Modulating glutathione thiol status alters pancreatic β-cell morphogenesis in the developing zebrafish (Danio rerio) embryo
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 (Nf...
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| Format: | Article |
| Language: | English |
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Elsevier
2021-01-01
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| Series: | Redox Biology |
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| Online Access: | http://www.sciencedirect.com/science/article/pii/S2213231720309939 |
| _version_ | 1819077396926562304 |
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| author | Archit Rastogi Emily G. Severance Haydee M. Jacobs Sarah M. Conlin Sadia T. Islam Alicia R. Timme-Laragy |
| author_facet | Archit Rastogi Emily G. Severance Haydee M. Jacobs Sarah M. Conlin Sadia T. Islam Alicia R. Timme-Laragy |
| author_sort | Archit Rastogi |
| collection | DOAJ |
| description | 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 co-ortholog) in islet morphogenesis. We delineated critical windows of susceptibility to redox disruption of β-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 μM; 10-min at 48 hpf) or tert-butylhydroquinone (1 μM; 48-56 hpf) decreased β-cell cluster area at 96 hpf. Conversely, exposures to antioxidant N-acetylcysteine (bolsters GSH pools; 100 μM; 48-72 hpf) or sulforaphane (activates Nrf2a; 20 μM; 48-72 hpf) significantly increased islet areas. Nrf2a was also stabilized in β-cells: 10-min exposures to 77.6 μM tert-butylhydroperoxide significantly increased Nrf2a protein compared to control islet cells that largely lack stabilized Nrf2a; 10-min exposures to higher (776 μM) 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 μM) 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 β-cell development impact morphogenesis, with implications for disease states at later life stages. Our work identifies a potential molecular target (Nrf2) that mediates abnormal β-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 β-cells, rendering them vulnerable to later-life stresses and disease. |
| first_indexed | 2024-12-21T18:56:32Z |
| format | Article |
| id | doaj.art-d79f35c1f213476c943ef65c9022d0b9 |
| institution | Directory Open Access Journal |
| issn | 2213-2317 |
| language | English |
| last_indexed | 2024-12-21T18:56:32Z |
| publishDate | 2021-01-01 |
| publisher | Elsevier |
| record_format | Article |
| series | Redox Biology |
| spelling | doaj.art-d79f35c1f213476c943ef65c9022d0b92022-12-21T18:53:36ZengElsevierRedox Biology2213-23172021-01-0138101788Modulating glutathione thiol status alters pancreatic β-cell morphogenesis in the developing zebrafish (Danio rerio) embryoArchit Rastogi0Emily G. Severance1Haydee M. Jacobs2Sarah M. Conlin3Sadia T. Islam4Alicia R. Timme-Laragy5Molecular & Cellular Biology Graduate Program, University of Massachusetts, Amherst, MA, 01003, USADepartment of Environmental Health Sciences, School of Public Health and Health Sciences, University of Massachusetts, Amherst, MA, 01003, USADepartment of Environmental Health Sciences, School of Public Health and Health Sciences, University of Massachusetts, Amherst, MA, 01003, USADepartment of Environmental Health Sciences, School of Public Health and Health Sciences, University of Massachusetts, Amherst, MA, 01003, USADepartment of Environmental Health Sciences, School of Public Health and Health Sciences, University of Massachusetts, Amherst, MA, 01003, USAMolecular & Cellular Biology Graduate Program, University of Massachusetts, Amherst, MA, 01003, USA; Department of Environmental Health Sciences, School of Public Health and Health Sciences, University of Massachusetts, Amherst, MA, 01003, USA; Corresponding author. University of Massachusetts Amherst, 686 N. Pleasant St., Goessmann 171B, Amherst, MA, 01003, USA.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 co-ortholog) in islet morphogenesis. We delineated critical windows of susceptibility to redox disruption of β-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 μM; 10-min at 48 hpf) or tert-butylhydroquinone (1 μM; 48-56 hpf) decreased β-cell cluster area at 96 hpf. Conversely, exposures to antioxidant N-acetylcysteine (bolsters GSH pools; 100 μM; 48-72 hpf) or sulforaphane (activates Nrf2a; 20 μM; 48-72 hpf) significantly increased islet areas. Nrf2a was also stabilized in β-cells: 10-min exposures to 77.6 μM tert-butylhydroperoxide significantly increased Nrf2a protein compared to control islet cells that largely lack stabilized Nrf2a; 10-min exposures to higher (776 μM) 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 μM) 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 β-cell development impact morphogenesis, with implications for disease states at later life stages. Our work identifies a potential molecular target (Nrf2) that mediates abnormal β-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 β-cells, rendering them vulnerable to later-life stresses and disease.http://www.sciencedirect.com/science/article/pii/S2213231720309939PancreasNfe2l2Antioxidant defensesIsletRedoxEmbryonic development |
| spellingShingle | Archit Rastogi Emily G. Severance Haydee M. Jacobs Sarah M. Conlin Sadia T. Islam Alicia R. Timme-Laragy Modulating glutathione thiol status alters pancreatic β-cell morphogenesis in the developing zebrafish (Danio rerio) embryo Redox Biology Pancreas Nfe2l2 Antioxidant defenses Islet Redox Embryonic development |
| title | Modulating glutathione thiol status alters pancreatic β-cell morphogenesis in the developing zebrafish (Danio rerio) embryo |
| title_full | Modulating glutathione thiol status alters pancreatic β-cell morphogenesis in the developing zebrafish (Danio rerio) embryo |
| title_fullStr | Modulating glutathione thiol status alters pancreatic β-cell morphogenesis in the developing zebrafish (Danio rerio) embryo |
| title_full_unstemmed | Modulating glutathione thiol status alters pancreatic β-cell morphogenesis in the developing zebrafish (Danio rerio) embryo |
| title_short | Modulating glutathione thiol status alters pancreatic β-cell morphogenesis in the developing zebrafish (Danio rerio) embryo |
| title_sort | modulating glutathione thiol status alters pancreatic β cell morphogenesis in the developing zebrafish danio rerio embryo |
| topic | Pancreas Nfe2l2 Antioxidant defenses Islet Redox Embryonic development |
| url | http://www.sciencedirect.com/science/article/pii/S2213231720309939 |
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