Grantee Research Project Results
2009 Progress Report: Integrating Innovative Biomarkers of Environmentally Induced Disease for Children in Agricultural Communities
EPA Grant Number: R832733Title: Integrating Innovative Biomarkers of Environmentally Induced Disease for Children in Agricultural Communities
Investigators: Faustman, Elaine , Griffith, William C. , Yu, Xiaozhong
Institution: University of Washington
EPA Project Officer: Callan, Richard
Project Period: October 1, 2005 through September 30, 2008 (Extended to September 30, 2010)
Project Period Covered by this Report: October 1, 2008 through September 30,2009
Project Amount: $749,997
RFA: Early Indicators of Environmentally Induced Disease (2004) RFA Text | Recipients Lists
Research Category: Biology/Life Sciences , Children's Health
Objective:
The purpose of this study is to develop an integrative tool for evaluating the importance of knowledge of genomic biomarkers of susceptibility and early response for identifying and characterizing the exposure-effect-disease relationship in adults and children in agricultural and non-agricultural communities.Progress Summary:
This study is coordinated with an ongoing community-based participatory research study in the Yakima Valley (CBPR), a part of the EPA/NIEHS funded Children’s Health Risk Research Center (CHC). This study uses biorepository samples collected as part of the CHC-CBPR study. We are evaluating potential new biomarkers of susceptibility (genotype for pesticide metabolism and oxidative protective pathways), biomarkers of early biological effect (gene expression responses in oxidative response and cell death), and biomarkers of effect (cholinesterase and disease expression pathways) in order to improve our early identification and prevention of pesticide-related health effects.
Paraoxonase (PON1) genotype and phenotype status is a biomarker of susceptibility for OP toxicity. We have completed a substrate-based geno/phenotype laboratory analysis of the approximately 200 farmworkers and non-farmworkers in our study population.
Buccal cells (collected from the inner cheek) offer an attractive non-invasive alternative for biospecimen collection. We have refined the collection techniques using soft oral brushes to produce a more applicable protocol for collecting specimens from children. In order to optimize the yield and quality of DNA isolated from buccal samples, we compared the performance of three extraction strategies. A “salting out” protocol with an incubation at -80 °C proved to be the best method for our needs. DNA was isolated from the children’s buccal samples amplified to provide sufficient material for multiplex genotyping on the Affymetrix Drug Metabolizing Enzyme and Transporter (DMET) platform. After cleanup, the amplification products were of sufficient yield and quality to proceed with genotyping. To genotype adult study participants, DNA was extracted from buffy coat samples using the same “salting out” protocol.
In addition to their wide use for DNA analysis, the evaluation of buccal cell specimens recently has been shown to have high potential for RNA expression analysis, chromosomal analysis, genotoxicity analysis, and proteomic analysis. We have established an mRNA extraction protocol that produces high quality mRNA in sufficient quantity to be used for microarray-based gene expression analysis.
Work has progressed on developing components of the integrative framework tool to describe exposure, dose, early biological effects, and altered structure/function as data have become available from the CHC-CBPR study. We also have used data from other toxicological studies for development of the model components. Using microarray data from a laboratory-based toxicological study of multiple doses measured at multiple time points, we have developed a systems-based gene ontology (GO) methodology (Yu, et al., 2006). We used information from GO databases and showed how information across multiple genes in biological pathways can be combined to provide a better understanding of toxicological modes of action. This approach has been integrated into the GenMAPP program (http://www.genmapp.org/tutorials/GO-Quant.htm).
We have used Bayesian-based mixed effects methodologies to model exposure to organophosphate pesticides (OPs) in farmworkers and their children during the thinning season. Exposure as measured by OP dialkyl phosphate urinary metabolites is a function of the OP parent compound concentrations in vehicle and house dust, the types of orchards worked in by the adult (pome or non-pome), and other factors. The mixed effects model shows that the day-to-day variability within an individual is larger than between individuals.
Components of the dose model to describe metabolism of OPs by cytochrome P450s have been developed to account for the effect of genotype/phenotypes in the CYP450s. Methods to identify early biological effects based on gene expression microarray data are being developed to determine whether multiple genes in biological pathways rather than single genes are altered by exposure. These methods provide an important component of the integrative framework tool to test whether “omic” biomarkers of disease are correlated with OP exposure and hence assess their potential for identifying and decreasing pesticide exposure.
Future Activities:
Genotyping of study samples is ongoing and will be completed during the coming year. Upon completion, this information will be used to evaluate genotypes for enzymes known to be associated with pesticide metabolism and assess their relationship with pesticide urinary metabolite profiles, cholinesterase inhibition, and other endpoints. The gene expression profiles will be modeled as an endpoint to evaluate gene-environment interactions between pesticide exposures and genotypes for pesticide-metabolizing enzymes. We will use the various results and data components that we have assembled to inform our integrative framework tool as a means to link exposure and dose. The early response portion of the tool is designed to allow for incorporation of pathway-specific molecular response data into the exposure-effect-disease paradigm. We will use our GO-Quant methodology to provide quantitative analysis of the response pathways
Journal Articles:
No journal articles submitted with this report: View all 20 publications for this projectSupplemental Keywords:
organophosphate pesticides, genomic biomarkers, children, neurodegenerative disease
, RFA, Health, Scientific Discipline, ENVIRONMENTAL MANAGEMENT, Health Risk Assessment, Biochemistry, Children's Health, Risk Assessment, pesticide exposure, Human Health Risk Assessment, assessment of exposure, children's vulnerablity, susceptibility, children's environmental health, biological markers, agricultural community, disease
Progress and Final Reports:
Original AbstractThe perspectives, information and conclusions conveyed in research project abstracts, progress reports, final reports, journal abstracts and journal publications convey the viewpoints of the principal investigator and may not represent the views and policies of ORD and EPA. Conclusions drawn by the principal investigators have not been reviewed by the Agency.