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2000 Progress Report: Genetic Susceptibility to Pesticides (Paraoxonase Polymorphism or PON1 Study)
EPA Grant Number: R826886C002Subproject: this is subproject number 002 , established and managed by the Center Director under grant R826886
(EPA does not fund or establish subprojects; EPA awards and manages the overall grant for this center).
Center: University of Washington
Center Director: Faustman, Elaine
Title: Genetic Susceptibility to Pesticides (Paraoxonase Polymorphism or PON1 Study)
Investigators: Faustman, Elaine
Institution: University of Washington
EPA Project Officer: Fields, Nigel
Project Period: August 1, 1998 through December 31, 2003
Project Period Covered by this Report: August 1, 1999 through July 31,2000
Project Amount: Refer to main center abstract for funding details.
RFA: Centers for Children's Environmental Health and Disease Prevention Research (1998)
Research Category: Health Effects , Children's Health
Description:
Objective:The general aim of this project remains that of determining the role of the enzyme paraoxonase (PON1) in the developmental toxicity and neurotoxicity of organophosphorus (OP) insecticides. The specific aims also remain as follows: (1) investigate the acute toxicity of a number of OPs in wild type and PON1- knockout mice; (2) investigate whether human PON1, either the Arg192 or Gln192 isozyme, would offer protection against the toxicity of OP s in PON1-/- mice; (3) determine the acute toxicity of OPs in developing mice; (4) assess the developmental neurotoxicity of OPs in wild type and PON1- knockout mice; and (5) determine the PON1 status in infants and children.
Progress Summary:During the second year studies were carried out to further understand the in vivo function of PON1 for detoxifying OP insecticides utilizing transgenic mice. We had previously found that PON1 -/- mice were extremely sensitive to the toxicity of diazoxon but not of paraoxon. In vitro assays indicated that the PON1R192 isoform hydrolyzed diazoxon less rapidly than did the PONQ192 isoform. In vivo experiments, where PON1-knockout mice received the same amount of either PON1192 isoform via i.p. injection 4 hours prior to exposure, showed that both isoforms provided a similar degree of protection against diazoxon, while PON1R192 conferred better protection against chlorpyrifos-oxon than PON1Q192. Injection of purified rabbit PON1 or either human PON1192 isoform did not protect PON1-knockout mice from paraoxon toxicity nor did over-expression of the human PON1R192 transgene in wild type mice. Kinetic analysis of the two PON1192 isoforms revealed that the catalytic efficiency (Vmax/Km) determines the in vivo efficacy of PON1 for organophosphate detoxication. The results indicate that PON1 plays a major role in the detoxication of diazoxon and chlorpyrifos oxon but not paraoxon.
Since a wide range of expression levels of PON1 among individuals has been observed, we examined the promoter region of PON1 for genetic factors that might affect PON1 activity levels. We conducted a deletion analysis of the PON1 promoter region in transient transfection assays and found that cell-type promoter elements for liver and kidney are present in the first 200 bases upstream of the coding sequence. Sequence analysis of DNA from several individuals identified five polymorphisms in the first 1000 bases upstream of the coding region at positions –108, –126, –162, –832, and –909. The two polymorphisms at –126 and –832 have no apparent effect on expression level in the reporter gene assay. The polymorphisms at position –909, –162, and –108 each have approximately a two-fold effect on expression level. The expression level effects of the three polymorphisms does not appear to be additive and may depend on context effects.
Finally, taking advantage of a study on children diagnosed with phenylketonuria, a number of serum samples are being collected from infants and children. These samples are analyzed for PON1 activity using different substrates. Preliminary results show an increase of serum PON1 activity with age.
Significance
Animal studies done so far are progressing toward the final goal of the project (i.e., the role of PON1 in the developmental neurotoxicity of OPs). The human study will provide unique information on the developmental profile of serum PON1 in children. Overall, the significance of the project lies in the understanding of whether the combined genetic background and developmental expression of PON1 may render children more at risk for OP toxicity and neurotoxicity.
Future Activities:In the third year of this project, we plan to continue pursuing the Specific Aims. In particular, we will continue the collection of samples and the assays of human serum from infants and children. Furthermore, we will conduct studies toward the goals of Aim 3. For this purpose, we will assess the developmental profile of serum and liver PON1 activity with different substrates and of liver PON1 mRNA in C57 mice, which are of the same strain as the PON1-/- mice we utilize. We will then conduct a series of studies to determine the acute effects of various doses of OPs in mice of different ages (postnatal days 4, 7, 10, 15, 21, and 28), comparing wild type and PON1- knockout mice. The end-point in these experiments will be, as always, inhibition of brain and diaphragm acetylcholinesterase activity. These experiments will also provide important information (e.g., doses of OPs) for the studies of developmental neurotoxicity.
Journal Articles on this Report: 4 Displayed | Download in RIS Format
| Other subproject views: | All 14 publications | 14 publications in selected types | All 11 journal articles |
| Other center views: | All 83 publications | 51 publications in selected types | All 47 journal articles |
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Costa LG, Li WF, Richter RJ, Shih DM, Lusis A, Furlong CE. The role of paraoxonase (PON1) in the detoxication of organophosphates and its human polymorphism. Chemico-Biological Interactions 1999;119-120:429-438. |
R826886C002 (2000) R831709 (2005) |
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Costa LG. The emerging field of ecogenetics. NeuroToxicology 2000;21(1-2):85-89. |
R826886C002 (2000) R831709 (2005) |
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Furlong CE, Li WF, Richter RJ, Shih DM, Lusis AJ, Alleva E, Costa LG. Genetic and temporal determinants of pesticide sensitivity: role of paraoxonase (PON1). NeuroToxicology 2000;21(1-2):91-100. |
R826886C002 (2000) R826886C002 (2001) R831709 (2005) |
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Li WF, Costa LG, Richter RJ, Hagen T, Shih DM, Tward A, Lusis AJ, Furlong CE. Catalytic efficiency determines the in- vivo efficacy of PON1 for detoxifying organophosphorus compounds. Pharmacogenetics 2000;10(9):767-779. |
R826886C002 (2000) R826886C002 (2001) R831709 (2005) |
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children, health, pesticide, exposure. , Toxics, Geographic Area, Scientific Discipline, Health, RFA, Susceptibility/Sensitive Population/Genetic Susceptibility, Risk Assessments, genetic susceptability, Health Risk Assessment, Children's Health, Biochemistry, pesticides, Environmental Chemistry, State, exposure assessment, insecticides, public health, environmental hazard exposures, health effects, Washington (WA), intervention, environmental toxicant, assessment of exposure, farmworkers, exposure prone behavior, harmful environmental agents, pesticide residues, agricultural community, community-based intervention, exposure pathways, pesticide exposure, sensitive populations, biological response, children, environmental risks, exposure, children's vulnerablity, environmental health hazard, dermal exposure, human exposure, Human Health Risk Assessment
Relevant Websites:
http://depts.washington.edu/chc/
Progress and Final Reports:
Original Abstract
2001 Progress Report
2002 Progress Report
Main Center Abstract and Reports:
R826886 University of Washington
Subprojects under this Center:
(EPA does not fund or establish subprojects; EPA awards and manages the overall grant for this center).
R826886C001 Molecular Mechanisms of Pesticide-Induced Developmental Toxicity
R826886C002 Genetic Susceptibility to Pesticides (Paraoxonase Polymorphism or PON1 Study)
R826886C003 Community-Based Participatory Research Project
R826886C004 Pesticide Exposure Pathways Research Project