Grantee Research Project Results

2013 Progress Report: Molecular Mechanisms

EPA Grant Number: R834514C003
Subproject: this is subproject number 003 , established and managed by the Center Director under grant R834514
(EPA does not fund or establish subprojects; EPA awards and manages the overall grant for this center).
Center: Predictive Toxicology Center for Organotypic Cultures and Assessment of AOPs for Engineered Nanomaterials
Center Director: Faustman, Elaine
Title: Molecular Mechanisms
Investigators: Faustman, Elaine
Current Investigators: Faustman, Elaine , Fenske, Richard , Griffith, William C. , Yost, Michael , Costa, Lucio G , Furlong, Clement , Thompson, Engelberta , Vigoren, Eric M. , Carr, Catherine J
Institution: University of Washington
EPA Project Officer: Callan, Richard
Project Period: September 25, 2008 through September 24, 2016
Project Period Covered by this Report: September 25, 2012 through September 24,2013
RFA: Children's Environmental Health and Disease Prevention Research Centers (with NIEHS) (2009) RFA Text |  Recipients Lists
Research Category: Children's Health , Human Health

Objective:

Since 1998, researchers of the University of Washington Center for Child Environmental Health Risks Research (the Center) have been using a multi-disciplinary research approach working in the lab, in the field, and in the community to understand the mechanisms that define children’s susceptibility to pesticides, identify the implications of this susceptibility for development and learning, and partner with our communities to translate our findings into risk communication, risk management, and prevention strategies. The Center is jointly funded by the National Institute of Environmental Health Sciences (NIEHS) and the Environmental Protection Agency (EPA).

The Center is administratively housed within the Institute for Risk Analysis and Risk Communication, also directed by Center Director Dr. Elaine M. Faustman, which is in the University of Washington’s School of Public Health. The Center includes partnerships with the Fred Hutchinson Cancer Research Center and the Yakima Valley community, located in the agricultural center of Washington State, to jointly sponsor a Community Based Participatory Research (CBPR) Project aimed at reducing childhood pesticide exposure.

All Center efforts are highly integrated with two field-based research projects, two laboratory-based research projects, three facility cores, an Administrative Core, and Faculty Development Investigator.

The specific objectives of the laboratory-based research projects——a molecular mechanisms research project and a genetic susceptibility research project—are:

1. To identify cellular, biochemical and molecular mechanisms that cause adverse developmental neurotoxicity of pesticides; and
2. To identify susceptibility factors for developmental neurotoxicity of pesticides.

One of the objectives of the Molecular Mechanisms research project is to understand how exposure to chemical and nonchemical stressors can impact children’s' neurological developmental processes. In order to prevent and predict developmental neurotoxicity, it is important to understand the “who, what, where and when” of chemical and non-chemical stressor biology and exposure. A key focus of this research project is to understand who is exposed, to which environmental chemicals, where the exposure is coming from and to identify biological impacts occurring across life-course. For example, the Center investigates how and exposure early in development may differ significantly from impacts seen later when exposures occur at a different stage of development of life-course (toddler versus early school age children). Once we have characterized the exposures, we can use this information to make predictions about chemicals that could adversely impact neurodevelopment and then take action to prevent those exposures, especially in susceptible populations, such as children or pregnant women.

Our Center is particularly interested in pesticide exposure across the life-course. Pesticides can have adverse impacts on brain and nervous system development by altering the regulatory dynamics of normal central nervous system cell proliferation, differentiation, and cell death; which can result in altered morphogenesis. These alterations are correlated with subsequent deficits in learning and development. A thorough understanding of mechanisms of toxicity at the molecular, cellular, and organism level is therefore important as these mechanisms may define unique life stages that may be “windows of susceptibility” for many neurodevelopmental toxicants as well as for pesticides. The Molecular Mechanism Research Project evaluates these mechanistic processes across several pesticide classes in order to understand the commonality and uniqueness of pesticide toxicity across dose, life-course, cell type, and behavioral domain.

This research project brings together the expertise from different laboratories to focus on the relationship between pesticide‐induced alterations in cell proliferation and death during embryonic/postnatal development and postnatal alterations in behavior. These projects use rodent models and follow both in vivo and in vitro impacts. The laboratory of Dr. Elaine Faustman addresses the impacts of pesticides and model neurotoxicants on prenatal processes generating neurons, known as neurogenesis, and the laboratory of Dr. Lucio Costa evaluates the postnatal effects of pesticides on processes of generating non-neuronal glial cells, known as gliogenesis.

Progress Summary:

In the last several years, the Faustman lab initiated studies to explore new in vitro models for the evaluation of neurogenesis including assessment of proliferation and differentiation processes for in vivo comparison. The highlights of this year include an in vitro human neural progenitor cell model which was established to study adverse effects of the OP pesticide chlorpyrifos (CP) during neuronal proliferation and differentiation. Using confocal imaging to characterize cell by cell expression of markers of differentiation and proliferations will help illuminate developmental processes occurring in these cell cultures through both normal development and when perturbed by pesticide exposure. Understanding these dynamics will help improve and stabilize culture conditions to more accurately reflect in vivo developmental processes. Towards this goal the researchers have characterized protein expression and this expression has been compared to the Allen Developing Mouse Brain Atlas reports on gene expression. The researchers have included not only in vitro human neuronal progenitor cells with in vivo murine brain development but also have added a comparison with their in vitro murine Central Nervous Systems micromass cell cultures.

Another highlight of recent work are the results of our studies suggesting that epigenetic factors play a role in the neurodevelopmental effects associated with CP exposure. For example, toxicogenomic profiling of CP exposed fetal and maternal mouse brains showed significant alteration to chromatin modifications. Exposure to CP during critical “windows of susceptibility” has been reported to alter behavior and development of the central nervous system by interfering with the regulatory pathways that control the production and selective cell loss of neurons. These results showing differential alterations of epigenetic markers during proliferation and differentiation can offer cell stage specific mechanistic information to help researchers tease out neurodevelopmental effects of the organophosphate pesticide CP.

The focus of Dr. Costa’s' project is the investigation of glial-neuronal interactions in neuronal differentiation. Work in Dr. Costa’s' laboratory has focused on the OP pesticide diazinon (DZ) and its active metabolite diazoxon (DZO). As both have been shown to induce oxidative stress in neurons, and based on previous studies with manganese, it was hypothesized that these OPs also may induce oxidative stress in astrocytes, and that this would impair the ability of these cells to induce normal neuronal differentiation, which is necessary for healthy neurological development. In our recent work both DZ and DZO induced oxidative stress as assessed by measuring levels of reactive oxygen species (ROS), in rat cortical astrocytes. In separate experiments the ability of OPs to directly affect neuritogenesis also was investigated. In this experimental protocol, neurons were cultured alone, in the presence of astrocyte-conditioned serum, but not in co-culture with astrocytes. Preliminary experiments indicate that when co-incubated with neurons, astrocytes are capable of increasing the levels of the protective antioxidant, glutathione (GSH). The increased GSH levels in neurons would thus explain their increased resistance to OPs. This was further confirmed by raising GSH levels in neurons with the use of GSH ethyl ester.

The complimentary objectives of the Faustman and Costa laboratories have contributed to the advancement of our understanding of the potential for, and magnitude of, impacts of pesticides on neurogenesis and gliogenesis. In both these essential neurodevelopmental pathways the balance between initial proliferation and subsequent specific differentiation is integral for proper neurodevelopment. The integration of the Faustman and Costa laboratories allows us to better understand the molecular mechanisms which can cause adverse neurological development in children, following pesticide exposure. Our investigation of critical molecular pathways facilitating proliferation and toxicant response allow us to better predict neurological toxicity and increase our understanding about the importance of timing of exposure. The results from this project can be used to identify particularly vulnerable or sensitive populations and can be translated into information relevant for establishing conditions promoting environmental and public health safety.

Future Activities:

The highlights above illustrate the high volume and quality of the Center's efforts aiming at reducing the adverse effects of environmental pesticide exposures in children. The Center continues to work to understand the mechanisms that define children's susceptibility to pesticides, identify the implications of this susceptibility for health impacts on development and learning, and partner with our communities to translate our findings into risk communication, risk management, and prevention strategies. Center researchers continue to work in the lab, in the field, and in the community to bring a unique and successful approach to the study of children's environmental health.

Journal Articles:

No journal articles submitted with this report: View all 55 publications for this subproject

Supplemental Keywords:

Health, RFA, Scientific Discipline, INTERNATIONAL COOPERATION, ENVIRONMENTAL MANAGEMENT, Environmental Policy, Biology, Children's Health, Biochemistry, Environmental Monitoring, Risk Assessment, biological markers, pesticides, age-related differences, pesticide exposure, agricultural community, children's vulnerablity, molecular research

Relevant Websites:

Center for Child Environmental Health Risks Research | Department of Environmental & Occupational Health Science | University of Washington School of Public Health Exit

Progress and Final Reports:

Original Abstract

2009

2010

2011 Progress Report

2012

2014

2015 Progress Report

Final Report

Main Center Abstract and Reports:

R834514    Predictive Toxicology Center for Organotypic Cultures and Assessment of AOPs for Engineered Nanomaterials

Subprojects under this Center: (EPA does not fund or establish subprojects; EPA awards and manages the overall grant for this center).
R834514C001 Community-Based Participatory Research
R834514C002 Pesticide Exposure Pathways
R834514C003 Molecular Mechanisms
R834514C004 Genetic Susceptibility