Details zur Publikation |
| Kategorie | Textpublikation |
| Referenztyp | Qualifizierungsarbeiten |
| Volltext | Publikationsdokument |
| Titel (primär) | Fate of xenobiotics inside the zebrafish embryo - mass spectrometry imaging for toxicokinetic studies |
| Autor | Halbach, K. |
| Quelle | PhD Dissertation |
| Erscheinungsjahr | 2021 |
| Department | ANA |
| Band/Volume | 8/2021 |
| Seite bis | 246, 10 |
| Sprache | deutsch; englisch |
| Topic | T9 Healthy Planet |
| Keywords | Zebrabärbling; Versuchstiere; Toxizitätstest; Massenspektrometrie; Zebrafish embryo; Quantitative bioimaging; Biotransformation; Bioaccumulation; MS imaging; Toxicokinetics; yolk; Organic anion transporting polypeptide; MALDI-MS imaging; LA-ICP-MS |
| UFZ Bestand | Leipzig, Bibliothek, Berichtsammlung, 00539807, 21-0246 F/E |
| Abstract | The zebrafish embryo (Danio rerio) has developed as a popular model organism in various research fields. In the European Union, the early developmental stages are a promising alternative to animal testing, reducing ethical concerns. One important application is in the risk assessment of chemicals to aquatic organisms in ecotoxicology and drug discovery research. Understanding and predicting toxicokinetic (TK) processes is an essential step towards replacing animal tests with the zebrafish embryo. TK comprises the ADME processes of xenobiotics: absorption, distribution, metabolism, and elimination. Systematic understanding and prediction of the accumulation of xenobiotics in specific tissue and organs in the zebrafish embryo are still missing. It was first aimed to develop a method and data analysis workflow of mass spectrometry (MS) imaging for the zebrafish embryo, taking morphological information into account and focusing on data reproducibility. Secondly, the MS imaging results should be underlined with mechanistic description of the accumulation patterns of the xenobiotics in the zebrafish embryo. MS imaging methods were successfully applied to study TK processes of xenobiotics in the zebrafish embryo. The focus was placed on the two techniques, matrix-assisted laser desorption/ionization-MS (MALDI-MS) imaging and especially laser ablation inductively coupled plasma MS (LA-ICP-MS). For LA-ICP-MS, a workflow consisting of the ablation of whole embryos with carbon as an internal standard was developed. Reproducibility, including biological variation, was shown with Kernel density estimates. Furthermore, the workflow was advanced, presenting the software FishImager. FishImager processes MS imaging data, performs clustering of the data, and imports biological features (such as eyes, yolk) as regions of interest (ROI). The intensities and amounts are visualized in heatmaps making statistical information per ROI and cluster easily visible. FishImager significantly reduces the data analysis time and allows objective data processing to go beyond a subjective visual inspection of the data. It will be a useful tool for zebrafish embryo research. The spatial distribution of xenobiotics was investigated, targeting different physico-chemical and biological processes. MS imaging was combined with bulk analysis and toxicity measurements. A mechanical sample preparation separating the yolk from the rest of the zebrafish embryo (termed the embryonic body) was developed to investigate the sorption of xenobiotics to the yolk and embryonic body. By this, we could demonstrate that five selected neutral chemicals preferentially accumulated in the yolk. The extent of the accumulation increased with increasing hydrophobicity. Looking at internal concentrations in the whole embryo and relating them to toxic effects, e.g., neurotoxicity, would overestimate the target site’s concentration. We could highlight this by studying the spatial distribution of a bromine-containing acetylcholinesterase (AChE) inhibitor. LA-ICP-MS was applied following the bromine distribution as a marker for the compound. The spatial distribution of bromine in the head and spine likely reflected the localization of the tissue rich in AChE. The quantitative imaging significantly correlated with the measured decline in AChE activity in the embryos. Our studies also pointed out that processes other than simple passive diffusions such as metabolism, transporter proteins, and dynamic development are presumably affecting the distribution and steady-state establishment in the zebrafish embryo. More quantitative description is urgently needed. Biotransformation of several of the here selected compounds occurred. Different tissues were shown to likely transform the parent compounds. These are first of all the expected important organs liver and kidney but also the whole gastrointestinal tract and yolk or yolk syncytial layer. We could localize one compound in the middle of the GI tract with MALDI-MS imaging. We confirmed that the organic anion transporter Oatp1d1 is mediating the transport of this compound. But also developmental changes might be a reason for the local accumulation. The presented MS imaging workflows can be applied to more xenobiotics. First mechanistic understanding of tissue-specific accumulations can be extended and particular processes investigated in more depth. This can improve the knowledge and prediction of the ADME processes and increase the application of the model organism, the zebrafish embryo. |
| dauerhafte UFZ-Verlinkung | https://www.ufz.de/index.php?en=20939&ufzPublicationIdentifier=25339 |
| Halbach, K. (2021): Fate of xenobiotics inside the zebrafish embryo - mass spectrometry imaging for toxicokinetic studies Dissertation, Universität Leipzig, Fakultät für Chemie und Mineralogie PhD Dissertation 8/2021 Helmholtz-Zentrum für Umweltforschung - UFZ, Leipzig, 246, 10 pp. |
|