Hepatic co-cultures in vitro reveal suitable to detect Nrf2-mediated oxidative stress responses on the bladder carcinogen o-anisidine

Franziska Wewering*, Florent Jouy, Sükran Caliskan, Stefan Kalkhof, Martin von Bergen, Andreas Luch, Sebastian Zellmer

*Corresponding author for this work

Research output: Contribution to journalJournal articleResearchpeer-review

4 Citations (Scopus)

Abstract

The azo dye o-anisidine is known as an industrial and environmental pollutant. Metabolites of o-anisidine remain in the liver for > 24 h. However, the toxicological impact of o-anisidine on the liver and its individual cell types, e.g., hepatocytes and immune cells, is currently poorly understood. A novel co-culture system, composed of HepG2 or Huh-7 cells, and differentiated THP-1 cells was used to study the metabolic capacity towards o-anisidine, and compared to primary murine hepatocytes which express high enzyme activities. As model compounds the carcinogenic arylamine o-anisidine and its non-carcinogenic isomer, p-anisidine, as well as caffeine were used. Global proteome analysis revealed an activation of eIF2 and Nrf2-mediated oxidative stress response pathways only in co-cultures after treatment with o-anisidine. This was confirmed via detection of reactive oxygen species. In addition, the mitochondrial membrane potential decreased already after 3 h treatment of cells, which correlated with a decrease of ATP levels (R2 > 0.92). In the supernatant of co-cultured, but not single-cultured HepG2 and Huh-7 cells, o-anisidine caused increases of damage-associated proteins, such as HMGB1 (high mobility group box-1) protein. In summary, only co-cultures of HepG2 and THP-1 cells predict o-anisidine induced stress responsive pathways, since the system has a higher sensitivity compared to single cultured cells.

Original languageEnglish
JournalToxicology in Vitro
Volume40
Pages (from-to)153-160
Number of pages8
ISSN0887-2333
DOIs
Publication statusPublished - 1 Apr 2017

Keywords

  • Co-culture
  • HepG2
  • Nuclear factor erythroid 2-related factor 2 (Nrf2)
  • o-Anisidine
  • ROS

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