HepaRG cells were acquired from Biopredic International (Rennes, France).The detailed process of HepaRG cell culture was described previously (McGill et al., 2011 (link)). HepG2 cells obtained from ATCC (Manassas, VA, USA) were cultured in DMSO (dimethyl sulfoxide)-free Williams’ E medium containing penicillin/streptomycin, insulin and 10% fetal bovine serum, and were 70%~80% confluent before treatment. Both cell lines were treated with 10 mM APAP (Sigma, St. Louis, MO, USA; Lot #36F-7005-1) dissolved in warm Williams’ E medium. ATP (Sigma) was dissolved in saline and added to the cell culture medium to a final concentration of 10 μM or 100 μM. Twenty-four hours after treatment of either ATP and/or APAP, medium and cell fractions were harvested, and cell viability was determined using the lactate dehydrogenase (LDH) assay (Bajt et al., 2004 (link)).
Protocol detail
Hepatocyte cell culture and APAP/ATP treatment
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After treatment
Apap
Assay
Cell
Cell culture
Cell lines
Cell process
Cell viability
Dimethyl sulfoxide
Fetal bovine serum
Hepg2 cells
Insulin
Lactate dehydrogenase
Lot 1
Penicillin
Saline
Streptomycin
Corresponding Organization : University of Kansas Medical Center
Other organizations : Academic Medical Center, University of Amsterdam
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Variable analysis
independent variables
- APAP (10 mM)
- ATP (10 μM or 100 μM)
- Cell viability (determined using the lactate dehydrogenase (LDH) assay)
- HepaRG cell culture conditions (described previously in McGill et al., 2011)
- HepG2 cell culture conditions (DMSO-free Williams' E medium containing penicillin/streptomycin, insulin and 10% fetal bovine serum)
- HepG2 cell confluency (70%~80% confluent before treatment)
- Positive control: Not explicitly mentioned
- Negative control: Not explicitly mentioned
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