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Determination of genotoxicity by the Comet assay applied to murine precision-cut lung slices

: Switalla, Simone; Knebel, Jan; Ritter, Detlef; Dasenbrock, Clemens; Krug, Norbert; Braun, Armin; Sewald, Katherina


Toxicology in vitro 27 (2013), Nr.2, S.798-803
ISSN: 0887-2333
Fraunhofer ITEM ()
comet assay; EMS; Formalin; precision-cut lung slices

Precision-cut lung slices (PCLSs) are an organotypic lung model that is widely used in pharmacological, physiological, and toxicological studies. Genotoxicity testing, as a pivotal part of early risk assessment, is currently established in vivo in various organs including lung, brain, or liver, and in vitro in cell lines or primary cells. The aim of the present study was to provide the three-dimensional organ culture PCLS as a new ex vivo model for determination of genotoxicity using the Comet assay.
Murine PCLS were exposed to increasing concentrations of ethyl methane sulfonate 'EMS' (0.03-0.4%) and formalin (0.5-5 mM). Tissue was subsequently dissociated, and DNA single-strand breaks were quantified using the Comet assay. Number of viable dissociated lung cells was between 4 × 105 and 6.7 × 105 cells/slice. Even treatment with EMS did not induce toxicity compared to untreated tissue control. As expected, DNA single-strand breaks were increased dose-dependently and significantly after exposure to EMS. Here, tail length rose from 24 µm to 75 µm. In contrast, formalin resulted in a significant induction of DNA cross-links.
The effects induced by EMS and formalin demonstrate the usefulness of PCLS as a new ex vivo lung model for genotoxicity testing in the early risk assessment of airborne substances in the future.