Powdered crf 1

Seven-week-old male wild-type and CARKO mice were divided into 4 groups consisting of 5 mice per group for each genotype. The animals were kept in plastic cages according to group and genotype, under a 12-hr light/dark cycle. The wild-type and CARKO mice in the control group were fed a basal diet (powdered CRF-1, Oriental Yeast Co., Ltd., Tokyo, Japan). The other 3 groups were given PBO at concentrations of 200, 1,000, or 5,000 ppm in the basal diet for 7 days. The highest dose was determined from previous reports in which the dose was sufficient to induce liver hypertrophy and liver tumors in wild-type mice (Fujitani et al., 1993; Takahashi et al., 1997; Sakamoto et al., 2013) .
All mice were examined daily to check for changes in clinical condition. Body weights were measured on days 0 (the first day of dosing), 1, 5, and 7 (the day of termination). Food intake per cage was measured on days 0 to 1 and days 5 to 6, and the values were divided by the number of animals in each cage. At termination of the experiment, all mice were anesthetized by isoflurane inhalation and euthanized by exsanguination. The livers were removed and weighed, then processed for use in subsequent analyses.

Nine-week-old male C57BL/6J mice were obtained from Charles River Japan, Inc., and were fed a normal rodent diet (powdered CRF-1; Oriental Yeast Co. Ltd.) with or without 0.75% BHA for 14 days, as described in a previous report (Banerjee et al., 1995) . At the 13th day of the feeding period, all of the mice were given either TA (100 mg/kg) or saline once intraperitoneally as described in Experiment 1, and necropsied 24 hr later. The livers were fixed in formalin, and histopathological examination including grading of hepatic necrosis and swelling was performed as described in Experiment 1.