, 1998 and Ohta et al., 2009) or the PPARγ-agonist and human-specific hepatotoxicant troglitazone at physiologically relevant concentrations (Loi et al., 1999 and Yokoi, 2010). The cytotoxicity of the tested drugs was DNA Damage inhibitor assessed by the release of LDH from cells into the media. The amount of viable and metabolically active cells upon drug-treatment was determined via quantitation of ATP (see Materials and methods section). Rat and human 3D liver cells were treated for 1 to 15 days and 2D hepatocyte monolayers for 2 days with

increasing concentrations of fenofibrate (including the human Cmax of 12.4 μM (Table 1, (Vlase et al., 2010)). Fenofibrate induced dose- and time-dependent toxicity in rat 3D liver model (Fig. 4A) as detected by increased LDH

release and decreased ATP levels upon 15 days treatment. Fenofibrate- induced cytotoxicity in the rat 3D liver model was apparent starting from day 8 of chronic drug treatment. However almost no cytotoxicity was detected after 1–2 days of fenofibrate treatment neither in the rat 3D liver model nor in the rat 2D hepatocyte monolayer cultures (Fig. 4A). Fenofibrate decreased the ATP levels by about 20% in rat 2D hepatocyte monolayers after 2 days of treatment and by 80% in rat 3D liver cells after 15 days of treatment (Fig. 4A). In human 3D liver cells and 2D hepatocytes monolayers, fenofibrate did not induce dose- or time-dependent toxicity (Fig. 4A). Next, we treated rat and human 3D liver cells for 8 days and 2D hepatocyte monolayer cultures for 2 days with increasing concentrations of troglitazone GSK-3 activation (including human Cmax of 6.3 μM (Table 1), (Loi et al., 1999)) and measured cytotoxicity and viability of the cells. Troglitazone caused a dose- and time-dependent increase in LDH release and a decrease in ATP levels

in human 3D liver cells but less toxicity was detected in human 2D hepatocyte monolayers (Fig. 4B). Troglitazone induced strong LDH release at physiological ID-8 relevant concentrations already after 1 day of treatment of human 3D liver cells. The LDH release was more pronounced after 1 day than after 8 days treatment at 50–100 μM, indicating an early effect of this drug on human hepatic cells. In contrast to the results obtained in human 3D liver cells, rat 3D liver cultures did not show marked cytotoxicity and no pronounced decrease in cell viability when incubated with similar concentrations of troglitazone. These results are in line with the data from previous studies demonstrating no troglitazone toxicity in rats at physiological relevant concentrations (Fig. 4B, (Li et al., 2002)). However, troglitazone induced strong increase in cytotoxicity and decrease in cell viability in rat 2D hepatocytes after 2 days of treatment (Fig. 4B). We investigated whether human 3D liver models would detect toxicity induced by drugs known to be hepatotoxic in the clinic (Kaplowitz, 2005).