Complex skin models and impedance spectroscopy as new tools for hazard identification and efficacy testing
Despite advances in the development of in-vitro-tissue-models such as reconstructed human epidermis (RHE), the number of endpoints in toxicity-testing, which can be addressed with these models, is limited. This is due to a lack of key cellular components and a restricted live time of the models. In addition, the analysis of the models is still dependent on invasive methods such as histological processing or 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) staining. To overcome these pitfalls, we achieved advanced culture systems and biomaterials which allow long term culture of complex tissue-equivalents. Using these technologies, we have developed the first full thickness skin-model with a perfused vascular network. Furthermore, as an alternative for destructive methods, we have established a non-destructive technology to analyze the integrity of the epidermal barrier based on impedance spectroscopy. RHE typically exhibits characteristic impedance spectra in a frequency ranging between 1 Hz and 100 kHz, which is comparable to the spectra of freshly isolated human epidermal biopsies. From these spectra, we extracted electrical parameters of the RHE such as the capacitance and the ohmic resistance. These parameters change significantly during epidermal differentiation and were used to quantify the effects of mechanical and chemical disruption of the epidermal integrity. Most relevant, impedance spectroscopy shows a sufficient sensitivity to detect a transient decreased ohmic resistance caused by 2-propanol, which is classified as a non-irritant by MTT assays. This result indicates that impedance spectroscopy can be employed as an additional method to assess mild irritative effects. In our work we could create new technologies for the generation and analysis of tissue-models which is a vital requirement to increase the success of in-vitro-test-methods.