Estimated effects of air pollution and space-time-activity on cardiopulmonary outcomes in healthy adults: A repeated measures study.

BACKGROUND: Exposure to air pollution is known to affect both short and long-term outcomes of the cardiopulmonary system; however, findings on short-term outcomes have been inconsistent and often from isolated and long-term rather than coexisting and short-term exposures, and among susceptible/unhealthy rather than healthy populations. AIMS: We aimed to investigate separately the annual, daily and daily space-time-activity-weighted effect of ambient air pollution, as well as confounding or modification by other environmental (including noise) or space-time-activity (including total daily physical activity) exposures, on cardiopulmonary outcomes in healthy adults. METHODS: Participants (N=57: 54% female) had indicators of cardiopulmonary outcomes [blood pressure (BP), pulse (HR) and heart rate variability (HRV {SDNN}), and lung function (spirometry {FEV1, FVC, SUM})] measured on four different mornings (at least five days apart) in a clinical setting between 2011 and 2014. Spatiotemporal ESCAPE-LUR models were used to estimate daily and annual air pollution exposures (including PM10, PMCoarse, but not Ozone {derived from closest station}) at participant residential and occupational addresses. Participants' time-activity diaries indicated time spent at either address to allow daily space-time-activity-weighted estimates, and capture total daily physical activity (total-PA {as metabolic-equivalents-of-task, METs}), in the three days preceding health measurements. Multivariate-adjusted linear mixed-effects models (using either annual or daily estimates) were adjusted for possible environmental confounders or mediators including levels of ambient noise and greenness. Causal mediation analysis was also performed separately considering these factors as well as total-PA. All presented models are controlled by age, height, sex and season. RESULTS: An increase in 5µg/m3 of daily space-time-activity-weighted PMCoarse exposure was statistically significantly associated with a 4.1% reduction in total heart rate variability (SDNN; p=0.01), and remained robust after adjusting for suspected confounders [except for occupational-address noise (ß=-2.7, p=0.20)]. An increase in 10ppb of annual mean Ozone concentration at the residential address was statistically significantly associated with an increase in diastolic BP of 6.4mmHg (p<0.01), which lost statistical significance when substituted with daily space-time-activity-weighted estimates. As for pulmonary function, an increase in 10µg/m3 of annual mean PM10 concentration at the residential address was significantly associated with a 0.3% reduction in FVC (p<0.01) and a 0.5% reduction in SUM (p<0.04), for which again significance was lost when substituted for daily space-time-activity-weighted estimates These associations with pulmonary function remained robust after adjusting for suspected confounders, including annual Ozone, as well as total-PA and bioaerosol (pollen and fungal spore) levels (but not residential-neighborhood greenness {ß=-0.22, p=0.09; ß=-0.34, p=0.15, respectively}). Multilevel mediation analysis indicated that the proportion mediated as a direct effect on cardiopulmonary outcomes by suspected confounders (including total-PA, residential-neighborhood greenness, and occupational-address noise level) from primary exposures (including PM10, PMCoarse, and O3) was not statistically significant. CONCLUSION: Our findings suggest that increased daily space-time-activity-weighted PMCoarse exposure levels significantly adversely affect cardiac autonomic modulation (as reduced total HRV) among healthy adults. Additionally, increased annual levels at the residential address of Ozone and PM10 significantly increase diastolic blood pressure and reduce lung function, respectively, among healthy adults. These associations typically remained robust when adjusting for suspected confounders. Occupational-address noise and residential-neighborhood greenness levels, however, were seen as mediators of cardiovascular and pulmonary outcomes, respectively. Total daily physical activity was not seen as a mediator of any of the studied outcomes, which supports the promotion of active mobility within cities.

Acute respiratory response to traffic-related air pollution during physical activity performance.

BACKGROUND: Physical activity (PA) has beneficial, whereas exposure to traffic related air pollution (TRAP) has adverse, respiratory effects. Few studies, however, have examined if the acute effects of TRAP upon respiratory outcomes are modified depending on the level of PA. OBJECTIVES: The aim of our study was to disentangle acute effects of TRAP and PA upon respiratory outcomes and assess the impact of participants TRAP pre-exposure. METHODS: We conducted a real-world crossover study with repeated measures of 30 healthy adults. Participants completed four 2-h exposure scenarios that included either rest or intermittent exercise in high- and low-traffic environments. Measures of respiratory function were collected at three time points. Pre-exposure to TRAP was ascertained from land-use-modeled address-attributed values. Mixed-effects models were used to estimate the impact of TRAP and PA on respiratory measures as well as potential effect modifications. RESULTS: We found that PA was associated with a statistically significant increases of FEV1 (48.5mL, p=0.02), FEV1/FVC (0.64%, p=0.005) and FEF25-75% (97.8mL, p=0.02). An increase in exposure to one unit (1µg/m3) of PMcoarse was associated with a decrease in FEV1 (-1.31mL, p=0.02) and FVC (-1.71mL, p=0.01), respectively. On the other hand, for an otherwise equivalent exposure an increase of PA by one unit (1%Heart rate max) was found to reduce the immediate negative effects of particulate matter (PM) upon PEF (PM2.5, 0.02L/min, p=0.047; PM10, 0.02L/min p=0.02; PMcoarse, 0.03L/min, p=0.02) and the several hours delayed negative effects of PM upon FVC (PMcoarse, 0.11mL, p=0.02). The negative impact of exposure to TRAP constituents on FEV1/FVC and PEF was attenuated in those participants with higher TRAP pre-exposure levels. CONCLUSIONS: Our results suggest that associations between various pollutant exposures and respiratory measures are modified by the level of PA during exposure and TRAP pre-exposure of participants.

Evaluation of a human neurite growth assay as specific screen for developmental neurotoxicants.

Organ-specific in vitro toxicity assays are often highly sensitive, but they lack specificity. We evaluated here examples of assay features that can affect test specificity, and some general procedures are suggested on how positive hits in complex biological assays may be defined. Differentiating human LUHMES cells were used as potential model for developmental neurotoxicity testing. Forty candidate toxicants were screened, and several hits were obtained and confirmed. Although the cells had a definitive neuronal phenotype, the use of a general cell death endpoint in these cultures did not allow specific identification of neurotoxicants. As alternative approach, neurite growth was measured as an organ-specific functional endpoint. We found that neurite extension of developing LUHMES was specifically inhibited by diverse compounds such as colchicine, vincristine, narciclasine, rotenone, cycloheximide, or diquat. These compounds reduced neurite growth at concentrations that did not compromise cell viability, and neurite growth was affected more potently than the integrity of developed neurites of mature neurons. A ratio of the EC50 values of neurite growth inhibition and cell death of >4 provided a robust classifier for compounds associated with a developmental neurotoxic hazard. Screening of unspecific toxicants in the test system always yielded ratios <4. The assay identified also compounds that accelerated neurite growth, such as the rho kinase pathway modifiers blebbistatin or thiazovivin. The negative effects of colchicine or rotenone were completely inhibited by a rho kinase inhibitor. In summary, we suggest that assays using functional endpoints (neurite growth) can specifically identify and characterize (developmental) neurotoxicants.

Assessment of chemical-induced impairment of human neurite outgrowth by multiparametric live cell imaging in high-density cultures.

Chemicals that specifically alter human neurite outgrowth pose a hazard for the development of the nervous system. The identification of such compounds remains a major challenge, especially in a human test system. To address this issue, we developed an imaging-based procedure in LUHMES human neuronal precursor cells to quantify neurite growth of unfixed cultures. Live imaging allowed the simultaneous evaluation of cell viability and neurite outgrowth within one culture dish. The procedure was used to test the hypothesis that inhibitors of specific pathways can impair neurite outgrowth without affecting cell viability. Although the cells were grown at high density to allow extensive networking, overall neurite growth in this complex culture was quantified with a signal-to-noise ratio of > 50. Compounds such as U0126 slowed the extension of neuronal processes at concentrations > 4 times lower than those causing cell death. High numbers of individual viable cells without neurites were identified under such conditions, and neurite outgrowth recovered after washout of the chemical. Also an extension-promoting compound, Y-27632, was identified by this unique multiparametric imaging approach. Finally, the actions of unspecific cytotoxicants such as menadione, cadmium chloride, and sodium dodecyl sulfate were tested to evaluate the specificity of the new assay. We always found a ratio of EC50 (cell death)/EC50 (neurites) < 4 for such chemicals. The described novel test system may thus be useful both for high-throughput screens to identify neuritotoxic agents and for their closer characterization concerning mode of action, compound interactions, or the reversibility of their effects.

Requirement of a dopaminergic neuronal phenotype for toxicity of low concentrations of 1-methyl-4-phenylpyridinium to human cells.

LUHMES cells are conditionally-immortalized non-transformed human fetal cells that can be differentiated to acquire a dopaminergic neuron-like phenotype under appropriate growth conditions. After differentiation by GDNF and cyclic adenosine monophosphate, LUHMES were sensitive to 1-methyl-4-phenylpyridinium (MPP(+)) toxicity at < or =5 microM, but resistant to the parental compound 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). The high homogeneity and purity of the cultures allowed the detection of metabolic changes during the degeneration. Cellular ATP dropped in two phases after 24 and 48 h; cellular glutathione (GSH) decreased continuously, paralleled by an increase in lipid peroxidation. These events were accompanied by a time-dependent degeneration of neurites. Block of the dopamine transporter by GBR 12909 or mazindol completely abrogated MPP(+) toxicity. Inhibition of de novo dopamine synthesis by alpha-methyl-l-tyrosine or 3-iodo-l-tyrosine attenuated toxicity, but did not reduce the initial drop in ATP. Inhibition of mixed lineage kinases by CEP1347 completely prevented the MPP(+)-induced loss of viability and intracellular GSH, but failed to attenuate the initial drop of ATP. For the quantitative assessment of neurite degeneration, an automated imaging-based high content screening approach was applied and confirmed the findings made by pharmacological interventions in this study. Our data indicate that inhibition of mitochondrial ATP synthesis is not sufficient to trigger cell death in MPP(+)-treated LUHMES.