Lastly, the biological and molecular functions of these genes were explored in IPA. To understand which of the BaP-perturbed biological pathways are directly targeted by differentially expressed miRNA, the results were compared to the biological and molecular functions of those genes that were differentially altered in response to BaP but not identified as targets of any of the miRNA analysed. Serum chemistry was analysed to determine

the hepatic effects of BaP. The results are summarized in Table 1. Administration of 150 or 300 mg/kg BaP for three consecutive days by oral gavage resulted in a small decrease in serum inorganic phosphorous in both treatment groups. A decrease in serum glucose and alkaline phosphatase was seen in either 150 mg/kg

or 300 mg/kg group, respectively, at the 4 h time point. Total protein, uric acid, blood urea nitrogen, albumin and cholesterol did not Enzalutamide datasheet change in any of the groups compared to matched controls. A significant decrease in body weight was found for animals at the time of necropsy (from 24 g to 22.5 g; p < 0.01) but no apparent difference was observed in the specific liver weight for any of the dose groups (data not shown) ( Yauk et al., 2010). The formation of bulky mTOR inhibitor DNA adducts in lung and liver tissues of mice exposed to 150 and 300 mg/kg BaP was analysed by 32P-postlabelling 4 h after the last exposure. Exposure to BaP resulted in an increase in

stable DNA adducts in both lungs and livers in a dose-dependent manner (Table 2). Overall, DNA adduct levels in lungs were similar to the levels observed in liver for both the doses. BaP–DNA adducts were below detection limits in lungs and livers of mice exposed to vehicle control. Exposure to BaP by oral gavage caused a large response in pulmonary mRNA transcription. Approximately 558 and 1267 genes were differentially expressed with a fold change greater than 1.5 and a FDR adjusted p-value ≤ 0.05 in the 150 and 300 mg/kg exposure groups, respectively ( Supplementary Table 1). The complete microarray dataset is available through Calpain the Gene Expression Omnibus at NCBI (http://www.ncbi.nlm.nih.gov/geo/), accession number GSE24751. Hierarchical cluster analysis on differentially expressed genes revealed that samples within a treatment group were clustered ( Supplementary Figure 1), thus, a clear treatment effect was found as a result of exposure to BaP. A large fold induction was observed for a number of genes involved in the metabolism of BaP at both the doses, suggesting that the BaP reached the pulmonary system despite its administration by oral gavage. These genes included Cyp1b1 (25 fold and 50 fold), Cyp1a1 (25 fold and 30 fold), NAD(P)H dehydrogenase, quinone 1 (21 fold) and aryl-hydrocarbon receptor repressor (17 fold and 20 fold) for 150 and 300 mg/kg, respectively.