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Fish Embryos Possess a Mechanism for Protection against Chemicals

UFZ researchers discover function of protein as transporter of toxic chemicals in the zebrafish embryo

Researchers at the Helmholtz Centre for Environmental Research (UFZ), together with colleagues from the Swiss Eawag aquatic research institute, have discovered a protein which transports chemicals out of the embryo of the zebrafish and in this way protects the embryo against toxic substances.

Images of zebrafish (Danio rerio) embryos illustrating the function of the transporter protein Abcb4 acting as effective chemical defense by keeping out chemical compounds from the embryos. When exposed to the red, fluorescent dye rhodamine B little dye accumulates in the cells and tissues of the fish embryos because Abcb4 prevents uptake of the dye as can be seen in the image above. When transporter function is disrupted by a chemical inhibitor considerably more dye accumulates in the embryo tissues – the embryo tissues appear brighter as the image below shows. Source: Stephan Fischer/Eawag

However, certain environmental chemicals render this protective mechanism ineffective, so that the fish embryos become more sensitive to toxic substances. The study, published in the scientific journal "BMC Biology", could prove to be of great importance for the future assessment of chemicals.

Fish possess a number of different mechanisms for protection against harmful substances in an aquatic environment. These include, for example, molecular transport systems, such as the so called ABC (ATP binding cassette) transporters, which prevent the penetration of toxic substances into cells. ABC transporters have been well investigated for mammals. For fish and their embryos, however, little is known about such transporters.

Ecotoxicologists Dr. Till Luckenbach (UFZ) and Dr. Stephan Fischer (Eawag) together with other colleagues have now found that the transport protein Abcb4 actively extrudes chemicals from the embryo of the zebrafish (Danio rerio). "A fish embryo already has effective protective capabilities", says Luckenbach. "The importance of such transport systems is generally underestimated in toxicological and ecotoxicological research - but they play a very important role."

When a substance is bound to zebrafish Abcb4 , this triggers cleavage of the energy transfer substance adenosine triphosphate (ATP). The energy which this sets free is utilised to expel this substance from the cell. Abcb4 can repel a multitude of different chemical compounds, as a result of which the embryo is resistant to a multitude of toxic substances. In humans the protein ABCB1 serves this function. Thus, it came as a surprise that it is Abcb4 that in zebrafish acts as "multidrug" or "multixenobiotic" transporter. The homologous ABCB4 of humans, by contrast, is incapable of transporting toxic chemical compounds. Instead, human ABCB4 has a specific function in liver where it channels certain fatty acids into the bile ducts in order to protect the liver cells against aggressive biliary acids.

In experiments with zebrafish embryos in which the expression of Abcb4 protein was suppressed, Luckenbach and his team found that the embryos were much more sensitive to toxic chemicals and that these substances were enriched to a greater extent in the embryo tissue. "Based on this data we concluded that the zebrafish Abcb4 protects the embryo against the toxic impact of chemicals by keeping them out", says Luckenbach.

In follow-on investigations the researchers measured the activity of the transport system, enabling the identification of the chemicals which Abcb4 transports. However, certain substances block the transport mechanism. This inhibition renders its function ineffective, and other harmful substances can penetrate into the organism. "Compounds which inhibit the transporter throw open the doorway for other toxic substances ", says Stephan Fischer." These are also referred to as chemosensitizers, as they make the organism more sensitive to harmful chemicals. This indirect toxic effect plays an important role, above all in mixtures of substances such as frequently found in our environment."

At the UFZ a wide range of environmentally relevant chemicals are currently being investigated for their impact on the Abcb4 transporter system - separately and in mixtures. Luckenbach: "Many effects of substance mixtures may be explained on the basis of the zebrafish Abcb4 protein activity. Zebrafish embryos are used for the assessment of chemicals and for investigations of environmental impact, so we hope that in future our study will contribute to an awareness of the need to incorporate Abcb4 transport processes in toxicological testing directives." Nicole Silbermann

Abcb4 acts as multixenobiotic transporter and active barrier against chemical uptake in zebrafish (Danio rerio) embryos
Stephan Fischer, Nils Klüver, Kathleen Burkhardt-Medicke, Mirko Pietsch, Anne-Marie Schmidt, Peggy Wellner, Kristin Schirmer and Till Luckenbach
BMC Biology 2013, 11:69doi:10.1186/1741-7007-11-69
Partial funding was provided for this work by the Deutsche Forschungsgemeinschaft (DFG), the Ministry of Environment, Agriculture and Geology of Saxony and the the German Federal Environmental Foundation (DBU).
Further information:
Dr. Till Luckenbach
Helmholtz Centre for Environmental Research (UFZ)
Telephone: 0341-235-1514
Dr. Stephan Fischer
Eawag / Aquatic Research Institute of the Swiss Federal Institute of Technology
Telephone: +41 (0)58 765 55 67
Tilo Arnhold / Susanne Hufe (UFZ Press Office)
Telephone: 0341-235-1635, -1630
At Helmholtz Centre for Environmental Research (UFZ), scientists research the causes and ramifications of far-reaching changes in the environment. They focus on water resources, biological diversity, the consequences of climate change and means of adaptation, environmental and bio-technologies, bio-energy, how chemicals behave in the environment, their repercussions on health, modelling and questions of social sciences. Their leitmotif: our research serves the sustainable use of natural resources and, under the influence of climate change, helps safeguard these foundations of life for posterity. UFZ employs a staff of 1,100 employees at its sites in Leipzig, Halle and Magdeburg. Its funding comes from the Federal Government and the states of Saxony and Saxony Anhalt.

The Helmholtz Association contributes to solving major and urgent questions within society, sciences and industry through the provision of academic excellence in six fields of research: energy, the earth and environment, health, key technologies, structure of matter and aviation, aerospace and transport. Employing a staff of 35,000 in 18 research centres and equipped with an annual budget in the region of 3.8 billion euro, the Helmholtz Association is Germany's largest scientific organisation. Its work stands in the tradition of the outstanding natural scientist Hermann von Helmholtz (1821-1894).

Nicole Silbermann/Tilo Arnhold | UFZ News
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