Primary human hepatocytes (hHeps) are still gold standard to perform human drug metabolism studies, but their availability is limited by donor organ scarcity. Therefore, hepatoma cell lines are widely used as alternatives, although their phases I and II drug-metabolizing activities are substantially lower compared with hHeps. The major advantage of these cell lines is immediate availability, standardized culture conditions and unlimited life span. Therefore, the aim of this study was to investigate the drug-metabolizing profile of five human hepatoma cell lines (HepG2, Hep3B, HCC-T, HCC-M and Huh-7) over a culture period of 10 passages. Fluorescent-based assays for seven different cytochrome P450 (CYP) isoforms and seven different phase II enzymes were performed and compared with enzymatic activities of hHeps. CYP activities were much lower in the cell lines (5-15% of hHeps), whereas phase II enzyme activities that are involved in buffering oxidative stress (e.g., Glutathione-S-transferase) reached levels comparable with hHeps. Furthermore, phases I and II enzyme activities in hepatoma cell lines vary strongly during culture time. Interestingly, the most constant results were obtained with Huh-7 cells. Huh-7 cells as well as HCC-T cells exhibited a drug-metabolizing profile closest to hHeps between passages two and four. Toxicity studies with Diclofenac, Paracetamol and Verapamil in both cell lines show dose-response characteristics and EC(50) values similar to hHeps. Therefore, we propose that due to the more consistent results throughout the passages, Huh-7 could be an alternative system to the limitedly available hHeps and frequently used HepG2 cell line in the study of drug metabolism.

ABSTRACT:Epidemiological studies have indicated that exposure of humans to inorganic arsenic in drinking water is associated with the occurrence of bladder cancer. The mechanisms by which arsenic induces this malignancy are still uncertain; however, arsenic metabolites are suspected to play a pivotal role. The aim of the present study was the investigation of uptake capabilities of human urothelial cells (UROtsa) compared with primary human hepatocytes (phH) as well as the intracellular distribution of the arsenic species. Additionally, we were interested in the cyto-and genotoxic potential (comet assay, radical generation) of the different arsenic compounds in these two cell types. Our results show that UROtsa cells accumulate higher amounts of the arsenic species than the phH. Differential centrifugation revealed that the arsenic compounds are preferentially distributed into nuclei and ribosomes. After 24-h exposure, arsenic is mainly found in the ribosomes of UROtsa cells and in the nuclei and mitochondria of phH. In contrast to the pentavalent arsenic species, the trivalent species induced a 4-to 5-fold increase of DNA damage in hepatocytes. Radical generation, measured by thiobarbituric acid reactive substances, was more pronounced in hepatocytes than in urothelial cells. In summary, the uptake of arsenic compounds appears to be highly dependent upon cell type and arsenic species. The nonmethylating urothelial cells accumulate higher amounts of arsenic species than the methylating hepatocytes. However, cyto-and genotoxic effects are more distinct in hepatocytes. Further studies are needed to define the implications of the observed accumulation in cellular organelles for the carcinogenic activity of arsenic.The association between arsenic exposure and urinary bladder cancer, typically transitional cell carcinomas, has been observed in the same endemic areas of the world in which skin cancer populations have been identified (Chiou et al., 1995). In addition to bladder and skin cancer, chronic arsenic exposure causes several malignant and nonmalignant human diseases [for review, see Tseng (2007) (Challenger, 1945) consists of a series of reductions and oxidative methylations. In the sequence of reactions, the ϩ5 oxidative arsenic species are always formed before the analogous ϩ3 arsenic species.Recently, Hayakawa et al. (2005) proposed a new metabolic pathway for arsenic biotransformation in which the ϩ3 arsenic species are formed before the respective ϩ5 species. The trivalent metabolites are oxidized by hydrogen peroxide or other agents to the pentavalent species, which are considered end products of arsenic metabolism.It is generally accepted that the ϩ3 methylated arsenic species are more cyto-and genotoxic (e.g., Styblo et al., 2000;Mass et al., 2001;Aposhian et al., 2003;Kligerman et al., 2003;Dopp et al., 2004) and are more potent enzyme inhibitors (e.g., Styblo et al., 1997aStyblo et al., , 2002Schuliga et al., 2002;Chang et al., 2003) than the pentavalent counThis work was kindly supported by the Ge...

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Hepatocyte transplantation is a practicable and successful treatment option in case of liver insufficiency because implantation of NeoHeps or primary rat hepatocytes had an improving effect on survival. The promising data of the present study warrants further analysis to elucidate the role of NeoHeps in treatment of acute postoperative liver failure to a greater extent.

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