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Michaela Aufderheide

Publications and source records attributed to Michaela Aufderheide.

9 recordsLinked to original sources
PubMed2005

Direct exposure methods for testing native atmospheres.

In vitro studies of adverse cellular effects induced by inhalable substances face a number of problems due to the difficulties in exposing cultured cells of the respiratory tract directly to test atmospheres composed of complex gases and particulate compounds. This paper discusses the characteristics of in vitro work and summarizes the use of different in vitro technologies to determine the adverse effects of inhaled pollutants. The exposure of cells to test atmospheres requires accurate control of the pollutant levels, as well as the close contact of cells and gas without interfering with the medium. Systems which rely on the solution of the gas in the medium overlay do not resemble the exposure conditions in vivo, and may not be suitable for studying, for example, the effects of poorly soluble gases. Exposure to gases or complex mixtures can be performed with roller bottles or flasks on rotating and rocking platforms and, using these techniques, the cells are periodically exposed to the test atmosphere. However, the most promising approach is based on a biphasic cell culture technique, where cells are grown on microporous membranes at an air-liquid interface. Here the cells are nutrified from the basal side of the membrane whilst the apical part with the cultivated cells is in direct contact with the test atmosphere. Based on this culture technique, different exposure systems have been developed and these are described and discussed. Exposure of cells from the respiratory tract to gases or particles is responsible for cell injury or cell activation associated with an overexpression of mRNA and the release of bioactive mediators. Therefore, in vitro studies using such a strategy, in combination with relevant and efficient exposure devices, open up new ways to test native complex gases and aerosols. Furthermore, such an experimental approach is not only suitable for cultivated cells, but it can also be used for exposing bacteria to inhalable test compounds. It is possible to analyze the mutagenic potency of in- and outdoor pollutants and several attempts have been made to determine the induction of revertants in a modified Ames assay after exposure to single gases or complex mixtures.

Air Pollutants↗
PubMed2004

A new rat type I-like alveolar epithelial cell line R3/1: bleomycin effects on caveolin expression.

The study of function and regulation of the phenotype of alveolar type I (AT I) epithelial cells is limited by the rareness of suitable cell lines or primary cultures of this cell type. We describe in the present study the type I-like rat epithelial cell line R3/1. This cell line displays in vitro a phenotype with several characteristic features of AT I cells. R3/1 cells were analysed for mRNA and protein content of markers related to the AT I cell type (T1alpha, ICAM-1, connexin-43, caveolins-1 and -2) and AT II phenotypes [surfactant proteins (SPs) A, B, C and D]. The mRNAs for SPs were found to be at a low level. Moderate protein levels for SP-A and SP-B were found, and SP-C and SP-D proteins were not detectable. R3/1 cells are positive for CD44s, E-cadherin, cytokeratin, vimentin and RAGE, and bind the lectins BPA and SBA. For demonstration of the suitability of R3/1 cells for in vitro studies on epithelial injury, the cells were treated with bleomycin. As shown by real-time RT-PCR and immunoblotting, bleomycin-treatment of R3/1 cells resulted in a decrease in mRNA and protein for both caveolin-1 and caveolin-2 in comparison with controls. The AT I-like cell line R3/1 may serve as a promising tool for the study of lung cell biology.

Animals↗
PubMed2004

A modified CULTEX system for the direct exposure of bacteria to inhalable substances.

Increasing attention is being paid to the impact on human health of inhaled gaseous compounds and complex mixtures such as cigarette smoke. The evaluation of the genotoxicity of such materials is mostly based on experiments with model substances or mixtures and condensates in the standard Ames assay. Due to the methodological difficulties of testing air contaminants in their natural gaseous or aerosolised state, there are no generally accepted concepts and techniques for effective exposure of bacteria under such conditions. Therefore, we established a novel experimental approach using an exposure device based on the cell exposure system CULTEX. This allows us to investigate chemically and physically unchanged atmospheres like mainstream cigarette smoke by exposing bacteria of Salmonella typhimurium strains directly on the surface of culture media. The CULTEX exposure device can be connected to gas or aerosol generating systems. The introduction of this exposure device in the field of inhalation genotoxicology offers new test strategies for the in vitro evaluation of a wide range of inhalable substances in both laboratory and ambient situations. A patent was applied for this technical solution.

Air Pollutants↗
PubMed2004

Comparative assessment of toxicities of mainstream smoke from commercial cigarettes.

Three cigarette types were compared using an experimental approach for quantifying selected toxicological effects of diluted fresh whole cigarette mainstream smoke in vitro. The test procedure involved automatic smoking of cigarettes according to the FTC/ISO standard, online monitoring of generated smoke aerosols with respect to particulate and gas-phase components, and direct exposure of a human type II-like lung cell line (A549) using exposure conditions relevant to human smoking. Test specimens were the K1R4F standard research cigarettes (9.2 mg tar/cigarette) and two commercial European light filter cigarettes (brand 1, brand 2) having the same tar content (7.0 mg/cigarette). As a representative of the toxicological effect of smoke, intracellular reduced glutathione was analyzed directly after exposure of cells. Results revealed statistically significant different quantitative effects with regard to glutathione depletion when comparing whole smoke and filtered smoke from all three cigarettes. ED50 values revealed a depletion of reduced glutathione by brand 1 cigarettes that was more than twice the depletion caused by brand 2 cigarettes on a per cigarette basis. Also, quantitatively different effects were found on a per particle and on a per CO concentration basis using whole or filtered smoke from the cigarettes. We conclude that the methods we employed provide sensitive and reproducible ways of detecting differences in the toxicological action of smoke from various types of cigarettes.

Cell Line, Tumor↗
PubMed2003

Novel approaches for studying pulmonary toxicity in vitro.

The in vitro study of adverse cellular effects induced by inhaled pollutants poses a special problem due to the difficulties of exposing cultured cells of the respiratory tract directly to test atmospheres that can include complex gaseous and particulate mixtures. In general, there is no widely accepted in vitro exposure system. However, in vitro methods offer the unique possibility for use of human cells, developed and validated cell culture and exposure device (CULTEX(1)) using the principle of the air/liquid exposure technique. Cells of the respiratory tract are grown on porous membranes in transwell inserts. After removal of the medium, the cells can be treated on their superficial surfaces with the test atmosphere, and at the same time they are supplied with nutrients through the membrane below. In comparison with other experimental approaches, the goal of our studies is to analyze the biological effects of test atmospheres under environmental conditions, i.e. without humidifying the atmosphere or adding additional CO(2). The system used is small and flexible enough independent of a cultivation chamber and thus offers the opportunity for onsite study of indoor and outdoor atmospheres in the field. The efficacy of the exposure device has already been demonstrated in the analysis of dose-dependent cytotoxic and genotoxic effects of exposure of epithelial lung cells to complex mixtures such as native diesel exhaust and side-stream smoke.

Air Pollutants↗
PubMed2003

An improved in vitro model for testing the pulmonary toxicity of complex mixtures such as cigarette smoke.

Numerous approaches have been employed for testing the biological activity of cigarette smoke in vitro. None of them has managed to expose cultured lung cells in a realistic manner to the complex gaseous and particulate mixture that constitutes cigarette smoke. We have devised a system that makes this possible. The system presented here enables the direct exposure of human lung cells to native, unmodified cigarette mainstream smoke. It consists of a smoking machine, a dilution device for the smoke, analytical devices for online monitoring and a specially adapted exposure module based on the Cultex** cell cultivation system that is equipped with a gas-exposure top. Due to the special design of the exposure device and the optimised exposure conditions, this equipment allows cultured human lung cells to be exposed to freshly generated cigarette mainstream smoke. Exploratory experiments revealed that the smoke could be diluted over a wide concentration range in a reproducible way with respect to gas and particulate phases, and also demonstrated reproducible particle deposition depending on smoke concentration. Furthermore, it was shown that the exposed cells maintained their viability. Native cigarette mainstream smoke induced dose-dependent cellular effects in exposed cells with respect to cellular viability (viable cell number monitored by tetrazolium salt cleavage) and intracellular parameters (ATP and glutathione content). Therefore, fresh, physically and chemically unmodified cigarette mainstream smoke can be tested using this novel system.

Aerosols↗
PubMed2003

Exposure of human lung cells to inhalable substances: a novel test strategy involving clean air exposure periods using whole diluted cigarette mainstream smoke.

An experimental approach was established for the validation of an in vitro test system for complex environmental test atmospheres consisting of both gaseous substances and particulates. Smoke from two different cigarette types (generated by an automatic cigarette-smoking machine) was employed to assess both the sensitivity and the specificity of the system. The smoke was diluted with synthetic air and used to expose human lung cells grown on microporous membranes. Cells were exposed alternately to diluted cigarette smoke and pure synthetic air. The effect of diluted smoke was assessed without humidification, addition of CO2, or any other physical or chemical modification of the smoke. The experimental setup included online monitoring of the gas phase (by analysis of CO concentration) and particulate phase (by light-scattering photometry). Replicate experiments confirmed a reproducible generation and dilution of the smoke and a smoke age of about 7 s at the time it came into contact with the cells. Experiments using human lung cells revealed that smoke from the two different cigarette types induced different levels of dose-dependent toxicity. A cell exposure of 6 min using 6 alternating smoke and synthetic air periods was sufficient to cause different effects as measured by intracellular glutathione content. The fact that the system could differentiate between two different types of cigarette smoke demonstrated its high sensitivity and specificity. The system offers new ways to test native complex gaseous and aerosol mixtures in vitro using short exposure times and very small amounts of test substances.

Aerosols↗
PubMedDate not supplied

A method for the in vitro exposure of human cells to environmental and complex gaseous mixtures: application to various types of atmosphere.

The application of in vitro methods to the analysis of the effects of airborne materials is still limited, because there are no generally accepted concepts and technologies for efficiently exposing adherent growing cells to test atmospheres, especially those comprising complex mixtures of gaseous and particulate phases. The introduction of in vitro research into the field of inhalation toxicology offers a unique possibility for using human cells and tissues for pre-screening studies, thus reducing the necessity for animal experiments, and cutting the numbers of animals used in toxicological testing. We therefore developed a novel experimental concept that uses an exposure device based on the cell cultivation system CULTEX (Patent No. DE 198011763; PCT/EP99/00295). This allowed us to investigate environmental atmospheres, which were chemically and physically unmodified, in an in vitro system, by exposing the target cells directly at the air/liquid interface. The exposure device itself is small and flexible enough to be connected to a variety of aerosol-generating systems without the need for an incubator, as it fulfils all the requirements for maintaining cell viability over a defined period. The general applicability and the sensitivity of this in vitro approach for testing various generated atmospheres under the same cell-exposure conditions were demonstrated by studying dose-dependent cytotoxic effects in human lung epithelial cells exposed to air contaminated with single gases or complex mixtures, such as diesel exhaust fumes and side-stream cigarette smoke.

Adenosine↗