Development of a new in vitro system for continuous in vitro exposure of lung tissue to complex atmospheres: Application to diesel exhaust toxicology

The purpose of this study was the development of a new incubation system that can allow continuous exposure of lung tissue to complex atmospheres as a tool for the assessment of aerial environmental lung toxicology. To assess the pertinence of this new exposure system, we studied the impact of diesel engine exhausts as a complex atmosphere containing both gaseous and particulate fractions and have been able to discriminate between the toxicological impacts of the gaseous phase and particulate matter from diesel exhausts. Continuous flow-through rotating chambers with controlled pO₂, pCO₂, and hygrometry have been designed in which lung slices are positioned in rolling inserts that allow free access of atmosphere to the exposed lung tissue. Under control conditions, cell viability was preserved for at least 48 h as assessed by intracellular ATP, GSH, and K⁺ levels and slice O₂ consumption levels. Short-term exposure (1 h) to diesel whole exhausts did not affect intracellular potassium or slice O₂ consumption, while intracellular ATP and GSH levels were markedly decreased. Exposure to filtered exhausts showed less marked effects on both ATP and GSH levels. Superoxide dismutase activity was decreased in a similar way by both total and filtered exhausts while Se⁺-dependent glutathione peroxidase activity was induced by filtered exhausts to a larger extent than after total exhaust exposure, showing different response patterns of lung tissue after exposure to whole or filtered exhausts. In conclusion, this newly designed model opens a promising area in in vitro environmental lung toxicology testing.     

Repeated inhalations of diesel exhaust particles and oxidatively damaged DNA in young oxoguanine DNA glycosylase (OGG1) deficient mice

DNA repair may prevent increased levels of oxidatively damaged DNA from prolonged oxidative stress induced by, e.g. exposure to diesel exhaust particles (DEP). We studied oxidative damage to DNA in broncho-alveolar lavage cells, lungs, and liver after 4 × 1.5 h inhalations of DEP (20 mg/m³) in Ogg1^- / -  and wild type (WT) mice with similar extent of inflammation. DEP exposure increased lung levels of 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodG) in Ogg1^- / -  mice, whereas no effect on 8-oxodG or oxidized purines in terms of formamidopyrimidine DNA glycosylase (FPG) sites was observed in WT mice. In both unexposed and exposed Ogg1^- / -  mice the level of FPG sites in the lungs was 3-fold higher than in WT mice. The high basal level of FPG sites in Ogg1^- / -  mice probably saturated the assay and prevented detection of DEP-generated damage. In conclusion, Ogg1^- / -  mice have elevated pulmonary levels of FPG sites and accumulate genomic 8-oxodG after repeated inhalations of DEP.     

The size distribution of primary biological aerosol particles in the multiphase atmosphere

Primary biological aerosol particles are a ubiquitouscomponent of the atmospheric aerosol and have greatimportance within the whole atmosphere. Besides theireffect on air hygiene (i.e. causing allergic diseases), theycontribute to cloud and rain development. They amountto almost 25% of the total number of aerosolparticles both in dry air and in cloudwater. They showno seasonal variation in concentration but incomposition.     

The role of a mitochondrial pathway in the induction of apoptosis by chemicals extracted from diesel exhaust particles

We are interested in the cytotoxic and proinflammatory effects of particulate pollutants in the respiratory tract. We demonstrate that methanol extracts made from diesel exhaust particles (DEP) induce apoptosis and reactive oxygen species (ROS) in pulmonary alveolar macrophages and RAW 264.7 cells. The toxicity of these organic extracts mimics the cytotoxicity of the intact particles and could be suppressed by the synthetic sulfhydryl compounds, N-acetylcysteine and bucillamine. Because DEP-induced apoptosis follows cytochrome c release, we studied the effect of DEP chemicals on mitochondrially regulated death mechanisms. Crude DEP extracts induced ROS production and perturbed mitochondrial function before and at the onset of apoptosis. This mitochondrial perturbation follows an orderly sequence of events, which commence with a change in mitochondrial membrane potential, followed by cytochrome c release, development of membrane asymmetry (annexin V staining), and propidium iodide uptake. Structural damage to the mitochondrial inner membrane, evidenced by a decrease in cardiolipin mass, leads to O-*2 generation and uncoupling of oxidative phosphorylation (decreased intracellular ATP levels). N-acetylcysteine reversed these mitochondrial effects and ROS production. Overexpression of the mitochondrial apoptosis regulator, Bcl-2, delayed but did not suppress apoptosis. Taken together, these results suggest that DEP chemicals induce apoptosis in macrophages via a toxic effect on mitochondria.     

Characterizing biological aerosol in a chamber: an approach to estimation of viable organisms in a single biological particle

There are practical and valid reasons topresent biological field trial referenceresults as agent containing particles per literof air (ACPLA). However, workers in biologicalaerosol research have a need to know how manyviable individual organisms are contained in asingle particle of a given diameter. Anecdotalevidence may exist suggesting that the task hasbeen accomplished but without a way toreplicate the measurements, it is difficult toaccept unsubstantiated claims. It is verydifficult to declare a finite number thatsatisfies all the experimental requirements, asthe problem is a statistical probability issue. This paper describes a method for estimatingthe number with practical instructions forreplicating the work in other laboratories.The test aerosol was contained in a 90 m³chamber at concentrations as low as 5 ACPLA. Amodified version of slit sampler collectedviable particles. A statistical method was usedto demonstrate sampling predictability at 95%confidence level. By using glass fiber filtersmounted in a dichotomous sampler, samplingefficiencies were estimated for a variety ofcommon aerosol collectors. The accumulated datapermitted the estimation of the number ofviable spores per particle. For a 2.5 to4 µm particle, arguments have beenpresented for considering 4.5 as the mostprobable ACPLA value.     

The use of antlers to monitor temporal variation in environmental lead levels: a case study from an industrialized area in Germany

In order to assess temporal changes in ambient lead levels, Pb concentrations were determined in roe deer (Capreolus capreolus) antlers (n=116) that had been collected in the industrialized area of Siegen (western Germany) in the period 1948–2000. Lead concentrations ranged between 0.3 and 166.3 mg/kg dry weight. An overall decline in antler Pb concentration occurred from the mid-1970s onward, and lead concentrations in the samples from the periods 1980–1989 and 1990–2000 were significantly lower than those in samples from previous periods. Our findings indicate a marked decline in lead exposure of the roe deer inhabiting the study area in the recent past. This decline is attributed to a decrease in atmospheric lead deposition, caused mainly by the phase-out of leaded gasoline, and, in addition, by a reduction of lead emissions from stationary sources. The results of the present study and of previous investigations demonstrate that analysis of antler samples obtained in an area over a certain period can be used to reconstruct temporal changes in environmental lead levels.