Biomedical Engineering Reference
In-Depth Information
8.6.1 Reporter Assays
Enzymes in the zebrafish larval liver and gut including biotin and carboxylase have
been successfully measured and used as a further assessment of liver function. Using
such an enzyme reporter assay, carbamate and merbarone, two mammalian liver
toxicants, have been demonstrated to cause similar organ-specific effect in the
zebrafish as in mice and humans (Zhang et al., 2003).
8.6.2 Liver Histopathology
Histology can be performed on large numbers of zebrafish larvae (Hill et al., 2001) and
hence support for the more gross readouts obtained from other screens can be
achieved relatively simply. However, the main drawback to these investigations is
the time it takes to process and assess the samples. In livermutant studies the change in
liver morphology previously described, namely, the altered size, shape, color, and
texture of the tissue, has been investigated, and at the cellular level degeneration of
hepatocytes, compressed sinusoids, and pooling of erythrocytes have been verified
(Pack et al., 1996). Likewise, larvae exposed to a variety of toxicants have also
exhibited similar changes (unpublished data). In addition, use of TUNEL staining has
also proved useful for the identification of apoptosis (Amali et al., 2006) and specific
conditions such as hepatosteatosis have been visualized in zebrafish using stains such
as Oil Red O that labels fat deposits. For example, when zebrafish are treated with
gamma-hexachlorocyclohexane, fat droplets and fibrosis are induced that phenocopy
the effects seen in mammals (Braunbeck et al., 1990), and similarly after treatment
with thioacetamide, steatosis has also been identified (Amali et al., 2006).
When assessing compound toxicity in adult zebrafish, more traditional mam-
malian techniques can be adapted such as the administration of drugs by injection or
oral intubation. Histopathology would then be a valid option to test for hepatotoxicity
but in addition serum samples can also be taken to measure the levels of certain liver
function enzymes such as alanine transaminase (Murtha et al., 2003).
8.6.3 Biomarkers
With the availability of vast amounts of molecular and genetic information, new
techniques now facilitate analysis of the effects of toxic compounds on a large array of
specific biomarker molecules. In recent years, biomarker approaches for assessing
organ-specific toxicities have been investigated in rodent models (Chisholm and
Dolphin, 1996) and human cells and to a lesser extent in zebrafish. Hepatotoxicity in
particular can benefit from such investigations due to the idiosyncratic nature of many
hepatotoxicants. As well as identifying new mechanisms of toxicity to advance of
understanding of each condition, it is hoped that personalized therapies may also
become possible. In addition, use of a biomarker approach instead of a full
physiological assessment of toxic effects may enable refinement of current toxicity
testing regimes such that animals are dosed with lower amounts of chemicals for a
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