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Comparative Study
. 2017 Nov 1;160(1):95-110.
doi: 10.1093/toxsci/kfx165.

Editor's Highlight: Comparative Dose-Response Analysis of Liver and Kidney Transcriptomic Effects of Trichloroethylene and Tetrachloroethylene in B6C3F1 Mouse

Yi-Hui Zhou  1   2 , Joseph A Cichocki  3 , Valerie Y Soldatow  4 , Elizabeth H Scholl  2 , Paul J Gallins  2 , Dereje Jima  2 , Hong-Sik Yoo  4 , Weihsueh A Chiu  3 , Fred A Wright  1   2   5 , Ivan Rusyn  3
Affiliations
Comparative Study

Editor's Highlight: Comparative Dose-Response Analysis of Liver and Kidney Transcriptomic Effects of Trichloroethylene and Tetrachloroethylene in B6C3F1 Mouse

Yi-Hui Zhou et al. Toxicol Sci. .

Abstract

Trichloroethylene (TCE) and tetrachloroethylene (PCE) are ubiquitous environmental contaminants and occupational health hazards. Recent health assessments of these agents identified several critical data gaps, including lack of comparative analysis of their effects. This study examined liver and kidney effects of TCE and PCE in a dose-response study design. Equimolar doses of TCE (24, 80, 240, and 800 mg/kg) or PCE (30, 100, 300, and 1000 mg/kg) were administered by gavage in aqueous vehicle to male B6C3F1/J mice. Tissues were collected 24 h after exposure. Trichloroacetic acid (TCA), a major oxidative metabolite of both compounds, was measured and RNA sequencing was performed. PCE had a stronger effect on liver and kidney transcriptomes, as well as greater concentrations of TCA. Most dose-responsive pathways were common among chemicals/tissues, with the strongest effect on peroxisomal β-oxidation. Effects on liver and kidney mitochondria-related pathways were notably unique to PCE. We performed dose-response modeling of the transcriptomic data and compared the resulting points of departure (PODs) to those for apical endpoints derived from long-term studies with these chemicals in rats, mice, and humans, converting to human equivalent doses using tissue-specific dosimetry models. Tissue-specific acute transcriptional effects of TCE and PCE occurred at human equivalent doses comparable to those for apical effects. These data are relevant for human health assessments of TCE and PCE as they provide data for dose-response analysis of the toxicity mechanisms. Additionally, they provide further evidence that transcriptomic data can be useful surrogates for in vivo PODs, especially when toxicokinetic differences are taken into account.

Keywords: agents; dose-response; kidney; liver; methods; risk assessment; systems toxicology; toxicogenomics; volatile organic compounds.

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Figures

Figure 1
Figure 1
Liver (A and B) and kidney (C and D) levels of trichloroacetic acid (TCA) measured in male B6C3F1 mice 24 h following oral gavage with equimolar doses of trichloroethylene (TCE, top row) or tetrachloroethylene (PCE, bottom row) in aqueous vehicle (5% Alkamuls EL-620 in saline). Spearman (ρ) correlation coefficients and corresponding significance (P) values are shown for each dose-response relationship.
Figure 2
Figure 2
Dose-dependent effects of TCE or PCE on mouse liver (A and B) and kidney (C and D) gene expression. Plots of logistic dose-response modeling of normalized gene expression values for each animal, with individual data shown as red (for TCE) and green (for PCE) dots and respective curve fits colored accordingly are shown. Representative genes are shown. A and C, Cyp4a14 is significantly upregulated by both TCE and PCE in both liver and kidney. B and D, Alox15 is upregulated by TCE in liver only but not PCE.
Figure 3
Figure 3
Correlation analysis of dose-response in gene expression in mouse liver (A–C) and kidney (D–F) following treatment with TCE or PCE. Plotted are −Log10(P values) for dose-responsive differential gene expression analysis, right and top of axes origin are genes with positive association, left and bottom of axes origin are genes with negative association. Dots are individual genes and the color scheme is as follows: red (genes that are significant with false discovery rate (FDR) q < 0.05 in TCE but not PCE), green (q < 0.05 in PCE but not TCE), blue (significant with q < 0.05 for both TCE and PCE), and black (not significant for both TCE and PCE). A, Genes significantly correlated with the administered dose of either PCE or TCE. B, Genes significantly correlated with both TCE dose and liver TCA levels. C, Genes significantly correlated with both PCE dose and liver TCA levels. Panels D–F are same as A–C but for kidney. Pearson (r) correlation coefficients are shown for each plot.
Figure 4
Figure 4
Correlation analysis of gene expression responses between mouse liver and kidney following treatment with TCE (A) or PCE (B). Plotted are −log10(P values) for dose-responsive differential gene expression analysis (compared with TCE or PCE dose). Colors and directionality in effects are same as in the legend to Figure 3. Pearson (r) correlation coefficients are shown for each correlation.
Figure 5
Figure 5
Analysis of the transcriptional effects of TCE (800 mg/kg or 6 mmol/kg) and PCE (1000 mg/kg or 6 mmol/kg) in mouse liver (A–C) and kidney (D–F). Shown are genes that were significantly (FDR q < 0.05) up- or downregulated by treatment with either chemical. A and D, Genes that were significant for both TCE and PCE in liver (n = 54, only genes with official gene symbols counted) or kidney (n = 16, only genes with official gene symbols counted). Maximum fold-change in gene expression, converted to log2 values, is plotted for each gene. Regression analysis slope, correlation coefficients and significance are shown. Select genes are highlighted. B and C, Histograms of log2 maximum fold-change values for genes that are significant in liver either for TCE (n = 97) or PCE (n = 545). E and F, Histograms of log2 maximum fold-change values for genes that are significant in kidney either for TCE (n = 12) or PCE (n = 290). Lists of genes shown in each panel are provided as Supplementary Table 12.
Figure 6
Figure 6
Association of Cyp2c29 exon “usage” proportion with TCE dose (P = 2.1 × 10−5, q value = 0.18). The result is largely driven by reads mapping to exon 5, for which the exon-specific usage was highly significant (P = 2.6 × 10−6) while for other exons it was not significant.
Figure 7
Figure 7
Dose-response analysis of the transcriptional and apical effects of TCE and PCE in mouse liver and kidney. A, Point of departure (POD) (median BMDL) for pathways (Kyoto Encyclopedia of Genes and Genomes [KEGG] or Reactome) that were significantly perturbed (Fisher’s exact 2-tailed P < 0.05) by treatment with TCE (red) or PCE (green) in mouse liver (circles) and kidney (squares). Open symbols are KEGG pathways and closed symbols are Reactome pathways. A complete list of pathways and associated dose-response PODs are included as Supplementary Table 19. Select common pathways are labeled. B, Comparison of the apical POD (noncancer endpoints are black vertical bars, cancer endpoints are blue vertical bars) and transcriptomic POD ranges for all pathways (red box and whisker plots showing distribution of median pathway BMDLs) and selected pathways (individual red vertical lines showing individual median pathway BMDLs) after treatment with TCE or PCE. Complete details of the apical endpoint types, studies from which they were derived, and PODs for the pathways are available in Supplementary Tables 19 and 20. To enable direct comparison of the apical and transcriptional PODs to human exposure, all doses were converted to human equivalent doses using the following metrics: liver oxidative metabolism (for TCE and PCE Liver), GSH conjugation metabolism (for TCE Kidney), and PCE area under the curve (for PCE Kidney). C, Relationship between transcriptional and apical PODs converted to human equivalent doses. Symbols and error bars are the geometric means and ranges, respectively, of the median transcriptional BMDLs plotted against the corresponding geometric means and range of the apical PODs, from each panel in (B). Symbol shapes and colors represent different treatment (PCE or TCE), target tissue (kidney or liver), and type of apical POD (noncancer or cancer) as shown in the legend of panel C. Blue dotted lines are ± 1 order of magnitude deviation from perfect correspondence.
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