Academic journal article Environmental Health Perspectives

Gene Expression Analysis of the Hepatotoxicant Methapyrilene in Primary Rat Hepatocytes: An Interlaboratory Study

Academic journal article Environmental Health Perspectives

Gene Expression Analysis of the Hepatotoxicant Methapyrilene in Primary Rat Hepatocytes: An Interlaboratory Study

Article excerpt

Genomics technologies are used in several disciplines, including toxicology. However, these technologies are relatively new, and their applications require further investigations. When investigators apply these technologies to in vitro experiments, two major issues need to be clarified: a) can in vitro toxicity studies, in combination with genomics analyses, be used to predict the toxicity of a compound; and b) are the generated toxicogenomics data reproducible between laboratories? These questions were addressed by an interlaboratory study with laboratories of four pharmaceutical companies. We evaluated gene expression patterns from cultured rat primary hepatocytes after a 24-hr incubation with methapyrilene (MP). Extensive data analysis showed that comparison of genomics data from different sources is complex because both experimental and statistical variability are important confounding factors. However, appropriate statistical tools allowed us to use gene expression profiles to distinguish high-dose-treated cells from vehicle-treated cells. Moreover, we correctly identified MP in an independently generated in vitro database, underlining that in vitro toxicogenomics could be a predictive tool for toxicity. From a mechanistic point of view, despite the observed site-to-site variability, there was good concordance regarding the affected biologic processes. Several subsets of regulated genes were obtained by analyzing the data sets with one method or using different statistical analysis methods. The identified genes are involved in cellular processes that are associated to the exposure of primary hepatocytes to MP. Whether they are specific for MP and are cause or consequence of the toxicity requires further investigations. Key words: hepatotoxicity, interlaboratory study, methapyrilene, microarray, rat hepatocytes, toxicogenomics. Environ Health Perspect 114:92-99 (2006). doi: 10.1289/ehp.7915 available via http://dx.doi.org/[Online 12 August 2005]

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In the last decade, genomic technologies have become gradually integrated into several phases of drug development. In the field of toxicology, drug safety laboratories have begun to use these technologies to assist research to conduct toxicity evaluations on as many potential lead compounds as feasible and to gain a better understanding of the mechanisms of toxicities. For investigators to be successful in the selection of compounds most likely to succeed during preclinical development, the methods they use should have a medium throughput, a short turnaround time, a good predictivity, and be reproducible.

In vitro systems are being used in toxicology studies to determine several kinds of toxicities. Mouse lymphoma cells, primary rat hepatocytes, and human lymphocytes are among the mammalian cell systems used to determine mutagenicity (Kilbey et al. 1984). Primary rat or human hepatocytes are used to determine cytotoxicity as well as metabolism of compounds or their ability to induce cytochrome P450 genes (Gomez-Lechon et al. 1988; Paillard et al. 1999). However, only a few laboratories have investigated whether in vitro systems can be used in the toxicogenomics evaluation of development compounds. Harries et al. (2001) used the human liver HepG2 cell line to investigate gene expression changes of two hepatotoxins. The results strongly suggested that different mechanisms of hepatotoxicity may be associated with specific markers of gene expression. Waring et al. (2001) showed that gene expression profiles for compounds with similar mechanisms of toxicity tested in vitro on primary rat hepatocytes formed clusters, suggesting a similar effect on transcription. Conversely, Boess et al. (2003) characterized several hepatic in vitro systems on the basis of gene expression profiling and concluded that the results were poorly comparable with the in vivo outcome, depending on the cell culture system used. It is therefore essential to obtain more knowledge on the in vitro system used to achieve better understanding and interpretation of genomics data. …

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