Timme-Laragy, Alicia

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Associate Professor, Department of Environmental Health Sciences, School of Public Health & Health Sciences
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Timme-Laragy
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Alicia
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Environmental Health and Protection
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2016 - 201912020 - 20211
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    Embryonic Exposures to Perfluorooctanesulfonic Acid (PFOS) Disrupt Pancreatic Organogenesis in the Zebrafish, Danio rerio
    (2016-01-01) Sant, Karilyn E.; Jacobs, Haydee M.; Borofski, Katrina A.; Moss, Jennifer B.; Timme-Laragy, Alicia R.
    Perfluorooctanesulfonic acid (PFOS) is a ubiquitous environmental contaminant, previously 16 utilized as a non-stick application for consumer products and firefighting foam. It can cross the 17 placenta, and has been repeatedly associated with increased risk for diabetes in epidemiological 18 studies. Here, we sought to establish the hazard posed by embryonic PFOS exposures on the 19 developing pancreas in a model vertebrate embryo, and develop criteria for an adverse outcome 20 pathway (AOP) framework to study the developmental origins of metabolic dysfunction. 21 Zebrafish (Danio rerio) embryos were exposed to 16, 32, or 64 μM PFOS beginning at the mid-22 blastula transition. We assessed embryo health, size, and islet morphology in Tg(insulin-GFP) 23 embryos at 48, 96 and 168 hpf, and pancreas length in Tg(ptf1a-GFP) embryos at 96 and 168 24 hpf. QPCR was used to measure gene expression of endocrine and exocrine hormones, digestive 25 peptides, and transcription factors to determine whether these could be used as a predictive 26 measure in an AOP. Embryos exposed to PFOS showed anomalous islet morphology and 27 decreased islet size and pancreas length in a U-shaped dose-response curve, which resemble 28 congenital defects associated with increased risk for diabetes in humans. Expression of genes 29 encoding islet hormones and exocrine digestive peptides followed a similar pattern, as did total 30 larval growth. Our results demonstrate that embryonic PFOS exposures can disrupt pancreatic 31 organogenesis in ways that mimic human congenital defects known to predispose individuals to 32 diabetes; however, future study of the association between these defects and metabolic 33 dysfunction are needed to establish an improved AOP framework.
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    Modulating Glutathione Thiol Status Alters Pancreatic β-cell Morphogenesis in the Developing Zebrafish (Danio rerio) Embryo
    (2021-01-01) Rastogi, Archit; Severance, Emily G.; Jacobs, Haydee M.; Conlin, Sarah M.; Islam, Sadia T.; Timme-Laragy, Alicia R.
    Emerging evidence suggests that redox-active chemicals perturb pancreatic islet development. To better understand potential mechanisms for this, we used zebrafish (Danio rerio) embryos to investigate roles of glutathione (GSH; predominant cellular redox buffer) and the transcription factor Nrf2a (Nfe2l2a; zebrafish Nrf2 coortholog) in islet morphogenesis. We delineated critical windows of susceptibility to redox disruption of beta-cell morphogenesis, interrogating embryos at 24, 48 and 72 h post fertilization (hpf) and visualized Nrf2a expression in the pancreas using whole-mount immunohistochemistry at 96 hpf. Chemical GSH modulation at 48 hpf induced significant islet morphology changes at 96 hpf. Pro-oxidant exposures to tert-butylhydroperoxide (77.6 mu M; 10-min at 48 hpf) or tert-butylhydroquinone (1 mu M; 48-56 hpf) decreased beta-cell cluster area at 96 hpf. Conversely, exposures to antioxidant N-acetylcysteine (bolsters GSH pools; 100 mu M; 48-72 hpf) or sulforaphane (activates Nrf2a; 20 mu M; 48-72 hpf) significantly increased islet areas. Nrf2a was also stabilized in beta-cells: 10-min exposures to 77.6 mu M tert-butylhydroperoxide significantly increased Nrf2a protein compared to control islet cells that largely lack stabilized Nrf2a; 10-min exposures to higher (776 mu M) tert-butylhydroperoxide concentration stabilized Nrf2a throughout the pancreas. Using biotinylated-GSH to visualize in situ protein glutathionylation, islet cells displayed high protein glutathionylation, indicating oxidized GSH pools. The 10-min high (776 mu M) tert-butylhydroperoxide exposure (induced Nrf2a globally) decreased global protein glutathionylation at 96 hpf. Mutant fish expressing inactive Nrf2a were protected against tert-butylhydroperoxide-induced abnormal islet morphology. Our data indicate that disrupted redox homeostasis and Nrf2a stabilization during pancreatic beta-cell development impact morphogenesis, with implications for disease states at later life stages. Our work identifies a potential molecular target (Nrf2) that mediates abnormal beta-cell morphology in response to redox disruptions. Moreover, our findings imply that developmental exposure to exogenous stressors at distinct windows of susceptibility could diminish the reserve redox capacity of beta-cells, rendering them vulnerable to later-life stresses and disease.