Novel Zebrafish Models to Study Toxicity of PCBs and Pesticides During Pregnancy

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DESCRIPTION (provided by applicant): Hypoplastic left heart (HLH) is a grave congenital condition, uniformly fatal if untreated and requiring multiple surgeries for survival. Recently a cluster of HLH was discovered in children in the Baltimore-Washington area that correlated with accidental release of dioxins and polychlorinated biphenyls (PCBs). In an unrelated incident in Eastern NC during 2004, three migrant workers gave birth to infants with severe congenital defects, including one baby born without limbs, another with severe facial defects and associated heart abnormalities, and a third lacking reproductive structures who died shortly after birth; all birth defects were associated with pesticide overexposure. Strikingly, the PCB- induced HLH and pesticide associated birth defects may be related because PCBs and some pesticides disrupt cells by the same mechanism: activation of the aryl hydrocarbon receptor (AHR) signaling pathway which leads to tissue specific cell cycle arrest. Recently we created a novel model for toxin induced HLH in zebrafish where the AHR agonist PCB-126 induces a stringy heart with a reduced ventricle and out flow tract defects identical to those in HLH; remarkably, PCB-126 up regulates p53 and down regulates a heart specific transcription factor tbx5, implicated in Holt-Oram Syndrome and in HLH. We also found that the common organophosphate pesticide chlorpyrifos, both a developmental neurotoxin and AHR agonist, induces an enlarged atrium and a diminished ventricle reminiscent of that seen in PCB-treated zebrafish. Indeed, the cardiotoxicity common to PCB-126 and chlorpyrifos may reflect the fact that both are AHR agonists. We propose to create fluorescent reporter strains in zebrafish that will signal in living embryos when an AHR agonist disrupts cardiovascular development, and will use this approach to evaluate pesticides for developmental cardiotoxicity, particularly for teratogenic synergy. In Aim 1, we test the hypothesis that PCB-126 or chlorpyrifos activated AHR leads to down regulation of tbx5 followed by p53 mediated cell cycle arrest in the heart by using genetic epistasis and DNA microarray analysis. In Aim 2, fluorescent reporters in living zebrafish hearts will be constructed and then validated for use in signaling cardiotoxicity from exposure to PCB-126 and chlorpyrifos. The reporter strains then will be used to screen pesticides used in agriculture, in combinations and sequence typical of a growing season. We expect that our studies will provide a new testing paradigm to more accurately predict risk to EPA approved toxins as they are encountered routinely in real life. We seek to protect those in our society most vulnerable to toxin exposure those living in poverty, those with inadequate health care and migrant farm workers suffering occupational over exposure to pesticides.Hypoplastic left heart (HLH) is a grave congenital condition, uniformly fatal if untreated and requiring multiple surgeries for survival. Recently a cluster of HLH was discovered in children in the Baltimore-Washington area that correlated with accidental release of banned dioxins and polychlorinated biphenyls (PCBs). We have created a model for HLH in zebrafish where the AHR agonist, PCB-126, induces a stringy heart with a reduced ventricle and out flow tract defects identical to those seen in children with HLH. We propose to create fluorescent reporter strains in zebrafish that will signal in living embryos when an AHR agonist disrupts cardiovascular development and will use this approach to evaluate pesticides for developmental cardiotoxicity; we seek to protect those in our society most vulnerable to toxin exposure those living in poverty, those with inadequate health care and migrant farm workers suffering occupational over exposure to pesticides.