Potential of predictive biomarkers for in vitro skin corrosion tests

To perform successful in vitro skin corrosion tests validated human 3D test systems and adequate predictive biomarkers are necessary. In this study a human full-thickness skin equivalent developed by Fraunhofer-IGB and excised human skin were compared as in vitro models. 4 OECD substances according to guideline 431 were applied to both tissues to determine interleukin release patterns and protein synthesis profiles. 2-tert-Butylphenol and 1,2-Diaminopropane served as corrosive agents, 4-Amino-1,2,4-triazol as a skin irritant and Eugenol as a sensitizing agent. Medium was collected to perform IL-1?, IL-6 and IL-8 analysis by ELISA, supported by WST-1 viability-assay, histological and immunohistological investigations. To detect cytochrome P450 protein (CYP) and heat shock protein 60 (Hsp60) expression Western blots were performed. Different levels of epidermal barrier damage could be shown for the 4 substances by histological staining. All substances caused viability reduction of 90% in the skin equivalent. All chemicals led to a significant upregulation of the measured interleukins. In the presence of the tested irritants, aberrant cytokine release profiles could be observed compared to the sensitizing agent. Induction of CYP isoform 3A4 was found for corrosive Butylphenol, Diaminopropane and Aminotriazol, not for sensitising Eugenol. An upregulation of stress marker Hsp60 could be observed for all test substances. Therefore, CYP isozymes and cytokine release pattern could be relevant indicators to discriminate between corrosive, irritant and sensitizing potentials of chemicals.