Spec. Issue 2006 - Article Summary

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Current status of the embryonic stem cell test: The use of recent advances in the field of stem cell technology and gene expression analysis

Andrea Seiler, Roland Buesen, Katrin Hayess, Katharina Schlechter, Anke Visan, Elke Genschow, Birgitta Slawik and Horst Spielmann
Federal Institute for Risk Assessment (BfR), National Center for Documentation and Evaluation of Alternative Methods to Animal Experiments (ZEBET), Berlin, Germany
Summary
All guidelines that are currently used for regulatory developmental toxicity testing of chemicals and drugs are based on animal experiments. The most promising alternative is based on embryonic stem cells of the mouse (mESC). Their ability to differentiate into numerous cell types have made ES cells a popular system to study gene function and developmental processes during differentiation in vitro. The embryonic stem cell test (EST) makes use of this capacity to detect developmental toxicants during differentiation of stem cells into cardiomyocytes.
In the present study our investigations were aimed at the further development of the validated EST protocol. We present improvements that focus primarily on (i) the quantitative assessment of drug effects at the cellular level, using a novel approach in which the expression of tissue-specific marker proteins under the influence of the test chemical is quantified by intracellular flow cytometry in ES cells, (ii) the development of protocols for ES cell differentiation into various cell types other than cardiomyocytes, e.g. neural cells, and (iii) the standardisation and optimisation of ES cell culture and differentiation conditions in chemically defined serum-free medium. An important strength of the molecular approach in combination with serum-free culture conditions is that in this way the ability of the test to monitor the cellular response to toxins could be expanded to proteins of many signal transduction pathways in a highly standardised form. Furthermore, these improvements now allow testing of substances known to interact with serum proteins.

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