TY - JOUR
T1 - Hepatic co-cultures in vitro reveal suitable to detect Nrf2-mediated oxidative stress responses on the bladder carcinogen o-anisidine
AU - Wewering, Franziska
AU - Jouy, Florent
AU - Caliskan, Sükran
AU - Kalkhof, Stefan
AU - von Bergen, Martin
AU - Luch, Andreas
AU - Zellmer, Sebastian
PY - 2017/4/1
Y1 - 2017/4/1
N2 - 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.
AB - 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.
KW - Co-culture
KW - HepG2
KW - Nuclear factor erythroid 2-related factor 2 (Nrf2)
KW - o-Anisidine
KW - ROS
UR - http://www.scopus.com/inward/record.url?scp=85009986023&partnerID=8YFLogxK
U2 - 10.1016/j.tiv.2017.01.006
DO - 10.1016/j.tiv.2017.01.006
M3 - Journal article
AN - SCOPUS:85009986023
SN - 0887-2333
VL - 40
SP - 153
EP - 160
JO - Toxicology in Vitro
JF - Toxicology in Vitro
ER -