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Effect signatures in zebrafish embryos exposed to compounds with potential developmental toxicity
Poster Abstracts / Reproductive Toxicology 72 (2017) 31–52
P-39 Human embryonic stem cells as a model for paracetamol induced epigenetic deregulation and neurotoxicity Spildrejorde 1,7,∗ ,
Falck 2,7,∗ ,
Mari Martin Magnus Leithaug 1,7 , Kristina Gervin 4,7 , Hege B. Fjerdingstad 3 , Hedvig Nordeng 4,5,7 , Ragnhild E. Paulsen 6,7 , Ragnhild Eskeland 2,3,7 , Robert Lyle 1,7
49
[3] A.B. Csoka, M. Szyf, Epigenetic side-effects of common pharmaceuticals: a potential new field in medicine and pharmacology, Med. Hypotheses 73 (5) (2009) 770–780.
http://dx.doi.org/10.1016/j.reprotox.2017.06.178 P-40 Effect signatures in zebrafish embryos exposed to compounds with potential developmental toxicity
1
Department of Medical Genetics, Oslo University Hospital, Norway 2 Department of Biosciences, University of Oslo, Norway 3 Norwegian Center for Stem Cell Research, Department of Immunology, Oslo University Hospital, Norway 4 Pharmacoepidemiology and Drug Safety Research Group, Department of Pharmacy, School of Pharmacy, University of Oslo, Norway 5 Department of Child Health, Norwegian Institute of Public Health, Oslo, Norway 6 Department of Pharmaceutical Biosciences, University of Oslo, Norway 7 PharmaTox Strategic Research Initiative, Faculty of Mathematics and Natural Sciences, University of Oslo, Norway Recent cohort studies have suggested an association between paracetamol usage during pregnancy and risk of neurodevelopmental disorders (e.g. attention deficit hyperactivity disorder, ADHD) in the child [1,2]. The mechanism by which paracetamol modulate the increased risk of developing ADHD is currently unknown, and we hypothesize that paracetamol is involved in changing the epigenetic landscape during the child’s neurodevelopment. Perinatal development is a critical time for the establishment of epigenetic patterns and some commonly used medications during pregnancy, such as analgesics, antidepressants and anticonvulsants, have previously been shown to alter epigenetic homeostasis [3]. A recent unpublished study from PharmaTox indicates an association between epigenetic changes in cord blood, long-term paracetamol exposure during pregnancy and ADHD diagnosis in the offspring. To study the mechanisms by which neonatal exposure to medications might affect foetal brain development, we use human embryonic stem cells (hESCs) as a model for screening of paracetamol induced neurotoxicity and the impact on the epigenetic landscape. We measured cell viability and oxidative stress level of hESC and derived neural progenitor cells upon 24hour exposure to physiological concentrations of paracetamol or its metabolite AM404 and controls. Furthermore, global transcription and DNA methylation profiles of untreated and paracetamol treated hESCs were characterised by RNA sequencing and wholegenome bisulphite analysis, respectively. Our preliminary results indicate that hESCs is a suitable model to evaluate paracetamol toxicity in early human development. Furthermore, this hESC model can potentially be used to screen other pharmaceuticals commonly used during pregnancy.
References [1] R.E. Brandlistuen, et al., Prenatal paracetamol exposure and child neurodevelopment: a sibling-controlled cohort study, Int. J. Epidemiol. 42 (6) (2013) 1702–1713. [2] J.M.D. Thompson, et al., Associations between acetaminophen use during pregnancy and ADHD symptoms measured at ages 7 and 11 years, PLoS ONE 9 (9) (2014) pe108210.
Elisabet Teixido 1,∗ , Tobias Kießling 1,2 , David Leuthold 1 , Stefan Scholz 1 1 UFZ – Helmholtz Centre for Environmental Research, Department Bioanalytical Ecotoxicology, Leipzig, Germany 2 Scientific Software Solutions, Leipzig, Germany
The zebrafish embryo model is especially suited for the screening of of chemicals for developmental toxicity. Because the embryo develops in an integrated manner, organizing and differentiating spatially, temporally and in a hierarchical manner, disturbances are anticipated to result in nonrandom patterns of anomalies. Moreover, it has been shown that developmental cascade effects triggered by chemical exposure can be recapitulated by analyzing the relationships among endpoints [1]. The present study was designed to study the relationships among different quantitative endpoints and chemical treatments to determine chemical induced phenotypic signatures in the zebrafish embryo. Images of zebrafish larvae were collected using the Vertebrate Automated Screening Technology (VAST Bioimager) to handle and proper orient embryos. To detect the morphological features of zebrafish larvae a standalone software named FishInspector was developed based on matlab scripts. The software provides a modular architecture that could be extended with new plugins. Using the analytical platform KNIME the coordinates of each feature provided by FishInspector was used to quantify eye size, body length, otolith-eye distance, tail and notochord malformations, swim bladder inflation, yolk sac and pericard size. To demonstrate the performance of this software, a training set of hundreds of embryos treated with chemicals of different modes of action was assessed. Morphological features were complemented by measurement of heart rate and behavioral effects, including spontaneous movements at 24 hpf and locomotor response at 96 hpf. For each endpoint concentrationresponse curves were derived in order to rank the toxicity, cluster responses and potentially derive diagnostic signatures. Some of the substances displayed a high toxic ratio, supporting a specific interaction with embryonic development. In some cases, quantitative endpoints were more sensitive than cumulative manual analysis. Furthermore, the comparative assessment allowed to distinguish developmental delay from other type of interactions. This approach contributes to understand the predictive capacity of the zebrafish embryo assay for developmental toxicity testing. Reference [1] G. Zhang, K.R. Roell, L. Truong, R.L. Tanguay, D.M. Reif, A data-driven weighting scheme for multivariate phenotypic endpoints recapitulates zebrafish developmental cascades, Toxicol. Appl. Pharmacol. 314 (2017) 109–117.
http://dx.doi.org/10.1016/j.reprotox.2017.06.179
P-39 Human embryonic stem cells as a model for paracetamol induced epigenetic deregulation and neurotoxicity Spildrejorde 1,7,∗ ,
Falck 2,7,∗ ,
Mari Martin Magnus Leithaug 1,7 , Kristina Gervin 4,7 , Hege B. Fjerdingstad 3 , Hedvig Nordeng 4,5,7 , Ragnhild E. Paulsen 6,7 , Ragnhild Eskeland 2,3,7 , Robert Lyle 1,7
49
[3] A.B. Csoka, M. Szyf, Epigenetic side-effects of common pharmaceuticals: a potential new field in medicine and pharmacology, Med. Hypotheses 73 (5) (2009) 770–780.
http://dx.doi.org/10.1016/j.reprotox.2017.06.178 P-40 Effect signatures in zebrafish embryos exposed to compounds with potential developmental toxicity
1
Department of Medical Genetics, Oslo University Hospital, Norway 2 Department of Biosciences, University of Oslo, Norway 3 Norwegian Center for Stem Cell Research, Department of Immunology, Oslo University Hospital, Norway 4 Pharmacoepidemiology and Drug Safety Research Group, Department of Pharmacy, School of Pharmacy, University of Oslo, Norway 5 Department of Child Health, Norwegian Institute of Public Health, Oslo, Norway 6 Department of Pharmaceutical Biosciences, University of Oslo, Norway 7 PharmaTox Strategic Research Initiative, Faculty of Mathematics and Natural Sciences, University of Oslo, Norway Recent cohort studies have suggested an association between paracetamol usage during pregnancy and risk of neurodevelopmental disorders (e.g. attention deficit hyperactivity disorder, ADHD) in the child [1,2]. The mechanism by which paracetamol modulate the increased risk of developing ADHD is currently unknown, and we hypothesize that paracetamol is involved in changing the epigenetic landscape during the child’s neurodevelopment. Perinatal development is a critical time for the establishment of epigenetic patterns and some commonly used medications during pregnancy, such as analgesics, antidepressants and anticonvulsants, have previously been shown to alter epigenetic homeostasis [3]. A recent unpublished study from PharmaTox indicates an association between epigenetic changes in cord blood, long-term paracetamol exposure during pregnancy and ADHD diagnosis in the offspring. To study the mechanisms by which neonatal exposure to medications might affect foetal brain development, we use human embryonic stem cells (hESCs) as a model for screening of paracetamol induced neurotoxicity and the impact on the epigenetic landscape. We measured cell viability and oxidative stress level of hESC and derived neural progenitor cells upon 24hour exposure to physiological concentrations of paracetamol or its metabolite AM404 and controls. Furthermore, global transcription and DNA methylation profiles of untreated and paracetamol treated hESCs were characterised by RNA sequencing and wholegenome bisulphite analysis, respectively. Our preliminary results indicate that hESCs is a suitable model to evaluate paracetamol toxicity in early human development. Furthermore, this hESC model can potentially be used to screen other pharmaceuticals commonly used during pregnancy.
References [1] R.E. Brandlistuen, et al., Prenatal paracetamol exposure and child neurodevelopment: a sibling-controlled cohort study, Int. J. Epidemiol. 42 (6) (2013) 1702–1713. [2] J.M.D. Thompson, et al., Associations between acetaminophen use during pregnancy and ADHD symptoms measured at ages 7 and 11 years, PLoS ONE 9 (9) (2014) pe108210.
Elisabet Teixido 1,∗ , Tobias Kießling 1,2 , David Leuthold 1 , Stefan Scholz 1 1 UFZ – Helmholtz Centre for Environmental Research, Department Bioanalytical Ecotoxicology, Leipzig, Germany 2 Scientific Software Solutions, Leipzig, Germany
The zebrafish embryo model is especially suited for the screening of of chemicals for developmental toxicity. Because the embryo develops in an integrated manner, organizing and differentiating spatially, temporally and in a hierarchical manner, disturbances are anticipated to result in nonrandom patterns of anomalies. Moreover, it has been shown that developmental cascade effects triggered by chemical exposure can be recapitulated by analyzing the relationships among endpoints [1]. The present study was designed to study the relationships among different quantitative endpoints and chemical treatments to determine chemical induced phenotypic signatures in the zebrafish embryo. Images of zebrafish larvae were collected using the Vertebrate Automated Screening Technology (VAST Bioimager) to handle and proper orient embryos. To detect the morphological features of zebrafish larvae a standalone software named FishInspector was developed based on matlab scripts. The software provides a modular architecture that could be extended with new plugins. Using the analytical platform KNIME the coordinates of each feature provided by FishInspector was used to quantify eye size, body length, otolith-eye distance, tail and notochord malformations, swim bladder inflation, yolk sac and pericard size. To demonstrate the performance of this software, a training set of hundreds of embryos treated with chemicals of different modes of action was assessed. Morphological features were complemented by measurement of heart rate and behavioral effects, including spontaneous movements at 24 hpf and locomotor response at 96 hpf. For each endpoint concentrationresponse curves were derived in order to rank the toxicity, cluster responses and potentially derive diagnostic signatures. Some of the substances displayed a high toxic ratio, supporting a specific interaction with embryonic development. In some cases, quantitative endpoints were more sensitive than cumulative manual analysis. Furthermore, the comparative assessment allowed to distinguish developmental delay from other type of interactions. This approach contributes to understand the predictive capacity of the zebrafish embryo assay for developmental toxicity testing. Reference [1] G. Zhang, K.R. Roell, L. Truong, R.L. Tanguay, D.M. Reif, A data-driven weighting scheme for multivariate phenotypic endpoints recapitulates zebrafish developmental cascades, Toxicol. Appl. Pharmacol. 314 (2017) 109–117.
http://dx.doi.org/10.1016/j.reprotox.2017.06.179