Improvements of the ALI in vitro testing method for inhalable compounds

In the course of legislation (REACH) and ethical standards the testing of inhalable substances by appropriate in vitro methods is required. In this context, the air-liquid interphase (ALI-) method has emerged as the standard technique for exposure of cell lines, primary cells or co-cultures of the respiratory tract. A currently finished round-robin pre-validation study with model gases revealed the sensitivity and relevance of this method in principle. At the same time, several weak points in the concept concerning robustness, practicability, applicability and efficacy of the method regarding aerosol applications were discovered, which presently prevent an acceptance of this promising method for routine testing. This study aimed on a fundamental improvement of the technology by focusing on two main aspects: (i) to re-design the workflow of the procedure for reduction of cellular stress and experimental complexity and (ii) to clearly improve the deposition of airborne particulate matter on the cellular surface during aerosol testing. As a result, an advanced procedure including a novel type of exposure device was developed. It permits a uniform and smooth processing of cells leading to higher robustness, practicability and ease of use. Repeated dose exposures and less time and material consuming routine use are enabled. To increase the applicability of the ALI-method in aerosol testing, numerical CFD simulations were applied for the optimization of the stagnation flow setup. A combination of different particle deposition mechanisms such as sedimentation, diffusion and thermophoresis was balanced. First results of simulation and laboratory experiments showed a clearly increased deposition efficiency of particles from aerosols in the test system without observation of adverse effects on the exposed cells. Therefore, the improvements of the procedure so far seem promising to further enhance the applicability and acceptance of alternative methods in the study of inhaled substances significantly