Unknown,Transcriptomics,Genomics,Proteomics

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Transcription profiling of rat molecular mechanisms associated with markers of hepatocarcinogenicity during a 14 day dietary study in male Fischer rat

ABSTRACT: To investigate the molecular mechanisms associated with initial dose-related events that are linked to the development of liver tumours: liver growth; cell proliferation; changes in histopathology such as hypertrophy Experiment Overall Design: We selected dose levels of phenobarbital to give a carcinogenic (1000ppm) and two non-carcinogenic doses (50 and 500ppm). This also gave two doses that increased liver weights (500 and 1000ppm) and one that did not (50ppm). Alterations to liver growth and proliferation are complete after 14 days of dietary treatment and so we chose intermediate time points of 1,3 7 and 14 days of treatement. Small sections of the left lateral lobe of the liver were processed to give three biological replicates selected from the 5 treated animals in each group, total RNA purified and mRNA levels measured using Affymetrix Rat Genome 230 v2 microarrays. In addition we also measured a braod range of standard histopathological, clincal chemsitry and metabolomic endpoints.

ORGANISM(S): Rattus norvegicus

SUBMITTER: Richard Currie

PROVIDER: E-GEOD-18753 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:Furan is a widely used industrial chemical and a common contaminant in heated foods. Chronic furan exposure has been shown to cause cholangiocarcinoma and hepatocellular tumors at doses of 2 mg/kg bw/day with gender differences in frequency and severity. The hepatic transcriptional alterations induced by low doses of furan (doses below those inducing liver tumors) and the potential mechanisms underlying gender differences are largely unexplored. We used DNA microarrays to examine the global hepatic mRNA and microRNA transcriptional profiles of male and female rats exposed to 0, 0.03, 0.12, 0.5 or 2 mg/kg bw/day furan over 90 days. Marked gender differences in gene expression responses to furan were observed, with many more altered genes in exposed males than females, confirming the increased sensitivity of males even at the low doses. Pathway analysis supported that key events in furan-induced hepatotoxicity in males included gene expression changes related to oxidative stress, apoptosis and inflammatory response, while pathway changes in females were consistent with primarily adaptive responses (regeneration). Pathway benchmark doses (BMDs) were estimated and compared to relevant apical endpoints. Transcriptional BMDs could be examined in males, they ranged from 0.08 – 1.43 mg/kg bw/day and approximated those derived from traditional histopathology. MiR-34a (a target of P53 signalling) was the only microRNA significantly increased at the 2 mg/kg bw/day, providing evidence in support of the importance of apoptosis and cell proliferation in furan hepatotoxicity. Overall, this study demonstrates the use of transcriptional profiling to discern mode of action and mechanisms involved in gender differences. Total RNAs from 16 liver samples (4 males and 4 females from control and 2 mg/kg bw/day dose groups) were processed using the miRNA Complete Labelling and Hybridization kit (Agilent Technologies Inc.). Labelled RNA was hybridized on 8 X 15K Agilent rat miRNA microarray slides. Arrays were scanned using an Agilent G2505B scanner (5 ?M resolution). Feature extraction (version 10.7.3.1, Agilent Tech. Inc.) was used to acquire the fluorescence intensity of each probe.

Project description:Furan is a widely used industrial chemical and a common contaminant in heated foods. Chronic furan exposure has been shown to cause cholangiocarcinoma and hepatocellular tumors at doses of 2 mg/kg bw/day with gender differences in frequency and severity. The hepatic transcriptional alterations induced by low doses of furan (doses below those inducing liver tumors) and the potential mechanisms underlying gender differences are largely unexplored. We used DNA microarrays to examine the global hepatic mRNA and microRNA transcriptional profiles of male and female rats exposed to 0, 0.03, 0.12, 0.5 or 2 mg/kg bw/day furan over 90 days. Marked gender differences in gene expression responses to furan were observed, with many more altered genes in exposed males than females, confirming the increased sensitivity of males even at the low doses. Pathway analysis supported that key events in furan-induced hepatotoxicity in males included gene expression changes related to oxidative stress, apoptosis and inflammatory response, while pathway changes in females were consistent with primarily adaptive responses (regeneration). Pathway benchmark doses (BMDs) were estimated and compared to relevant apical endpoints. Transcriptional BMDs could be examined in males, they ranged from 0.08 M-bM-^@M-^S 1.43 mg/kg bw/day and approximated those derived from traditional histopathology. MiR-34a (a target of P53 signalling) was the only microRNA significantly increased at the 2 mg/kg bw/day, providing evidence in support of the importance of apoptosis and cell proliferation in furan hepatotoxicity. Overall, this study demonstrates the use of transcriptional profiling to discern mode of action and mechanisms involved in gender differences. A total of 50 RNA samples (5 female and 5 male rats per dose group, 5 dose groups total) were labelled with Cyanine 5-CTP using Low Input Quick Amp Labelling kits (Agilent Technologies Inc.) following the manufacturerM-bM-^@M-^Ys instruction. Universal rat reference total RNA (Agilent Technologies Inc.) was labelled with Cyanine 3-CTP. Cy5-sample cRNA and Cy3-reference cRNA were hybridized to Agilent G4853A SurePrint G3 Rat GE 8 X 60K microarrays (Agilent Technologies Inc.). One microarray from the male group did not pass the quality control and was eliminated from further analyses

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Transcription profiling of rat molecular mechanisms associated with markers of hepatocarcinogenicity during a 14 day dietary study in male Fischer rat

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