Modelling of the "toxome" of cultivated human hepatocytes

12/2013-11/2015

This third-party funded project is conducted in the framework of the BfR research program for modern methods in toxicology.

Support code of the Federal Ministry of Education and Research: FKZ 3R-474-007

The LivSys joint research project, funded by the Ministry of Education and Research, aims to predict hepatotoxicity based on in vitro systems and mathematical modelling.. To reach this goal, a combination of transcriptomics and simulation techniques (including PBPK and spatio-temporal modelling) is used by the partner research groups from BfR and the Technical University of Dortmund under the lead of the Leibniz Research Centre for Working Environment and Human Factors.

It has been well established that upon exposure to toxic compounds hepatocytes show patterns of gene signatures, which relate to ‘stress response pathways’ in order to re-establish cellular homeostasis. Using ‘omics’ data it is possible to analyze if and to which extent ‘stress response pathways’ are activated. Moreover, the pattern and intensity of activated stress pathways provides opportunities to derive possible mechanisms of toxicity. The current project is based on a well established database of hepatotoxic compounds tested in human hepatocytes by Affymetrix gene arrays. Exploiting and modeling these data, activation of specific stress pathways can be analyzed and evaluated, which ones may be useful as alerts for toxicological hazard evaluation.

Since it cannot be expected that hepatocyte in vitro systems maintain all mechanisms relevant for hepatotoxicity in vivo, it is critical to know which mechanisms still function in an in vivo-like manner and which should be interpreted with caution. Therefore, the objectives of this project are:

• to establish a model of the ‘human toxome’ of the liver. This will include the entity of all known mechanisms (‘stress response pathways’) relevant for hepatotoxicity. The pathways (including feedback loops, thereby forming a network) will be presented as a reaction network in agreement with recently defined standards of mathematical models in systems biology.
• to identify which ‘pathways’ (or parts of the network) of the toxome are still functional and which are compromised (and if compromised to which degree) in 3D collagen sandwich cultured human hepatocytes. The required information will be obtained by analysis of our already established ‘omics’ database, by functional analyses and by in vitro/in vivo comparisons.
• to identify the minimally required set of biomarkers (e.g. mRNA species measurable by qRT-PCR or proteins that can be measured by ELISA) to quantify the degree of activity of individual ‘stress pathways’ and to define thresholds when a result is considered as significant
• to characterize the most frequently used human liver cells (cell lines) using the novel systems biology approach of the ‘human toxome’. This analysis will include HepG2 and HepaRG cells.

The expected delivery of this project will be an analysis of ‘stress response pathways’ still active in human hepatocyte in vitro systems in an in vivo-like manner and can therefore be utilized as descriptors for hazard evaluation of compounds.

References

• Heise T, Schug M, Storm D, Ellinger-Ziegelbauer H, Ahr H. J, Hellwig B, Rahnenführer J, Ghallab A, Guenther G, Sisnaiske J, Reif R, Godoy P, Mielke H, Gundert-Remy U, Lampen A, Oberemm A, Hengstler J G (2012) In Vitro - In Vivo Correlation of Gene Expression Alterations Induced by Liver Carcinogens. Curr. Med. Chem. 19, 1721-1730
• Grinberg M, Stöber RM, Edlund K, Rempel E, Godoy P, Reif R, Widera A, Madjar K, Schmidt-Heck W, Marchan R, Sachinidis A, Spitkovsky D, Hescheler J, Carmo H, Arbo MD, van de Water B, Wink S, Vinken M, Rogiers V, Escher S, Hardy B, Mitic D, Myatt G, Waldmann T, Mardinoglu A, Damm G, Seehofer D, Nüssler A, Weiss TS, Oberemm A, Lampen A, Schaap MM, Luijten M, van Steeg H, Thasler WE, Kleinjans JC, Stierum RH, Leist M, Rahnenführer J, Hengstler JG (2014) Toxicogenomics directory of chemically exposed human hepatocytes. Arch. Toxicol. 12, 2261-2287