ZeTox – Toxicology Services

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icon-hepato-tox HepatoTox – Hepatotoxicity Service

Drug-induced liver injury is a major cause of attrition during both the early and late stages of drug development and postmarketing. Zebrafish liver is fully developed and functional by 5 days post fertilization. Moreover, metabolic pathways and detoxification mechanisms are similar to those found in humans. The use of zebrafish larvae for the assessment of compound-induced liver toxicity allows to streamline the process of drug evaluation and reduce the risk of drug attrition in the following phases.

HepatoTox - Hepatotoxicity Service

3Rs-aligned method for high throughput in vivo hepatotoxicity screening.

Advantages

High concordance between zebrafish and mammals after exposure to hepatotoxic and non-hepatotoxic compounds.

Bridge between cell-based and mammalian methods with results in short times.

Simultaneous assessment of cardiotoxicity and neurotoxicity through the service ZeGlobalTox.

Method description

Transgenic zebrafish larvae expressing red fluorescent protein in the hepatocytes are incubated with the No Observable Effect Concentration (NOEC) of the compound of interest from 96 to 120 hours post fertilization (hpf). The fluorescent signal is used to measure the hepatic area. Specific fat staining is later performed to quantify the lipid content in the yolk and in the liver.

Readouts

Evaluation of hepatic morphological and functional parameters at 120 hpf:

  • Liver area: liver necrosis and hepatomegaly.
  • Yolk lipid retention: yolk decreases its size during development due to its consumption. Hepatotoxicity impairs that natural metabolization.
  • Steatosis (fat accumulation in the liver): hepatotoxic compounds promote liver steatosis.
Figure 1. Quantification of liver necrosis. (A) Liver area of zebrafish larvae treated with negative and positive controls and a study compound that shows little effect on the liver size (area). (B) Microscope images of transgenic zebrafish liver expressing RFP in hepatocytes after the exposure of the above-mentioned compounds. 
Figure 2. Evaluation of hepatic steatosis. (A) Proportion of zebrafish larvae that show increased liver fat after treatment with a candidate molecule compared to negative and positive controls. (B) Representative images of healthy zebrafish larva (image on the left) and steatotic zebrafish livers (middle and right images) after specific fat staining.

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References

  1. Cornet C, Calzolari S, Miñana-Prieto R, Dyballa S, van Doornmalen E, Rutjes H, Savy T, D'Amico D, Terriente J. ZeGlobalTox: An Innovative Approach to Address Organ Drug Toxicity Using Zebrafish. Int J Mol Sci. 2017 Apr 19;18(4):864.