Zebrafish (Danio rerio) fed vitamin E-deficient diets produce embryos with increased morphologic abnormalities and mortality

The Linus Pauling Institute, Oregon State University, Corvallis, OR 97331, USA.
The Journal of nutritional biochemistry (Impact Factor: 3.79). 06/2011; 23(5):478-86. DOI: 10.1016/j.jnutbio.2011.02.002
Source: PubMed

ABSTRACT Vitamin E (α-tocopherol) is required to prevent fetal resorption in rodents. To study α-tocopherol's role in fetal development, a nonplacental model is required. Therefore, the zebrafish, an established developmental model organism, was studied by feeding the fish a defined diet with or without added α-tocopherol. Zebrafish (age, 4-6 weeks) were fed the deficient (E-), sufficient (E+) or lab diet up to 1 years. All groups showed similar growth rates. The exponential rate of α-tocopherol depletion up to ~80 day in E- zebrafish was 0.029±0.006 nmol/g, equivalent to a depletion half-life of 25±5 days. From age ~80 days, the E- fish (5±3 nmol/g) contained ~50 times less α-tocopherol than the E+ or lab diet fish (369±131 or 362±107, respectively; P<.05). E-depleted adults demonstrated decreased startle response suggesting neurologic deficits. Expression of selected oxidative stress and apoptosis genes from livers isolated from the zebrafish fed the three diets were evaluated by quantitative polymerase chain reaction and were not found to vary with vitamin E status. When E-depleted adults were spawned, they produced viable embryos with depleted α-tocopherol concentrations. The E- embryos exhibited a higher mortality (P<.05) at 24 h post-fertillization and a higher combination of malformations and mortality (P<.05) at 120 h post-fertillization than embryos from parents fed E+ or lab diets. This study documents for the first time that vitamin E is essential for normal zebrafish embryonic development.

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Available from: Robert L Tanguay, Sep 25, 2015
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    • "We have previously demonstrated that E-embryos appear normal at 36 hpf, but by 48 hpf, 50% of E-embryos display developmental defects [3]. We hypothesized that Į-tocopherol is needed to mediate the production of hydroxy-PUFAs, specifically HETEs and HDHAs. "
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    ABSTRACT: To test the hypothesis that embryogenesis depends upon α-tocopherol (E) to protect embryo polyunsaturated fatty acids (PUFAs) from lipid peroxidation, new methodologies were applied to measure α-tocopherol and fatty acids in extracts from saponified zebrafish embryos. A solid phase extraction method was developed to separate the analyte classes, using a mixed mode cartridge (reverse phase, π-π bonding, strong anion exchange), then α-tocopherol and cholesterol were measured using standard techniques, while the fatty acids were quantitated using a novel, reverse phase liquid chromatography-mass spectrometry (LC-MS) approach. We also determined if α-tocopherol status alters embryonic lipid peroxidation products by analyzing 24 different oxidized products of arachidonic or docosahexaenoic (DHA) acids in embryos using LC with hybrid quadrupole-time of flight MS. Adult zebrafish were fed E- or E+ diets for 4 months, and then were spawned to obtain E- and E+ embryos. Between 24 and 72 hours post-fertilization (hpf), arachidonic acid decreased 3-times faster in E- (21 pg/h) compared with E+ embryos (7 pg/h, P<0.0001), while both α-tocopherol and DHA concentrations decreased only in E- embryos. At 36 hpf, E- embryos contained double the 5-hydroxy-eicosatetraenoic acids and 7-hydroxy-DHA concentrations, while other hydroxy-lipids remained unchanged. Vitamin E deficiency during embryogenesis depleted DHA and arachidonic acid, and increased hydroxy-fatty acids derived from these PUFA, suggesting that α-tocopherol is necessary to protect these critical fatty acids.
    12/2013; 2(1):105-13. DOI:10.1016/j.redox.2013.12.007
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    • "Previously, we utilized the zebrafish model to separate the maternal and embryonic requirements, and to characterize the molecular defect of embryonic vitamin E deficiency. We reported that α-tocopherol-deficient fish spawn and produce viable eggs, but within days the embryos and larvae display developmental impairment and increased risk of mortality [7], establishing a critical embryonic need for α-tocopherol. Zebrafish nutrients are derived solely from the yolk sac for the initial 4–5 days post fertilization. "
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    ABSTRACT: The hepatic α-tocopherol transfer protein (TTP) is required for optimal α-tocopherol bioavailability in humans; mutations in the human TTPA gene result in the heritable disorder ataxia with vitamin E deficiency (AVED, OMIM #277460). TTP is also expressed in mammalian uterine and placental cells and in the human embryonic yolk-sac, underscoring TTP's significance during fetal development. TTP and vitamin E are essential for productive pregnancy in rodents, but their precise physiological role in embryogenesis is unknown. We hypothesize that TTP is required to regulate delivery of α-tocopherol to critical target sites in the developing embryo. We tested to find if TTP is essential for proper vertebrate development, utilizing the zebrafish as a non-placental model. We verify that TTP is expressed in the adult zebrafish and its amino acid sequence is homologous to the human ortholog. We show that embryonic transcription of TTP mRNA increases >7-fold during the first 24 hours following fertilization. In situ hybridization demonstrates that Ttpa transcripts are localized in the developing brain, eyes and tail bud at 1-day post fertilization. Inhibiting TTP expression using oligonucleotide morpholinos results in severe malformations of the head and eyes in nearly all morpholino-injected embryos (88% compared with 5.6% in those injected with control morpholinos or 1.7% in non-injected embryos). We conclude that TTP is essential for early development of the vertebrate central nervous system.
    PLoS ONE 10/2012; 7(10):e47402. DOI:10.1371/journal.pone.0047402 · 3.23 Impact Factor
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    ABSTRACT: Seven-month-old zebrafish (Danio rerio) were fed four different diets to test the hypothesis that diet affects spawning success and resulting characteristics of eggs and offspring. The diets were: the recommended feeding regime for zebrafish (a mixture of Artemia, flake feed, and liver paste); Artemia; a flake feed; and a commercially available trout diet. The number of eggs laid and average egg diameter were significantly different as functions of male, female, and individual matings. Fish fed the flake diet produced significantly fewer eggs (mean, 116) than fish fed all other diets (means, 166-187). However, the percent hatch of eggs from fish fed the flake diet (62.5%) was significantly higher than from fish fed the trout diet (19.5%). The percentages of hatched eggs from fish fed the control diet (36.2%) or Artemia (35.6%) were not significantly different from each other or from fish fed the other two diets. Wet weight and diameter of eggs were not significantly affected by diet. Larval length was significantly higher from parents fed the flake diet (14.5 mm) compared to larvae from parents fed Artemia (13.7 mm). Length of larvae from fish fed the control or trout diets was intermediate and not significantly different from fish fed the flake diet or Artemia. Larval weight was not significantly affected by dietary treatment, but offspring from fish fed the flake diet were heavier than larvae from adults fed any of the other diets. Feeding adult zebrafish the flake diet alone resulted in more viable offspring and larger larvae and is a simpler feeding regime than the current recommendation. The authors recommend feeding adult zebrafish flake diets to satiation three times daily for maximum production of viable offspring.
    Zebrafish 02/2007; 4(1):69-74. DOI:10.1089/zeb.2006.9993 · 1.95 Impact Factor
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