The embryonic development of vertebrates is highly susceptible to disruption by exposure to chemicals, such as pharmaceuticals, pesticides, and nanoparticles. Xenobiotics can also cause developmental cardiotoxicity through generation of reactive oxygen species (ROS) and oxidative stress (1), defined as """"""""a disruption of redox signaling and control"""""""" (2). The zebrafish is a powerful model in developmental toxicology and has great potential for use in screening and understanding possible human teratogens, including those that act via oxidative stress. Much remains to be learned about the oxidative stress response during development. We have found that while embryos grow increasingly resistant to pro-oxidant exposure from 18-52 hours post fertilization (hpf), this is followed by a surprising window of heightened sensitivity between 72-96 hpf. Taking a collaborative and multidisciplinary approach, the objective of this proposal is to understand the mechanisms involved in the differential sensitivity to pro-oxidants during embryogenesis.
Aim 1 will identify critical windows of sensitivity, the role of the antioxidant response element transcription factor NRF2, and the impact of pro-oxidant exposure on cardiac progenitor cell fate decisions. These experiments will involve ROS detection by chemiluminescence, cardiac function and cell fate analysis using calcium imaging, as well as embryo survival studies and assessment of the role of NRF2 through loss and gain of function experiments.
Aim 2 will examine the role of glutathione, the most abundant component of antioxidant defenses, using quantitative real-time PCR, measurements of total glutathione, GSH:GSSG ratios, and modulation of GSH content.
Aim 3 will examine the contribution of a physiologic increase in respiration and associated ROS towards oxidant sensitivity during development by measuring oxygen consumption using self-referencing oxygen sensing microelectrodes and reducing endogenous sources of ROS by over-expressing antioxidant enzymes on the outer mitochondrial membrane. Given the growing movement of government agencies and private industry to employ zebrafish for teratogenicity and developmental toxicity, it is especially important to comprehend the critical windows of sensitivity to oxidative stress and the underlying mechanisms. This research will provide a detailed and mechanistic understanding of how oxidative stress can impact embryonic development, and identify important considerations necessary when examining chemicals (and chemical mixtures that cause oxidative stress. It will also result in a better understanding of how oxidative stress influences the development and progression of human embryotoxicity and resulting teratogenesis and diseases.

Agency
National Institute of Health (NIH)
Institute
National Institute of Environmental Health Sciences (NIEHS)
Type
Postdoctoral Individual National Research Service Award (F32)
Project #
5F32ES017585-03
Application #
8097330
Study Section
Special Emphasis Panel (ZRG1-F05-K (20))
Program Officer
Humble, Michael C
Project Start
2009-07-01
Project End
2012-06-30
Budget Start
2011-07-01
Budget End
2012-06-30
Support Year
3
Fiscal Year
2011
Total Cost
$53,042
Indirect Cost
Name
Woods Hole Oceanographic Institution
Department
Type
DUNS #
001766682
City
Woods Hole
State
MA
Country
United States
Zip Code
02543
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