Developmental Malformations in Zebrafish Caused by Exposure to Environmental Pollutants



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It is estimated that there are about 80,000 man-made chemicals released to the environment with little to no toxicity information. Exposure to these chemicals is identified as one of the risk factors for causing birth defects. We used zebrafish as a model to identify and predict environmental chemicals causing congenital malformations. High-throughput screening of 292 unique chemicals from the ToxCast Phase I library identified 38 skeletal disruptor compounds (SDCs) producing bone and craniofacial cartilage malformations in zebrafish. We used ToxCast in vitro data and RT-qPCR to predict that these compounds affect vitamin D metabolism and dopamine transporters resulting skeletal malformations. Transcriptomics data identified that exposure to cyproconazole, a prominent SDC, induce adipogenesis, while repressing osteo and chondrogenesis. Next, we investigated vascular disruptor compounds (VDCs) that cause malformations in vivo in zebrafish vasculature and in vitro in human umbilical cord endothelial cells (HUVECs). ToxCast data correlations identified that VDCs alter signaling of nuclear receptors like ER, AR and transcription factors like Hif1α, causing vascular perturbations. The ToxCast in vitro assays that correlated with either the identified SDCs or VDCs were used to predict chemicals with in vivo toxicity in zebrafish using the ToxPi tool. Exposures to predicted SDCs and VDCs were found to induce bone and vascular malformations, respectively. We confirmed that chemicals predicted to not have any negative effect on the skeleton or vasculature were indeed inert in zebrafish. The predicted VDCs affected in vitro HUVEC tube formation, while predicted non-VDCs did not. Lastly, we investigated the effects of a widely used insecticide, pyriproxyfen, which is suspected to cause microcephaly. We identified that pyriproxyfen exposure causes significantly smaller heads, smaller midbrains and a prominent gap along the mid-dorsal margin in the brains in zebrafish compared to controls. Transcriptomic data proposed that pyriproxyfen affects neuronal and axonal migration and cerebrovascular development. In conclusion, we used zebrafish embryo toxicity in combination with pathway-based in vitro data to identify environmental chemicals that cause developmental malformations and their mechanisms of action. Most of the identified molecular mechanisms are conserved among vertebrates, suggesting that these compounds could potentially be harmful to humans and other mammals.



Zebrafish, Transgenics, Environmental chemicals, High-throughput, Skeletal malformation, Skeletal disruptors, Computational toxicology, Angiogenesis, Vascular disruptors, Pyriproxyfen, Brain malformation, Neuronal migration