In vitro gene expression analysis of hepatotoxic drugs in rat primary hepatocytes

Abstract

The study examined the feasibility of screening for hepatotoxicity by an in vitro gene expression analysis using rat primary hepatocytes and Affymetrix Rat Toxicology U34 arrays. Hepatocytes were exposed for 6 or 24 h to eight drugs, with different mechanisms of hepatotoxicity, at one third of the cytotoxic concentration TC 50 , i.e. acetaminophen, cyclophosphamide, clofibrate, chlorpromazine, lithocholic acid, cisplatin, diclofenac and disulfiram. The types of transcriptional changes observed in this study were generally consistent with previously reported in vivo data, although there were some differences. In hierarchical cluster analysis, drugs formed clusters depending on their mode of toxicity against cells. The number of transcripts affected by the cholestatic hepatotoxicants (lithocholic acid and chlorpromazine) or the drugs that rarely cause of hepatotoxicity (cisplatin, diclofenac and disulfiram) were limited compared with the other drugs (acetaminophen, clobifibrate and cyclophosphamide), where they did not induce transcriptional changes apparently related to toxicity. It is concluded that in vitro gene expression analysis of hepatocytes using microarray is a useful tool for evaluating the toxicological profile of drugs and in screening for the direct toxicity of drugs against hepatocytes.

Transcriptional profiling of reactive metabolites for elucidating toxicological mechanisms: a case study of quinoneimine-forming agents

“…Microarray studies looking a diclofenac-induced hepatic gene expression have been performed in mice (Chung et al, 2006), rat primary hepatocytes (Suzuki et al, 2008), and fish (Oncorhynchus mykiss) (Cuklev et al, 2011). In both mice and fish, numerous transcripts were found to have differential expression.…”

“…The first toxicogenomic study involving APAP was published in 2000 with the intent of identifying an ideal data analysis method to better visualize important transcriptomic profiles in response to toxicant exposure in rats (Cunningham et al, 2000). Subsequently, numerous microarray experiments investigating APAPinduced liver injury have been performed in rats and ratbased models (Elferink et al, 2008;Fukushima et al, 2006;Heinloth et al, 2004;Huang et al, 2008;Irwin et al, 2004;Minami et al, 2005;Morishita et al, 2006;Powell et al, 2006;Suzuki et al, 2008;Zidek et al, 2007). However, recent evidence suggests that toxicologic studies in rats treated with APAP have limited clinical relevance for humans due to mechanistic differences and that mice are the preferred species for such studies .…”

New Models for the In Vitro Study of Liver Toxicity: 3D Culture Systems and the Role of Bioreactors

“…prothrombin, CYP1A1, CYP1A2, CYP2E1, CYP3A, etc.) -Ureagenesis varies according to the model, and may be 6-7 times as in classical 2D culture -High level of hepatic transporters -Fluidic dynamic of some bioreactor favours shear stress and support liver-specific gene expression -Improve microenvironment features -Hepatocytes are much more sensitive to some xenobiotics than in static 2D culture -After optimisation of culture and microenvironment conditions, these models could be used for chronic toxicology necrotic cores: -According to the 3D model, specific hepatic function are maintained -Currently unsuitable for HTS (need the development of adequate methods/devices for increasing sensitivity and reliability, lowering cost and time-consuming features) Evenou et al, 2007Kienhuis et al, 2007Liu Tsang et al, 2007Maguire et al, 2007Meng, 2010Miranda et al, 2010Suzuki et al, 2008Walker & Woodrooffe, 2001 3D co-cultures of liverderived cell types -Longer cell viability, as compared to monotypic 3D culture (up to 57 days) -Sustain some liver-specific function from 3 days to 7 up weeks (albumin, urea secretion, expression of CYP1A1/2,CYP2B1, CYP3A …”

Application of RNA interference to improve mechanistic understanding of omics responses to a hepatotoxic drug in primary rat hepatocytes

“…Toxicogenomics studies are often designed to investigate drug induced liver injury in vivo and in vitro in order to improve mechanistic understanding or to develop predictive models of drug-induced liver injury (Chen et al, 2012;de Longueville et al, 2003;Heinloth et al, 2004;Huang et al, 2004;Kienhuis et al, 2009;Laifenfeld et al, 2014;Suzuki et al, 2008;Uehara et al, 2008;Waring et al, 2001;Zidek et al, 2007). Within the frame of the InnoMed PredTox project, which aimed at improving preclinical safety evaluation by integrating "omics" technologies, transcriptomics studies were conducted to characterize changes in gene expression in rat liver in response to several hepatotoxic drug candidates, with the overall aim of identifying molecular events leading to the observed toxicities Suter et al, 2011).…”