Gene expression profiling reveals indicators of potential adverse effects

The study presented here tested the hypothesis that gene expression analysis after exposure to sub-toxic doses would allow prediction of adverse effects that only become manifest after exposure to toxic doses. Therefore

Figure 2.2.

Electron micrographs of centrilobular hepatocytes. (A) Hepatocyte from control animal. (B) Hepatocyte from animal after treatment with 150 mg kg-1 APAP. Arrows point to mitochondria.

we treated rats with sub-toxic (50 and 150 mg kg-1) and toxic (1500 mg kg-1) doses of APAP and performed microarray analysis on total RNA isolated from livers of those animals. As demonstrated, the observed gene expression changes indicated cellular energy depletion at doses of APAP that did not cause any traditional manifestations of toxicity. Ultrastructural studies

Microarray hybridization scheme

Pooled control RNA sample

Time-matched vehicle treated control animals

Time-matched vehicle treated control animals

Differentially regulated genes

Hypothesis: Gene expression analysis indicates mitochondrial damage with subsequent ATP depletion of liver tissue

Measurement of ATP levels

Electron microscopy

revealed mitochondrial damage after exposure to 150 mg kg-1, but not to 50 mg kg-1 APAP, although gene expression analysis suggested some modest cellular energy loss even after exposure to 50 mg kg-1 APAP. This indicates that gene expression changes are very sensitive markers of cellular stress, even in the absence of any apparent phenotypic changes (8).

Our results have great implications for future toxicological research. On the one hand, the prediction of toxicity based on, in the traditional sense, sub-toxic exposure levels has great potential for identification of compounds that have toxicological potential. Comparison of gene expression profiles retrieved from unknown toxicants against extensive toxicogenomics databases might reveal toxic potential of those compounds - and save the effort of doing prolonged dose-finding studies to establish phenotypic anchors. This is even more important in the case of novel and genetically engineered compounds for which limited amounts may be available.

On the other hand, with the advent of toxicogenomics, it is becoming apparent that slight alterations in the environment will manifest themselves in gene expression changes compared with sham-treated controls. The challenge is to determine which of those changes are truly meaningful and indicative of potential harm and which are pharmacological, adaptive, or confounding events - related to animal housing, animal handling, feeding and the circadian cycle.

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