes for patients with severe infections improve when antibiotics are appropriately chosen and given early. The objective of this study was to estimate the association of prior healthcare exposure on time to appropriate antibiotic therapy in patients with gram-negative BSI.MethodWe performed a multicenter cohort study of adult, hospitalized patients with gram-negative BSI using time to event analysis in nine community hospitals from 2003-2006. Event time was defined as the first administration of an antibiotic with in vitro activity against the infecting organism. Healthcare exposure status was categorized as community-acquired, healthcare-associated, or hospital-acquired. Time to appropriate therapy among groups of patients with differing healthcare exposure status was assessed using Kaplan-Meier analyses and multivariate Cox proportional hazards models.ResultsThe cohort included 578 patients with gram-negative BSI, including 320 (55%) healthcare-associated, 217 (38%) community-acquired, and 41 (7%) hospital-acquired infections. 529 (92%) patients received an appropriate antibiotic during their hospitalization. Time to appropriate therapy was significantly different among the groups of healthcare exposure status (log-rank p=0.02). Time to first antibiotic administration regardless of drug appropriateness was not different between groups (p=0.3). The unadjusted hazard ratios (HR) (95% confidence interval) were 0.80 (0.65-0.98) for healthcare-associated and 0.72 (0.63-0.82) for hospital-acquired, relative to patients with community-acquired BSI. In multivariable analysis, interaction was found between the main effect and baseline Charlson comorbidity index. When Charlson index was 3, adjusted HRs were 0.66 (0.48-0.92) for healthcare-associated and 0.57 (0.44-0.75) for hospital-acquired, relative to patients with community-acquired infections.ConclusionsPatients with healthcare-associated or hospital-acquired BSI experienced delays in receipt of appropriate antibiotics for gram-negative BSI compared to patients with community-acquired BSI. This difference was not due to delayed initiation of antibiotic therapy, but due to the inappropriate choice of antibiotic

Effects of High-Volume Versus High-Load Resistance Training on Skeletal Muscle Growth and Molecular Adaptations

We evaluated the effects of higher-load (HL) versus (lower-load) higher-volume (HV) resistance training on skeletal muscle hypertrophy, strength, and muscle-level molecular adaptations. Trained men (n = 15, age: 23 ± 3 years; training experience: 7 ± 3 years) performed unilateral lower-body training for 6 weeks (3× weekly), where single legs were randomly assigned to HV and HL paradigms. Vastus lateralis (VL) biopsies were obtained prior to study initiation (PRE) as well as 3 days (POST) and 10 days following the last training bout (POSTPR). Body composition and strength tests were performed at each testing session, and biochemical assays were performed on muscle tissue after study completion. Two-way within-subject repeated measures ANOVAs were performed on most dependent variables, and tracer data were compared using dependent samples t-tests. A significant interaction existed for VL muscle cross-sectional area (assessed via magnetic resonance imaging; interaction p = 0.046), where HV increased this metric from PRE to POST (+3.2%, p = 0.018) whereas HL training did not (−0.1%, p = 0.475). Additionally, HL increased leg extensor strength more so than HV training (interaction p = 0.032; HV \u3c HL at POST and POSTPR, p \u3c 0.025 for each). Six-week integrated non-myofibrillar protein synthesis (iNon-MyoPS) rates were also higher in the HV versus HL condition, while no difference between conditions existed for iMyoPS rates. No interactions existed for other strength, VL morphology variables, or the relative abundances of major muscle proteins. Compared to HL training, 6 weeks of HV training in previously trained men optimizes VL hypertrophy in lieu of enhanced iNon-MyoPS rates, and this warrants future research