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Development of novel in vitro photosafety assays focused on the Keap1–Nrf2–ARE pathway
Author(s) -
TsujitaInoue Kyoko,
Hirota Morihiko,
Atobe Tomomi,
Ashikaga Takao,
Tokura Yoshiki,
Kouzuki Hirokazu
Publication year - 2016
Publication title -
journal of applied toxicology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.784
H-Index - 87
eISSN - 1099-1263
pISSN - 0260-437X
DOI - 10.1002/jat.3234
Subject(s) - phototoxicity , keap1 , in vitro , sensitization , chemistry , cell culture , microbiology and biotechnology , pharmacology , immunology , biochemistry , biology , gene , genetics , transcription factor
Although photoallergens require UV energy for antigen formation, the subsequent immune response is considered to be the same as in ordinary skin sensitization. Therefore, in vitro tests for skin sensitization should also be applicable for photoallergy testing. In this study, we examined whether activation of the Keap1 (Kelch‐like ECH‐associated protein 1)–Nrf2 (nuclear factor‐erythroid 2‐related factor 2)–ARE (antioxidant response element) pathway could be used to assess the photoallergenic potential of chemicals, using the reporter cell line AREc32 or KeratinoSens TM . First, we identified an appropriate UVA irradiation dose [5 J cm –2 irradiation in phosphate‐buffered saline (PBS)] by investigating the effect of UV irradiation on ARE‐dependent gene induction using untreated or 6‐methylcoumarin (6‐MC)‐treated cells. Irradiation of well‐known photoallergens under this condition increased ARE‐dependent gene expression by more than 50% compared with both vehicle and non‐irradiated controls. When the cut‐off value for detecting photoallergens was set at 50% induction, the accuracy of predicting photoallergenic/phototoxic chemicals was 70% in AREc32 cells and 67% in KeratinoSens TM cells, and the specificity was 100% in each case. We designate these assays as a photo‐ARE assay and photo‐KeratinoSens TM , respectively. Our results suggest that activation of the Keap1‐Nrf2‐ARE pathway is an effective biomarker for evaluating both photoallergenic and phototoxic potentials. Either of the above tests might be a useful component of a battery of in vitro tests/ in silico methods for predicting the photoallergenicity and phototoxicity of chemicals. Copyright © 2015 John Wiley & Sons, Ltd.