Untersuchungen zum biologischen Effekt inhalierbarer Aerosole in vitro

This bachelor thesis is concerned with a model system for the investigation of biological effects of inhalable aerosols in vitro. This model must show an in-vitro exposure system for test aerosols, a biological test system, a model aerosol as well as analytical methods that are geared to the test system as necessary features. The P.R.I.T. Air/Liquid Interface (ALI) System, which was developed at the Fraunhofer ITEM was used as the exposure system. This system allows a complete separation of a basal medium phase and the apical exposure phase by the use of inserts and cell culture membranes, which are covered with air/lifted-cultivated cells like the water/air barrier in the lung. Human alveolar epithelial cells (A-549) serve as the biological test system. These cells also come into contact with inhalable aerosols like cigarette smoke in the real situation in vivo. The smoke aerosol of a reference cigarette (3R4F) was used as the model aerosol. The cells were exposed to the test aerosol for 1h in a procedure that was based on the smoking behavior of consumers and analyzed immediately or after further incubation for 24h under submerged conditions. The cellular endpoints thereby consider toxicological parameters (Living Cell Count, WST-1), the detection of oxidative stress (oxidized/reduced glutathione) and immunological effects (interleukin-8) that are induced by the smoke aerosol and detectable in the model system. The investigation of different dilutions of the test aerosol allows the establishment of dose-response curves and the determination of EC50 values. Furthermore, a filtration of the smoke aerosol provides the possibility to selectively expose the cells against the gaseous phase of the smoke aerosol and therefore to narrow its toxic components to the cell system. A comparison with the conventional method of testing solvent-extracted cigarette smoke shows that a higher sensitivity can be reached by using the new model system. The aim of these investigations is the establishment of a model system for the in vitro testing of virtually all airborne substances with test cells from different areas of the human lung in combination with the associated analytical methods