lri annual science meeting - american chemistry · pdf filehoneywell, a member of acc’s...

May 5-6, 2004Hotel Inter-Continental

Miami, Florida

AMERICAN CHEMISTRY COUNCIL’S

PROGRAM AND ABSTRACTS

LRI Annual Science Meeting

Stay Ahead of the CURVE

Long-Range Research Initiative Annual Science Meeting May 5-6, 2004

Miami Hotel InterContinental, Miami, FL

WELCOME

On behalf of the American Chemistry Council’s (ACC) Strategic Science Team, we would like to welcome you to the second Annual Science Meeting of the ACC’s Long-Range Research Initiative (LRI). This marks the fifth-year anniversary of the LRI. Through the LRI, the chemical industry sponsors independent research to expand scientific knowledge of the potential impacts that chemicals may have on the health of human and wildlife populations and the environment. The LRI Annual Science Meetings highlight LRI-funded research projects underway and focus on significant, overarching research issues. The 2004 LRI Science Meeting explores the future of research on the impacts of chemicals on development and reproduction. LRI-funded research is based on highest scientific merit and relevance to the program's mission and objectives, with projects sponsored at leading research institutes, government agencies, academic institutes, and research organizations. For example, CIIT Centers for Health Research conducts research in the areas of its expertise, specifically research to advance the science of human health risk assessment. All the research is performed independently. The investigators are responsible for experi-mental design, selection of chemicals, conduct of the work, and publication of the results in peer-reviewed scientific journals. All the results are made public, regardless of the outcome. The LRI oper-ates under the auspices of the International Council of Chem-ical Associations (ICCA). The ACC, the European Chemical Industry Council (Cefic), and the Japan Chemical Industry Association (JCIA), through the ICCA, have similar principles and goals and coordinate research projects and research strategies in key areas. The LRI involves industry and non-industry scientists from all over the world. This meeting presents the projects spon-sored through ACC’s LRI Program.

CONTENTS Welcome .............................................................. 1

Meeting Highlights................................................. 3

Agenda................................................................. 5

Day I Presentations ............................................... 7

Day II Presentations.............................................13

Poster Sessions and Map ......................................19

Poster Abstracts ...................................................27

Author Index........................................................63

2 2004 LRI Annual Science Meeting

We hope this meeting will provide a forum for you to meet many of the scientists and learn more about LRI projects underway. Carol J. Henry, Ph.D., D.A.B.T. James S. Bus, Ph.D., D.A.B.T. ACC Vice President, Science and Research Dow Chemical Company Co-Leader, LRI Strategic Science Team Co-Leader, LRI Strategic Science Team

Meeting Organizing Committee Members

Dr. Jack Moore, Hollyhouse, Inc.

Dr. Robert Kavlock, Consultant

Dr. Judy Graham, American Chemistry Council

Cheryl Morton, American Chemistry Council

2004 LRI Annual Science Meeting 3 Highlights

MEETING HIGHLIGHTS • Welcome and Introduction: Dr. Nance Dicciani, President & CEO, Specialty Materials,

Honeywell, a member of ACC’s Board of Directors and Executive Committee, and chair of the ACC Board Committee with oversight responsibilities for the LRI, will kick off the LRI meeting with her talk, Enhancing the Foundation for Decision-Making Through Research and Partnerships, at a joint session with the Responsible Care® Conference.

• Discussions on Meeting Theme: Envisioning the Future of Research on the Impacts of Chemicals on Development and Reproduction. Developmental defects from a wide variety of causes, known and unknown, are a significant human health problem. In a report entitled, Scientific Frontiers in Developmental Toxicology and Risk Assessment issued by the National Academy of Sciences, the NAS evaluates current approaches used to assess risk for developmental defects and identifies areas of uncertainty in those approaches. Speakers at the LRI Annual Science Meeting will address the report as follows:

Keynote Speaker: Dr. John Gerhart (University of California at Berkeley), a preeminent developmental biologist, will be the keynote speaker. Dr. Gerhart was Vice-Chair of the NAS Committee on Developmental Toxicology, which wrote the report. He will present the findings from the Academy report. Panel Discussion: How Close Are We to the “Frontiers” of Developmental Toxicology? Four esteemed scientists from government, academe, and industry will participate in a panel discussion following Dr. Gerhart’s presentation, to explore how new approaches and information are being applied to the field of developmental toxicology.

• Poster Sessions: Thirty-four scientists conducting research sponsored by the LRI will

present posters and be available to discuss their work during two poster sessions. This is a unique opportunity to meet the researchers and engage in discussion with them.

• LRI Research Showcases: Six investigators will make formal in-depth presentations to

highlight the progress and significance of their LRI-funded projects. Selections are drawn from hot topics related to endocrine system effects, susceptibility factors, fetal/child development, systems biology, fetal origins of adult disease, and genomics. Human health and ecosystems programs are included.

Highlights 4 2004 LRI Annual Science Meeting

NOTES

2004 LRI Annual Science Meeting 5 Agenda

AGENDA

Day I – Wednesday, May 5, 2004

9:00 AM – 5:00 PM Registration

11:00 AM – 12:00 PM Welcome and Introduction: Enhancing the Foundation for Decision-Making Through Research and Partnerships Nance Dicciani, President and CEO, Specialty Materials, Honeywell and ACC Board Research Committee Chair

Versailles

12:00 PM – 1:30 PM Lunch Chopin Room

1:30 PM – 2:10 PM Long-Range Research Initiative: Staying Ahead of the Curve Chair: Carol Henry, American Chemistry Council (ACC)

Trianon Room

• LRI Overview – Carol Henry, ACC • LRI Research Programs and Projects – Jim Bus, Dow

Chemical

2:10 PM – 3:00 PM Keynote Speech: The National Academy of Sciences (NAS) Report - Scientific Frontiers in Developmental Toxicology and Risk Assessment John Gerhart, University of California, Berkeley

Trianon Room

3:00 PM – 3:30 PM Break

3:30 PM – 4:45 PM Panel: How Close Are We to the “Frontiers” of Developmental Toxicology? Chair: John Gerhart, University of California, Berkeley

• Sid Hunter, U.S. Environmental Protection Agency • George Daston, Procter & Gamble • Tom Knudsen, University of Louisville • Philip Mirkes, University of Washington

Trianon Room

5:15 PM – 7:00 PM Poster Session I and Reception Chair: Cheryl Morton, ACC

Mezzanine

7:00 PM – 9:00 PM Dinner Bayfront

Agenda 6 2004 LRI Annual Science Meeting

AGENDA

Day II – Thursday, May 6, 2004

7:30 AM – 9:00 AM Continental Breakfast and Poster Viewing Mezzanine

9:00 AM – 10:30 AM LRI Research Showcase I Chair: Tina Bahadori, ACC • Sexually Dimorphic Forebrain Development: A

Potential Target for Endocrine Disruption Eva Polston, CIIT Centers for Health Research • Fetal Origins of Adult Disease: Do In Utero

Exposures to Environmental Toxicants Play a Role? Warren Foster, McMaster University

• Effects of a Potent Estrogen Mimic on Aquatic

Populations: Results of a Whole-Lake Experiment Vince Palace, Department of Fisheries and Oceans, Canada

Trianon Room

10:30 AM – 12:15 PM Poster Session II Chair: Cheryl Morton, ACC

Mezzanine

12:15 PM – 1:15 PM Lunch Chopin Room

1:15 PM – 2:45 PM LRI Research Showcase II Chair: Tina Bahadori, ACC • Systems Biology and Dose-Response Relationships

Mel Andersen, CIIT Centers for Health Research • Application of Functional Genomics Research to

Toxicology Russell Thomas, CIIT Centers for Health Research • Investigating How Age and Gender Affect Internal

Doses of Chemicals Harvey Clewell, ENVIRON

Trianon Room

2:45 PM – 3:00 PM Closing Remarks Jim Bus, Dow Chemical

Trianon Room

DAY I PRESENTATIONS

2003 LRI Annual Science Meeting Day I Presentations 7

Welcome and Introduction

Speaker Biographies

Dr. Nance Dicciani, Specialty Materials, Honeywell

Enhancing the Foundation for Decision-Making Through Research and Partnerships Dr. Nance Dicciani is President and CEO of Specialty Materials, a strategic business group of Honeywell that serves as a world leader in the production of nylon, polyester, polyethylene, fluorocarbons, caprolactam, electronic materials, and specialty chemicals. Specialty Materials' headquarters are located in Morristown, New Jersey. Nance joined the company in November of 2001 from Rohm and Haas where she held the position of Senior Vice President and Business Group Executive of Chemical Specialties and Director, European Region. She was responsible for business strategy and worldwide operations of five business units (Consumer and Industrial Specialties, Organic Specialties, Inorganic Specialties, Ion Exchange Resins, and Primenes). As Regional Director, Europe, she was responsible for the company's operations and infrastructure in Europe, the Middle East and Africa. Nance joined Rohm and Haas in 1991 and during her tenure served as Vice President and General Manager of the Petroleum Chemicals Division and headed the company’s worldwide Monomers business. Prior to joining Rohm and Haas, Nance worked for Air Products and Chemicals, Inc., for 14 years where she held positions of increasing responsibility in research, engineering and research management. In 1986, she became general manager for a stand-alone manufacturer of specialty cryogenic containers. Additionally, she held leadership roles in their Chemicals Groups, Industrial Chemicals and Specialty Chemicals Divisions. Nance holds a B.S. degree in Chemical Engineering from Villanova University, a M.S. degree in Chemical Engineering from the University of Virginia and a Ph.D. in Chemical Engineering from the University of Pennsylvania. She also received an M.B.A. from the Wharton School of the University of Pennsylvania. She currently serves as a Vice President of SCI, the Society of Chemical Industry. Nance previously served on the Board of Advisors of Cefic, the Board of Directors of PP&L Resources Inc., and the Board of Trustees of Villanova University.

LRI – Staying Ahead of the Curve

Dr. Carol Henry, American Chemistry Council

Overview Long-Range Research Initiative (LRI)

Dr. Henry serves as Vice President for Science and Research at ACC. She directs and manages the ACC’s multi-million dollar Long-Range Research Initiative that is designed to study the potential impacts of chemicals on health and the environment. She received her undergraduate degree in Chemistry from the University of Minnesota and Doctorate in Microbiology from the University of Pittsburgh. In addition, Dr. Henry held postdoctoral fellowships in biochemistry at the Max Planck Institute in Germany, in biology at Princeton University, and biochemistry/cancer research at the Sloan Kettering Institute. Prior to joining the American Chemistry Council in May 1999, Dr. Henry served as director of the Health and Environmental Sciences Department of the American Petroleum Institute (API) and as API’s chief scientist. Before joining API in 1997, Dr. Henry completed five years of public service, serving as Associate Deputy Assistant Secretary for Science and Risk Policy at the U.S. Department of Energy, and as director of the Office of Environmental Health Hazard Assessment at the California Environmental Protection Agency. Prior to that appointment, she was Executive Director of the International Life Sciences Institute’s Risk Science Institute. A diplomate of the American Board of Toxicology, Dr. Henry is a member of the American College of Toxicology, of which she has been president, the Society of Toxicology, the American Association for

DAY I PRESENTATIONS

Day I Presentations 2003 LRI Annual Science Meeting 8

Cancer Research, the American Association for the Advancement of Science, and the American Chemical Society. She served on the Board of Scientific Counselors of the National Toxicology Program and the EPA Clean Air Act Science Advisory Committee’s Blue Ribbon Panel on Oxygenates in Gasoline. She has served on the Board on Environmental Studies and Toxicology of the National Research Council, the Roundtable on Environmental Health Sciences, Research, and Medicine of the Institute of Medicine, the Chemical Sciences Roundtable of the National Research Council, the DOD/EPA/DOE Strategic Environmental Research and Development Program’s Science Advisory Board, and as a Consultant to the EPA’s Science Advisory Board’s Executive Committee.

Dr. James Bus, The Dow Chemical Company

LRI Research Programs and Projects Dr. Bus currently holds the position of Director of External Technology and serves as a member of the Leadership Team in the Toxicology and Environmental Research and Consulting group at the Dow Chemical Company in Midland, Michigan. Prior to joining Dow Chemical in 1989, he held positions of Associate Director of Toxicology and Director of Drug Metabolism at the Upjohn Company (1986-1989), Research Scientist at the Chemical Industry Institute of Toxicology (1977-1986), and Assistant Professor of Toxicology at the University of Cincinnati (1975-1977). Dr. Bus currently is Adjunct Professor of Pharmacology and Toxicology at Michigan State University and previously has held the position of Adjunct Associate Professor of Toxicology at the University of North Carolina.

In 1996-1997 Dr. Bus served as President of the Society of Toxicology, a 4,000 member scientific society, and in 1986-1987 as President of the American Board of Toxicology, an organization which credentials practicing toxicologists. He has served on the U.S. Environmental Protection Agency Office of Research and Development Board of Scientific Counselors (1996-2003), the National Toxicology Program Board of Scientific Counselors, Bioassay Review Subcommittee (1996-2000), the ACGIH Chemical Substances TLV Committee (1993-2002), as a Director of the International Union of Toxicology (1998-2001), and on the Board of Trustees of the International Life Sciences Institute (ILSI) Health and Environmental Sciences Institute (2002-2004; currently Past-Chair of the Emerging Issues Committee). Dr. Bus is Co-Chair of the American Chemistry Council LRI. He has also been a member of the Board of Directors (1997-2003) and is currently Co-Chair of the Science Program Committee of the CIIT Centers for Health Research, a research institute founded by and primarily supported by the chemical industry.

Dr. Bus received his Ph.D. in Pharmacology from Michigan State University in 1975 and a B.S. degree in Medicinal Chemistry from the University of Michigan in 1971. His research interests have focused on mechanisms of chemical toxicity. In 1987 he was the recipient of the Society of Toxicology Achievement Award, granted for outstanding contributions to the science of toxicology by a young scientist. In 1999 he received the Robert A. Scala Award from the Environmental and Occupational Health Sciences Institute (Rutgers University) for outstanding contributions to toxicology by an industrial scientist, and in 2001 the Distinguished Alumnus Award from the Michigan State University Department of Pharmacology and Toxicology. He has authored or coauthored over 90 research papers, reviews, and books.

DAY I PRESENTATIONS

2003 LRI Annual Science Meeting Day I Presentations 9

Keynote Speaker

Dr. John Gerhart, The University of California, Berkeley

The National Academy of Sciences Report – Scientific Frontiers in Developmental Toxicology and Risk Assessment

Abstract Cell-cell signaling pervades all aspects of development, not just of vertebrates but all

animals (metazoa). It is an evolutionary innovation of the multicellular kingdoms, animals, plants, and fungi, which diverged about 1.2 billion years ago from a common ancestor descended from a lineage of unicellular eukaryotes. In metazoa at least 17 kinds of signal transduction pathways operate, each distinguished by its transduction intermediates. Five kinds predominate in early embryonic development, namely, the Wnt, TGF-b, Hedgehog, RTK, and Notch pathways. Five more are used in late development, and seven more in the functions of differentiated cells. The pathways must have evolved and become conserved in pre-Cambrian times before the divergence of basal members of most of the modern phyla. The responses made by cells to intercellular signals in development and physiology are also conserved from this time. These responses include cell proliferation, secretion, motility, and transcription. Many responses date back even to the unicellular eukaryotic forms of two billion years ago and some even to prokaryotic cells of three billion years ago. Individual developmental processes consist of specifically associated signals and responses, and many of these processes (their networks of signals and responses) have been conserved widely in metazoa, shared for example by insects and mammals. Genome sequencing of disparate animals has confirmed and deepened this recognition of conserved genes and encoded cell components. The study of model organisms, even those of non-vertebrate groups, is expected to continue to contribute greatly to the understanding of mammalian development and to offer opportunities to analyze the effects of toxicants on development, as well as opportunities to devise incisive assays for toxicants.

Biography John Gerhart has served on the faculty of the University of California, Berkeley since 1962, in the Department of Cell and Molecular Biology. He currently holds an emeritus position, Professor of the Graduate School. His research has concerned, in successive decades, allosteric enzymes, the cell cycle, the early development of the amphibian Xenopus laevis, and most recently comparative studies on the development of hemichordates, the phylum most closely related to ours, the chordates. His work on development in the 1990's concerned particularly the formation and function of Spemann's organizer. This was the time when researchers were isolating and identifying inducers for the first time, as well as discovering the means of response of cells to inducers. The similarities of development across the vertebrates, the chordates, and even all bilateral animals was an emerging realization in that period, and Dr. Gerhart has written on this subject of conserved molecular mechanisms of development and its implications for understanding mechanisms of animal evolution. He is a member of the National Academy of Sciences and has served as co-Chair with Dr. Elaine Faustman on the National Research Council's committee to review "Scientific Frontiers in Developmental Toxicology and Risk Assessment" (National Academy Press, 2000; 327pp.). He gave the Warkany Lecture in 1998.

DAY I PANEL DISCUSSION

Day I Panel Discussion 10 2003 LRI Annual Science Meeting

How Close Are We to the “Frontiers” of Developmental Toxicology?

Panel Member Biographies

Dr. Sid Hunter, U.S. Environmental Protection Agency Dr. Hunter’s degrees are from Hampden Sydney College (B.S. Chemistry, 1980), Old Dominion University (M.S. Toxicology, Dept. Chemical Sciences, 1983) and University of North Carolina at Chapel Hill (Ph.D., Anatomy and Embryology, 1986). Dr. Hunter continued his research training as a postdoctoral fellow and Research Assistant Professor at UNC-CH in the Department of Cell Biology and Anatomy. In 1990 Dr. Hunter moved to the National Institute of Environmental Health Sciences in the National Toxicology Program where he work as a Developmental Toxicologist under Dr. Bern Schwetz. Dr. Hunter moved to the U.S. EPA in the Reproductive Toxicology Division in 1993 that is directed by Dr. Robert Kavlock. Dr. Hunter’s research in the area of developmental toxicology has focused on the potential for disinfection by-products to produce developmental effects and the mechanisms responsible for those defects during early craniofacial development. Dr. Hunter is Adjunct Assistant Professor in Toxicology at UNC-CH where he lectures in developmental toxicology and co-directs a course in Embryology and Teratology. Dr. Hunter has mentored three Masters and Ph.D. students and three postdoctoral trainees. He has served on two NIH Study sections and several peer review panels and committees. Dr. Hunter previously served as a Section Editor for Teratology. Dr. Hunter’s laboratory continues to study mechanisms and morphogenetic events during early craniofacial development using a variety of genomic and proteomic tools.

Dr. George Daston, Procter & Gamble George Daston has spent his entire career in research to understand the effects of exogenous chemicals on the developing embryo, fetus, and child. His research interests include teratogenic mechanisms, in vitro methodologies, and risk assessment. He has published over 90 peer-reviewed articles, reviews, and book chapters, and has edited three books. His most recent research includes (1) toxicant-nutrient (especially zinc) and maternal-embryonal interactions in developmental toxicity; (2) genomic approaches to endocrine disrupter screening; and (3) improvements in risk assessment methodology for non-cancer endpoints. Dr. Daston received his B.S. and Ph.D. from the University of Miami and received post-doctoral training at the U.S. EPA’s laboratories in Research Triangle Park, North Carolina. He was an Assistant Professor of Biological Sciences at the University of Wisconsin-Milwaukee from 1983-85. He joined Procter & Gamble in 1985, where he has risen to the rank of Research Fellow. Dr. Daston's professional activities include serving as Chair of the Developmental and Reproductive Toxicology Technical Committee of ILSI-Health Effects Sciences Institute; President (1994-95) of the Society of Toxicology's Reproductive and Developmental Toxicology Specialty Section; President (1999-2000) of the Teratology Society; member of the National Academy of Sciences Board on Environmental Studies and Toxicology (1995-98); Councilor of the Society of Toxicology (2001-2003); member of the EPA Board of Scientific Counselors; member of the U.S. National Toxicology Program Board of Scientific Counselors; and member of EPA's Endocrine Disrupter Screening and Testing Advisory Committee (EDSTAC). Dr. Daston has served on the organizing committees for: ILSI/EPA/AIHC workshops on benchmark dose methodology and human variability in toxic response; an EPA workshop on endocrine-mediated toxicity; and as chair or co-chair of: AIHC/EPA workshop on Leydig cell tumors, an ILSI/EPA workshop on interpreting reproductive toxicity endpoints, and NIEHS workshops on the state of validation of the FETAX assay for teratogen screening and receptor binding and transcriptional activation assays for estrogens and androgens. Dr. Daston is Editor-in-Chief of Birth Defects Research: Developmental and Reprodutive Toxicology, an Associate Editor of Toxicological Sciences, on the Editorial Board of Human and Ecological

DAY I PANEL DISCUSSION

2003 LRI Annual Science Meeting Day I Panel Discussion 11

Risk Assessment, and an ad hoc reviewer for Journal of Nutrition, FASEB Journal and other journals. Dr. Daston is an adjunct professor in the Department of Pediatrics and Developmental Biology Program at the University of Cincinnati and Children's Hospital Research Foundation. Dr. Daston was a Visiting Scientist at the Salk Institute, Molecular Neurobiology Laboratory, 1993-94. Dr. Daston was elected a Fellow of AAAS in 1999.

Dr. Thomas Knudsen, University of Louisville Dr. Thomas Knudsen’s degrees are from Albright College (B.S., Biology, 1976) and Thomas Jefferson University (Ph.D., Anatomy, 1981), and he obtained specialized research training as a postdoctoral fellow in cell biology at the Children’s Hospital of Cincinnati and in developmental biology at Emory University in Atlanta. In 1986 Dr. Knudsen was appointed Assistant Professor at E. Tennessee State University. He was recruited back to Philadelphia in 1990 and was promoted through the ranks to Professor (tenured) in the Department of Pathology, Anatomy, and Cell Biology at Jefferson Medical College of Thomas Jefferson University. For his research on birth defects, Dr. Knudsen has received over $7M in federal research grants from the National Institutes of Health (NIH) and U.S. EPA. His work entails understanding the regulation and control of mitochondrial mtDNA genomic expression during early embryo development. His educational activities at Jefferson included human medical gross anatomy, embryology and teratology, and bioinformatics; he mentored eight Masters and Ph.D. students, and directed a training grant in birth defects research from the NIH - National Institute of Environmental Health Sciences. Dr. Knudsen belongs to seven professional societies, has served numerous NIH study sections, and has served numerous university and national committees. He is presently Editor-in-Chief of Reproductive Toxicology, the official journal of the European Teratology Society. Dr. Knudsen joined the faculty at the University of Louisville School of Dentistry in December 2003 as a tenured Professor of Molecular, Cellular, and Craniofacial Biology. Part of his responsibilities is to establish and direct a Systems Analysis Laboratory at the Birth Defects Center, which applies functional genomics and computational biology to solve basic problems in birth defects research.

Dr. Philip Mirkes, University of Washington Philip E. Mirkes received his Ph.D. in Zoology from the University of Michigan in 1970 and then completed a one-year postdoctoral fellowship with Dr. Arthur Whiteley in the Department of Zoology, followed by a two-year fellowship with Dr. David Stadler in the Department of Genetics at the University of Washington. After a brief tenure as an Assistant Professor in the Department of Biology at the University of South Carolina, Dr. Mirkes was awarded a career development award in toxicology and returned to the University of Washington to join the faculty in the Department of Pediatrics where he is now a Research Professor. Dr. Mirkes’ research in developmental toxicology has focused on the molecular mechanisms by which teratogens, primarily hyperthermia and cyclophosphamide, disrupt normal development. His recent research has focused on the molecular mechanisms by which teratogens activate the apoptotic pathway in mouse embryos and the role of this cell death in the etiology of specific malformations, e.g., neural tube defects. In addition, his research has also focused on the role of heat shock proteins as modulators of developmental toxicity. Dr. Mirkes is a member of the Teratology Society and served as President from 1997-98. Since January 1 of 2003, Dr. Mirkes has been editor of Birth Defects Research: Clinical and Molecular Teratology. On July 1, 2004, Dr. Mirkes will move to Texas A&M University where he will assume the directorship of the NIEHS-funded Center for Environmental and Rural Health.

Day I Presentations

Day I Presentations 12 2003 LRI Annual Science Meeting

Poster Session Organization

Cheryl Morton, American Chemistry Council

Meeting Organizer and Chair of Poster Sessions

Cheryl Morton serves as Managing Director of the LRI. Prior to joining the Council, Ms. Morton was the Director of Regulatory and Technical Affairs at the Synthetic Organic Chemical Manufacturers Association (SOCMA). At SOCMA, she managed the government relations’ staff and served as SOCMA’s expert on compliance assistance and regulatory advocacy related to the Toxic Substances Control Act (TSCA). Earlier in her career, she was Director of Environmental Affairs at a public relations firm in Washington, D.C., where she managed three associations focusing on air, water, and hazardous waste. Also, she worked on U.S. Environmental Protection Agency contract projects at ENVIRON and Booz, Allen and Hamilton. Ms. Morton received a B.S., Environmental Science and B.S., Chemistry, George Washington University, where she has taken selected graduate courses in the Regulatory Affairs Program.

DAY II PRESENTATIONS

2003 LRI Annual Science Meeting 13 Day II Presentations

LRI Research Showcase I

Dr. Tina Bahadori, American Chemistry Council

Chair for Research Showcase Tina Bahadori, Sc.D., is a Senior Scientist/Senior Director of the Long-Range Research Initiative (LRI) at the American Chemistry Council with primary responsibilities for research programs in the areas of Exposure and Risk Assessment. Prior to joining the Council, she was the Manager of Air Quality Health Integrated Programs at Electric Power Research Institute (EPRI), where her charge was to bring together lead scientists from the areas of atmospheric science, epidemiology, exposure assessment, and health/toxicology to examine the fundamental health effects of air pollution. At Arthur D. Little, Inc., she was a Consultant in the Risk Assessment Unit, where she assisted clients with technical problems related to management of environment, health, and safety exposures. Dr. Bahadori is on the Chemical Exposure Work Group of the National Child Study and has been a member of numerous peer review groups. She is on the Board of the International Society of Exposure Analysis (ISEA). She holds a Sc.D. in Environmental Science and Engineering from the Harvard University School of Public Health. She also received a M.S. in Chemical Engineering and Technology and Policy and a B.S. in Chemical Engineering from MIT.

Dr. Eva Polston, CIIT Centers for Health Research

Sexually Dimorphic Forebrain Development: A Potential Target for Endocrine Disruption

Abstract Exposure of the perinatal mammalian brain to estrogen has profound consequences on the structural and functional development of the brain circuits that mediate reproduction. In males, estradiol (E2) is synthesized de novo in brain from an early surge of fetal/neonatal testosterone. The developing female brain receives little exposure to estradiol, thus remains protected from the masculinizing effects of the hormone. Because the brains of both sexes are sensitive to E2 exposure during the perinatal critical period, chemicals that influence the levels of endogenous E2 or that act as agonists or antagonists at the estrogen receptor may significantly alter normal brain development. Today, little is known about the specific mechanisms by which estrogen exposures of these brain tissues regulate cellular level responses such as apoptosis or cell survival and proliferation. The long-term goal of our research is to understand the risks posed by perinatal exposure to endocrine-active compounds (EACs) on brain development. Our initial studies will determine the doses of estradiol and estrogenic EACs that are required to perturb proper sexual differentiation of brain morphology. Studies will be devoted to developing dose-response curves for brain masculinization in the neonatal rat pup of both sexes by assessing estrogen receptor binding and activation in brain and correlating these parameters to cellular level responses and to altered reproductive function.

Biography Dr. Eva K. Polston has recently joined the Division of Biological Sciences at the CIIT Centers for Health Research (CIIT-CHR) as an Assistant Investigator. Dr. Polston received a B.A. degree in Psychobiology from Oberlin College in 1986 and a Ph.D. degree in Neuroscience from Boston University in 1997. She was a postdoctoral fellow in behavioral neuroscience at Dartmouth College from 1997 to 1999 and recently completed a postdoctoral fellowship in developmental hypothalamic neuroanatomy at the Oregon National Primate Research Center. The focus of Dr. Polston’s research in recent years has been on the hormone-dependent development of sexually dimorphic forebrain circuits that


Day II Presentations 14 2003 LRI Annual Science Meeting

control reproductive function. Her work investigates how estrogen actions in the neonatal brain have a permanent ‘masculinizing’ effect that structurally and functionally differentiates reproductive neuroendocrine and behavioral systems. At CIIT-CHR, Dr. Polston’s research focuses on examining whether exposure to endocrine-active compounds in the environment may disrupt these hormone-mediated developmental processes in the brain and determining the levels of exposure that would pose risks to exposed individuals.

Dr. Warren Foster, McMaster University

Fetal Origins of Adult Disease: Do In Utero Exposures to Environmental Toxicants Play a Role?

Abstract Increasingly fetal insult has been linked with neonatal and adult diseases such as obesity, type-II diabetes, and cardiovascular disease. Reports in the literature of an increase in the prevalence of developmental abnormalities of the male reproductive tract, hormone dependent cancers, and asthma have given rise to concerns that in utero exposure to environmental contaminants may be associated with adverse health outcomes in children and adults. However, numerous limitations of contemporary studies preclude establishment of a link between exposure to any environmental contaminant or group of contaminants and adverse health outcomes in the human population. A major limitation of studies to date is accurate assessment of in utero exposure to environmental contaminants. Furthermore, exposure studies have tended to measure chemical contaminants without consideration of potential mechanism of action for the adverse outcome of concern. We hypothesize that exposure to estrogenic agents in the environment are too low to induce adverse health outcomes in the human population but exposure to aryl hydrocarbon receptor agonists could potentially change maternal thyroid function and contribute to developmental abnormalities in the fetus and neonate. To address the hypothesis, 310 pregnant women attending a prenatal diagnosis clinic at McMaster University Medical Centre were invited to participate in this study from which 150 women were recruited. The study was conducted in accordance with the McMaster University ethics committee approval and all participants completed signed consent, before completing intake, food frequency, and obstetrical history questionnaires. The mean age of women enrolling in this study was 38.0 ± 0.2 years (range 34-44 years). A single 20 ml blood sample was collected during the second trimester prenatal visit from each woman and allowed to clot at 4° C, serum collected, and aliquoted for subsequent analyses. Serum samples were subsequently analyzed for aryl hydrocarbon receptor activity using an established chemically activated luciferase expression assay (CALUX). Maternal thyroid function was assessed by commercially available radioimmunoassays for thyroid stimulating hormone (TSH) and thyroxine (T4) as well as the presence of thyroid peroxidase antibodies (TPO). Birth weight was recorded for each neonate as were clinical assessments of neonatal development. Preliminary CALUX results reveal mean (± SEM) serum levels of 0.34 ± 0.01 and range of 0.19 to 0.73 TEQ pg 2,3,7,8 tetrachlorodibenzo-p-dioxin (TCDD)/g serum lipid. Of this study cohort 6 of 150 women (4%) were clinically hypothyroid (TSH > 7.0 µIU/ml and T4 < 61.7 nmol/L) and 12/150 women (8%) were classified with sub-clinical hypothyroidism (TSH > 4.5 µIU/ml and T4 < 57.9 nmol/L). Five neonates were classified as low birth weight babies (less than 2500 g/birth). Multiple regression analysis of our preliminary results failed to demonstrate a statistically significant relationship between maternal serum TCDD TEQ level, thyroid hormone function, and birth weight after controlling for potential confounding factors. Therefore, our preliminary results fail to support the hypothesis and we conclude that in utero exposure to AhR agonists fail to affect maternal thyroid function or induce adverse health effects in neonates. It should be noted however that these are preliminary results and the study is ongoing. Furthermore,



infants continue to be followed and markers of immune function during the first postnatal year are being recorded to assess both immune system development.

Biography A graduate of McMaster University, Dr. Foster is the Director of the Reproductive Biology Division in the Department of Obstetrics & Gynecology at McMaster University. Prior to joining the faculty at McMaster, Dr. Foster served as the Associate Director of Women’s Health and Director of Research at Cedars-Sinai Medical Center in Los Angeles and as the Head of Reproductive Toxicology, Health Canada, Ottawa.

His research interests are broadly focused on investigation of the impact of environmental toxicant exposure on human health. Selected examples of funded studies being conducted by Dr. Foster include the following:

• Explore the relationship between developmental exposure to environmental toxicants

and thyroid and immune function in children; • Determine the relative potency of environmental toxicants on aromatase expression

and activity; • Environmental toxicant induced modulation of tissue remodelling enzyme expression

in estrogen sensitive target tissues; • Dioxins and dioxin-like chemicals in endometriosis: towards understanding mechanism

of action; and

• Biomarkers of breast cancer.

Dr. Vince Palace, Department of Fisheries and Oceans, Canada

Effects of a Synthetic Estrogen on Aquatic Populations: A Whole Ecosystem Study

Abstract The potent estrogen mimic used in birth control pills has been added continuously over three summers (2001-03) to a lake at the Experimental Lakes Area in northwestern Ontario. Effects on the microbial through fish populations have been assessed in this lake during additions, and contrasted to pre-addition and reference lake data. Male minnows exposed to the environmentally-relevant concentrations of 17α-ethynylestradiol (EE2; 5-6 ng/L) produced high concentrations of the egg protein precursor vitellogenin and have developed gonadal abnormalities (including intersex). Female fish are being impacted at the biochemical level with increased production of vitellogenin and at the tissue level with delayed development of oocytes. Population monitoring indicates that the fathead minnow have not successfully spawned in the treated lake in the last two years resulting in a loss of the smaller size classes of this species; no changes in the abundances of the other three species (lake trout, white sucker, pearl dace) have yet been observed. Some impacts on lower-trophic-level organisms have also been found. Female zooplankton of some species produced fewer eggs in both 2001 and 2002, and total zooplankton abundances were higher in 2003 likely due to decreased predation by minnows. Intersex was also observed in mink frog tadpoles caged in the experimental lake but not in the reference systems. Low concentrations of a potent estrogen mimic can have adverse impacts on both vertebrates and invertebrates in aquatic ecosystems either through direct or indirect effects.



Biography Vince Palace has been a research scientist with the Department of Fisheries and Oceans' Habitat Impacts Research Section in Winnipeg, Canada for the past five years. His research is primarily concerned with studying the impacts of domestic and industrial effluents on reproductive physiology of wild fish populations. In addition to the work that he will present today regarding the effects of estrogens, he is also currently involved in projects examining the effects of elevated environmental concentrations of selenium, arsenic, pesticides, and some emerging contaminants of concern, including PBDEs and other flame retardant chemicals.

LRI Research Showcase II

Dr. Melvin Andersen, CIIT Centers for Health Research

Systems Biology and Dose-Response Relationships

Abstract Non-cancer risk assessments utilize safety factors and benchmark doses to estimate human risks. Mechanistic pharmacokinetic modeling of target tissue dosimetry is easily used in these assessments; however, results from mechanistic studies that link initial interactions through to toxic outcomes, are not routinely used for dose-response assessments. New systems biology approaches at CIIT Centers for Health Research (CIIT-CHR) are combining mechanistic research, high-throughput genomic methodologies, computational pathway modeling, and bioinformatics to identify chemical targets and to create mathematical models of signaling networks that are perturbed by chemical exposures. These systems biology approaches offer promise to (1) understand the shape of dose-response curves in regions of low incidence, (2) provide a stronger basis for human health risk assessment for a wide variety of responses, and (3) provide alternatives to defaults. Success in developing these mechanistic dose-response models from a systems biology approach requires work by interdisciplinary teams devoted to unraveling the basic biology of toxic responses and the quantitative description of this biology in simulation models. This talk focuses on specific risk assessment related research by the author on activation of genetic circuits in liver by aryl hydrocarbon (Ah) receptor agonists and examines the types of data needed for dose-response modeling of cell responses. In projects at CIIT-CHR we are implementing systems biology approaches in two programs: developmental toxicity of anti-androgens and respiratory toxicity of irritant vapors in pulmonary epithelium. The systems biology directions for these projects will also be highlighted.

Biography Dr. Andersen is the Director, Computational Biology Division, CIIT Centers for Health Research, Research Triangle Park, NC. His responsibilities include imparting a systems biology emphasis to research on the health effects of environmental chemicals. Dr. Andersen was Professor of Environmental Health at Colorado State from 1999 to 2002. From 1994-1998, Dr. Andersen was Vice-President of the K.S. Crump Group of ICF Kaiser International Consulting. Between 1971 and 1994, he held positions in toxicology research and research management in the federal government (DoD and U.S. EPA) and in private industry (Chemical Industry Institute of Toxicology). Throughout his career, he has developed biologically realistic models of the uptake, distribution, metabolism, and biological effects of drugs and toxic chemicals and applied these models in safety assessments and quantitative health risk assessments. Dr. Andersen is an author or co-author of 240 papers, 30 book chapters, and numerous reports and abstracts. In June 2002, Dr. Andersen was recognized as a ‘highly cited’ scientist by the Institute for Scientific Information. His professional awards include the Herbert Stokinger Award



(American Conference of Industrial Hygienists, 1988), the Kenneth Morgareidge Award (International Life Sciences Institute, 1989), the George Scott Award (Toxicology Forum, 1993), and the Frank R. Blood (1982), Achievement (1984), and Arnold J. Lehman (2004) Awards from the Society of Toxicology. Dr. Andersen is board certified in industrial hygiene and in toxicology. His current research interests are in developing mathematical descriptions of signaling networks in the developing and adult organism and the dose- response and risk assessment implications of perturbations of these control processes.

Dr. Russell Thomas, CIIT Centers for Health Research

Application of Functional Genomics Research to Toxicology

Abstract “Functional genomics” is a new catchphrase that has a number of commonly used definitions. A broad definition of the term is that it is the branch of genomics that studies the genome in a cellular and functional context and is usually characterized by high-throughput, large-scale experimental methods. The overarching goals of the Functional Genomics Research Program at CIIT Centers for Health Research are to develop tools that delineate the cellular signaling pathways involved in toxicity and identify the cause-and-effect relationships among altered genes. To accomplish this objective, the research in this project will focus on three primary areas: Area 1: Develop functional genomic screens to dissect upstream cellular signaling pathways. Dissecting the upstream cellular signaling pathway is important for understanding the potential targets of toxicity, identifying key nodal points in the signaling pathway, and potential crosstalk among pathways known to be involved in other biological endpoints (e.g., development, inflammation). Area 2: Perform functional genomic screens to identify genes involved in susceptibility. Applying gain-of-function and loss-of-function screens in cells exposed to a marginally toxic or sub-toxic dose can identify potential modifiers of susceptibility. Area 3: Use a combination of gene expression analysis and functional genomic tools to map primary, secondary, and tertiary gene expression responses following toxicant exposure. In the cascade model of gene regulation, genes altered early in the process are the primary regulators, while secondary and tertiary changes in expression are more directly involved in the biochemical and structural response to the stimulus. Biologically, the sum of the connections that link the primary regulators with the secondary subnetworks determines the physiological response of an organism or cell to a particular stimulus. Identifying these connections will be an important part of investigating the toxicological mechanisms. Through the application of functional genomic tools, a novel approach to toxicology will be developed to provide critical mechanistic understanding of toxic responses that is essential for predicting the shape of the dose-response curves and supporting risk assessments.

Biography Russell Thomas is an Associate Investigator at CIIT Centers for Health Research in the Division of Biomathematics and Physical Sciences. He is the Director of the Functional Genomics Research Program and the Gene Expression Core Facility. Dr. Thomas completed his Ph.D. in Toxicology at Colorado State University and focused on constructing pharmacokinetic and pharmacodynamic models for the effects of chlorinated benzenes. Following his doctoral studies, Dr. Thomas performed postdoctoral research in molecular biology and genomics at the McArdle Cancer Research Laboratory at the



University of Wisconsin. In addition to the doctoral degree, he has earned a Masters degree in radiation ecology and an undergraduate degree in chemistry. Prior to coming to CIIT-CHR, Dr. Thomas worked in bioinformatics in the biotech industry and was the head of the genomics and toxicology groups at a biopharmaceutical company. At present, Dr. Thomas’ research is focused on developing functional genomic tools for delineating the cellular signaling networks involved in specific toxicological responses and determining the cause-and-effect relationships among altered genes.

Harvey Clewell, ENVIRON Health Sciences Institute

Exposure Research: The LRI Perspective

Abstract The objective of this research was to identify age- and gender-specific differences in physiological and biochemical processes that affect tissue dosimetry and integrate them into a predictive pharmacokinetic (PK) framework. Three case studies were conducted based on information available in the literature. The first case study focused on physiological and biochemical determinants of systemic toxicity, and tested the hypothesis that many of the pharmacokinetic differences observed across age and gender may be attributed to fundamental changes in physiological and biochemical processes. The second case study focused on the impact of age- and gender-specific differences in lung morphology and ventilation rate on both local and systemic inhalation toxicity as a function of the properties of the chemical. The third case study focused on the perinatal period, using physiologically based PK models to evaluate in utero exposure via placental transfer versus infant exposure via lactational transfer to demonstrate critical periods of exposure from a pharmacokinetic perspective. These studies were exercised for multiple compounds to provide information on pharmacokinetic behaviors as a function of the physicochemical properties of a compound. In general, variations in pharmacokinetic dose metrics for a chemical over a lifetime were within a factor of two; however, exceptions were observed in the neonatal period where the dose metrics varied by as much as an order of magnitude from adult values. Perinatal simulations demonstrated the importance of in utero, as compared to neonatal, exposure, even in the case of chemicals with a high potential for lactation transfer.

Biography Harvey J. Clewell III is a professional research manager with over twenty-five years of experience in environmental quality research, toxicology research, and hazardous materials management. He has gained an international reputation for his work in the application of physiologically based pharmacokinetic (PBPK) modeling to chemical risk assessment and pharmaceutical safety assessment. He has played a major role in the first uses of PBPK modeling in cancer and non-cancer assessments by EPA, ATSDR, OSHA, and FDA, for such chemicals as methylene chloride, trichloroethylene, vinyl chloride, and retinoic acid. He served for 20 years as an officer in the U.S. Air Force; his duties included Deputy Director of the Air Force Toxic Hazards Research Unit, Director of Hazardous Materials Safety for the Air Force Aeronautical Systems Center, and consultant to the Air Force Surgeon General on Chemical Risk Assessment. He is currently a Principal with ENVIRON Health Sciences Institute.

POSTER SESSIONS

2004 LRI Annual Science Meeting 19 Poster Abstracts

The LRI research posters are organized according to the three focus areas that were developed to frame the research strategy for the LRI and to stimulate and encourage strategic, multidisciplinary approaches to sponsoring research of highest quality and relevance to the chemical industry. These focus areas are:

Improved Methods: Building the scientific foundation to evaluate the potential risks of chemicals to public health and the environment

• Human health and ecological effects screening and testing methods − Toxicity test methods emphasizing development − Integration and implications of new and emerging approaches in health effects

research − Methods to detect effects on wildlife

• Human exposure methods − Methods for characterizing and estimating exposures − Interpreting and using biomonitoring data

• Interspecies, intraspecies, and target organ determinants of dose-response − Prediction of target tissue dose − Understanding toxicodynamics − Health hazard assessment methodologies

Susceptibility Factors: Identifying vulnerable groups (including children) and characterizing factors that may place them at higher risk

• Human health sensitivity factors • Human exposure factors

Chemicals in the Environment: Understanding how they move and change along pathways from sources to humans and wildlife

• Human exposure assessment and analysis • Ecosystem exposure analysis

Scientists will be presenting their LRI-funded research projects during two poster sessions and will be present at their posters during the following times:

Posters Attended Poster Session I (Wednesday Evening)

Poster Session II (Thursday Morning)

Odd-numbered posters 5:15 - 6:00 PM 11:30 - 12:15 PM

Even-numbered posters 6:15 - 7:00 PM 10:30 - 11:15 AM

The posters may also be viewed during breaks and meals on Wednesday and during breakfast on Thursday. A map indicating the location of the posters by poster number is presented on the following page.

MAP OF POSTER LOCATIONS

Poster Abstracts 20 2004 LRI Annual Science Meeting

POSTER SESSIONS


LRI Research Posters Sorted by Focus Area Themes/Subthemes Poster

No.

Improved Methods: Human Health and Ecological Screening and Testing Methods The 2004 Long-Range Research Initiative (LRI) Portfolio at the CIIT Centers for Health Research: An Overview

F.J. Miller CIIT Centers for Health Research 27

Improved Methods: Toxicity Test Methods Emphasizing Development Statistical Issues in Modeling Pregnancy Outcome Data

G.M. Buck, V.M. Dukic, P.J. Heagerty, T.A. Louis, C.D. Lynch, L.M. Ryan, E.F. Schisterman, A. Trumble, and the Pregnancy Modeling Working Group

National Institute of Child Health and Human Development, University of Chicago, University of Washington, Johns Hopkins University, and Harvard University

10

Developmental Neurotoxicity Testing with Methyl Mercury and Methylazoxy Methanol: Neuropathology Endpoints Reconsidered

D.M.G. de Groot, M.H.M. Bos-Kuijpers, H-J.G. Gundersen, W.S.H. Kaufmann, J.H.C.M. Lammers, B. Pakkenberg, J.P. O’Callaghan, and D.H. Waalkens-Berendsen

TNO Nutrition and Food Research, the University of Aarhus, BASF, Research Laboratory for Stereology & Neuroscience, Copenhagen, DK, NIOSH

18

Comparison of Fetal vs. Adult-induced Immunotoxicity with Cyclosporin-A

I. Hussain, M. Piepenbrink, K. Fitch, J. Marsh, and R. Dietert

Cornell University 21

Endocrine-Active Compounds (EACs) and Development of Sexually Dimorphic Forebrain Circuits

E.K. Polston CIIT Centers for Health Research 23

Topical Sensitization and Intranasal Challenge to Trimellitic Anhydride Induces an Allergic Rhinitis Similar to that Induced by Intranasal Sensitization and Challenge in A/J Mice

A. Farraj, J.R. Harkema, and N.E. Kaminski

Michigan State University 28

Improved Methods: Integration and Implications of New and Emerging Approaches in Health Effects Research Methods and Applications of Functional Genomics to Health Effects Research

R.S. Thomas CIIT Centers for Health Research 25

Improved Methods: Methods to Detect Effects on Wildlife A Diagnostic Approach for Identifying Biological Impairment and Dominant Stressors

G.A. Burton, W. Clements, and J. Oris

Wright State University, Colorado State University, and Miami University

2


K. Kidd, V. Palace, B. Evans, K. Mills, P. Blanchfield, C. Podemski, M. Paterson, A. Salki, and D. Findlay

Fisheries and Oceans Canada, Freshwater Institute

19/20

Investigation of Chemical Mixtures in the Upper Ocklawaha River Basin: Reproduction, Development and Endocrine Status in Alligators, Fish, and Mussels

C.J. Borgert and T.S. Gross Applied Pharmacology and Toxicology, Inc., U.S. Geological Survey, and University of Florida

22

Field-Deployable Methods for Evaluating Exposure and Effects of Endocrine-Active Substances in Wild Bird Populations

L. Brewer, S. Tank, N. Murray, and K. Hunt

Springborn Smithers Laboratories, and University of Washington

24

POSTER SESSIONS



No. A Multi-endpoint Strategy for Determining Mechanism of Action of Thyroid Active Chemicals

S.J. Degitz and J.E. Tiegte U.S. Environmental Protection Agency

36

Development of a Reptile Egg Screening Assay for Endocrine-Disrupting Chemicals

L.G. Talent and D.M. Janz Oklahoma State University and the University of Saskatchewan

37

Improved Methods: Interpreting and Using Biomonitoring Data Biomarkers of Exposure to Hexamethylene Diisocyanate

L.A. Nylander-French, K. Jayaraj, D. Klapper, L.M. Ball, and A. Gold

The University of North Carolina 6

Statistical Methods for Evaluating Exposure-Biomarker Relationships

S.M. Rappaport, L.L. Kupper, D.J. Taylor, B.A. Johnson, and Y. Lin

The University of North Carolina 9

Fetal Origins of Adult Disease: Do In Utero Exposures to Environmental Toxicants Play a Role?

W.G. Foster McMaster University 12

Improved Methods: Prediction of Target Tissue Dose Mechanisms of Adaptive and Adverse Responses in the Respiratory Tract Following Low-Level Exposure to Inhaled Reactive Gases

J.S. Kimbell, D.C. Dorman, P.M. Schlosser, A.M. Jarabek, and M.E. Andersen

CIIT Centers for Health Research 7

Dosimetry of Inhaled Particles in Humans B. Asgharian, J. Kimbell, B. Wong, and O. Moss


Modeling Inter-Individual Variation in Physiological Factors Used in PBPK Models of Humans

P.S. Price, R.B. Conolly, C.F. Chaisson, J.S. Young, D.R. Tedder, and E.T. Mathis

The LifeLine Group Inc., CIIT Centers for Health Research, and Hebrew University

17

Physiologically Based Pharmacokinetic (PBPK) Modeling of Genistein in Rats

P.M. Schlosser, S.J. Borghoff, N.G. Coldham, H.T. Tran, and M.G. Zager

CIIT Centers for Health Research, Veterinary Laboratories Agency, and North Carolina State University

26

Dose-Response Modeling with Monoalkylphthalates: Assessing Cellular Targets for Mechanisms-Based Risk Assessment

M.E. Andersen and R.A. Clewell CIIT Centers for Health Research 33

Potential Activation of Human Estrogen Receptors in HepG2 Cells by Genistein, Genistein-Glucuronide, and Genistein-Sulfate

S.J. Borghoff, H.D. Parkinson, S.M. Ross, K. Gaido, and M. Sochaski


Improved Methods: Understanding Toxicodynamics A Computer Simulation Model of Low-Dose-Response Relations for VOC Risks

L.A. Cox Cox Associates and the University of Colorado

13

Interaction Analysis of Synthetic Chemicals and Phytoestrogens In Vitro

E.W. Carney, G.D. Charles, C. Gennings, B.B. Gollapudi, and T.R. Zacharewski

Dow Chemical Company, Virginia Commonwealth University, and Michigan State University

14

Interaction Analysis of Synthetic Chemicals and Phytoestrogens In Vivo

S.D. Seidel, D.R. Boverhof, E.W. Carney, G.D. Charles, C. Gennings, B.B. Gollapudi, and T.R. Zacharewski


15

Comparison of Empirical and Mechanistic Models to Identify Relevant Toxicodynamic and Toxicokinetic Interactions Between Chemicals in a Mixture

A.P. Freidig, W.H.M. Heijne, R.H. Stierum, H.M. Wortelboer, M.W. Schut, H.A. El-Masri, D. Moffett, and J.P. Groten

TNO Nutrition and Food Research and ATSDR

30

POSTER SESSIONS



No. Non-Linear Dose-Response Relationships for Developmental Responses: An Example with Defeminization by Estrogenic Xenobiotics

W.C.J. Chung, M.E. Andersen, and R.J. Handa

Colorado State University and CIIT Centers for Health Research

34

Improved Methods: Health Assessment Methodologies Dose-Response Analysis: Bridging from the Laboratory to Science-Based Risk Assessment

R.B. Conolly CIIT Centers for Health Research 29

Susceptibility Factors: Human Health Sensitivity Factors Correlation of Nasal Surface-Area-to-Volume Ratio with Predicted Inhaled Gas Uptake Efficiency in Humans

R.A. Segal, G.M. Kepler, D.L. Kalisak, R.B. Richardson, and J.S. Kimbell

CIIT Centers for Health Research and the University of North Carolina

8

Investigating How Age and Gender Affect Internal Doses of Chemicals

H. Clewell ENVIRON Health Sciences Institute 16

An Analysis of the Need for an Additional Toxicokinetic Uncertainty Factor for Neonates

N.V. Corea and A.G. Renwick University of Southampton 32

Chemicals in the Environment: Human Exposure Assessment and Analysis Analysis of North Atlantic Aircraft Data on Oxygenated Intermediate Species Using an Adapted Regional Chemistry-Transport Model

R. Chatfield, R. Mathur, K. Alapaty, A. Hanna, F. Binkowski, H. Guan, and R. Esswein

NASA Ames Research Center, the University of North Carolina at Chapel Hill, and Bay Area Environmental Research Institute

1

Innovative Experimental Techniques to Help Understand Exposure to Volatile Organic Air Toxics

H.E. Jeffries, K.G. Sexton, I. Jaspers

University of North Carolina 5

Chemicals in the Environment: Ecosystem Exposure Analysis On the Use of TRI and NPRI Data in Air Dispersion Modeling for Ecological Exposure Assessment

R.E. Clinkenbeard, D.L. Johnson, N.A. Esmen, T.A. Hall, and D.V. Shinde

University of Oklahoma 3

Integration of Habitat Models Database with a Spatially Explicit Wildlife Foraging Model

L.A. Kapustka, I. Linkov, M. McVey, A.J. Grebenkov, A.G. Trifonov, H. Galbraith, M. Luxon, and J. Yocum

ecological planning and toxicology, inc., ICF Consulting, Galbraith Environmental Sciences, and Windward Environmental, LLC

4

POSTER SESSIONS


LRI Research Posters Sorted by Poster Number Poster No.

1 Analysis of North Atlantic Aircraft Data on Oxygenated Intermediate Species Using an Adapted Regional Chemistry-Transport Model

R. Chatfield, R. Mathur, K. Alapaty, A. Hanna, F. Binkowski, H. Guan, and R. Esswein

NASA Ames Research Center, the University of North Carolina at Chapel Hill, and Bay Area Environmental Research Institute

2 A Diagnostic Approach for Identifying Biological Impairment and Dominant Stressors

G.A. Burton, W. Clements, and J. Oris

Wright State University, Colorado State University, and Miami University

3 On the Use of TRI and NPRI Data in Air Dispersion Modeling for Ecological Exposure Assessment

R.E. Clinkenbeard, D.L. Johnson, N.A. Esmen, T.A. Hall, and D.V. Shinde

University of Oklahoma

4 Integration of Habitat Models Database with a Spatially Explicit Wildlife Foraging Model

L.A. Kapustka, I. Linkov, M. McVey, A.J. Grebenkov, A.G. Trifonov, H. Galbraith, M. Luxon, and J. Yocum

ecological planning and toxicology, inc., ICF Consulting, Galbraith Environmental Sciences, and Windward Environmental, LLC

5 Innovative Experimental Techniques to Help Understand Exposure to Volatile Organic Air Toxics

H.E. Jeffries, K.G. Sexton, I. Jaspers

University of North Carolina

6 Biomarkers of Exposure to Hexamethylene Diisocyanate

L.A. Nylander-French, K. Jayaraj, D. Klapper, L.M. Ball, and A. Gold

The University of North Carolina

7 Mechanisms of Adaptive and Adverse Responses in the Respiratory Tract Following Low-Level Exposure to Inhaled Reactive Gases

J.S. Kimbell, D.C. Dorman, P.M. Schlosser, A.M. Jarabek, and M.E. Andersen

CIIT Centers for Health Research

8 Correlation of Nasal Surface-Area-to-Volume Ratio with Predicted Inhaled Gas Uptake Efficiency in Humans

R.A. Segal, G.M. Kepler, D.L. Kalisak, R.B. Richardson, and J.S. Kimbell

CIIT Centers for Health Research and the University of North Carolina

9 Statistical Methods for Evaluating Exposure-biomarker Relationships

S.M. Rappaport, L.L. Kupper, D.J. Taylor, B.A. Johnson, and Y. Lin

The University of North Carolina

10 Statistical Issues in Modeling Pregnancy Outcome Data

G.M. Buck, V.M. Dukic, P.J. Heagerty, T.A. Louis, C.D. Lynch, L.M. Ryan, E.F. Schisterman, A. Trumble, and the Pregnancy Modeling Working Group

National Institute of Child Health and Human Development, University of Chicago, University of Washington, Johns Hopkins University, and Harvard University

11 Dosimetry of Inhaled Particles in Humans B. Asgharian, J. Kimbell, B. Wong, and O. Moss


12 Fetal Origins of Adult Disease: Do In Utero Exposures to Environmental Toxicants Play a Role?

W.G. Foster McMaster University

13 A Computer Simulation Model of Low-Dose-Response Relations for VOC Risks

L.A. Cox Cox Associates and the University of Colorado

14 Interaction Analysis of Synthetic Chemicals and Phytoestrogens In Vitro

E.W. Carney, G.D. Charles, C. Gennings, B.B. Gollapudi, and T.R. Zacharewski


15 Interaction Analysis of Synthetic Chemicals and Phytoestrogens In Vivo

S.D. Seidel, D.R. Boverhof, E.W. Carney, G.D. Charles, C. Gennings, B.B. Gollapudi, and T.R. Zacharewski


16 Investigating How Age and Gender Affect Internal Doses of Chemicals

H. Clewell ENVIRON Health Sciences Institute

POSTER SESSIONS



17 Modeling Inter-Individual Variation in Physiological Factors Used in PBPK Models of Humans

P.S. Price, R.B. Conolly, C.F. Chaisson, J.S. Young, D.R. Tedder, and E.T. Mathis

The LifeLine Group Inc., CIIT Centers for Health Research, and Hebrew University

18 Developmental Neurotoxicity Testing with Methyl Mercury and Methylazoxy Methanol: Neuropathology Endpoints Reconsidered

D.M.G. de Groot, M.H.M. Bos-Kuijpers, H-J.G. Gundersen, W.S.H. Kaufmann, J.H.C.M. Lammers, B. Pakkenberg, J.P. O’Callaghan, and D.H. Waalkens-Berendsen

TNO Nutrition and Food Research, the University of Aarhus, BASF, Research Laboratory for Stereology & Neuroscience, Copenhagen, DK, NIOSH

19/20 Effects of a Synthetic Estrogen on Aquatic Populations: A Whole Ecosystem Study

K. Kidd, V. Palace, B. Evans, K. Mills, P. Blanchfield, C. Podemski, M. Paterson, A. Salki, and D. Findlay

Fisheries and Oceans Canada, Freshwater Institute

21 Comparison of Fetal vs. Adult-induced Immunotoxicity with Cyclosporin-A

I. Hussain, M. Piepenbrink, K. Fitch, J. Marsh, and R. Dietert

Cornell University

22 Investigation of Chemical Mixtures in the Upper Ocklawaha River Basin: Reproduction, Development and Endocrine Status in Alligators, Fish, and Mussels

C.J. Borgert and T.S. Gross Applied Pharmacology and Toxicology, Inc., U.S. Geological Survey, and University of Florida

23 Endocrine-Active Compounds (EACs) and Development of Sexually Dimorphic Forebrain Circuits

E.K. Polston CIIT Centers for Health Research

24 Field-Deployable Methods for Evaluating Exposure and Effects of Endocrine-Active Substances in Wild Bird Populations

L. Brewer, S. Tank, N. Murray, and K. Hunt

Springborn Smithers Laboratories, and University of Washington

25 Methods and Applications of Functional Genomics to Health Effects Research

R.S. Thomas CIIT Centers for Health Research

26 Physiologically Based Pharmacokinetic (PBPK) Modeling of Genistein in Rats

P.M. Schlosser, S.J. Borghoff, N.G. Coldham, H.T. Tran, and M.G. Zager

CIIT Centers for Health Research, Veterinary Laboratories Agency, and North Carolina State University

27 The 2004 Long-Range Research Initiative (LRI) Portfolio at the CIIT Centers for Health Research: An Overview

F.J. Miller CIIT Centers for Health Research

28 Topical Sensitization and Intranasal Challenge to Trimellitic Anhydride Induces an Allergic Rhinitis Similar to that Induced by Intranasal Sensitization and Challenge in A/J Mice

A. Farraj, J.R. Harkema, and N.E. Kaminski

Michigan State University

29 Dose-Response Analysis: Bridging from the Laboratory to Science-Based Risk Assessment

R.B. Conolly CIIT Centers for Health Research

30 Comparison of Empirical and Mechanistic Models to Identify Relevant Toxicodynamic and Toxicokinetic Interactions Between Chemicals in a Mixture

A.P. Freidig, W.H.M. Heijne, R.H. Stierum, H.M. Wortelboer, M.W. Schut, H.A. El-Masri, D. Moffett, and J.P. Groten

TNO Nutrition and Food Research and ATSDR

32 An Analysis of the Need for an Additional Toxicokinetic Uncertainty Factor for Neonates

N.V. Corea and A.G. Renwick University of Southampton

POSTER SESSIONS



33 Dose-Response Modeling with Monoalkylphthalates: Assessing Cellular Targets for Mechanisms-Based Risk Assessment

M.E. Andersen and R.A. Clewell CIIT Centers for Health Research

34 Non-Linear Dose-Response Relationships for Developmental Responses: An Example with Defeminization by Estrogenic Xenobiotics

W.C.J. Chung, M.E. Andersen, and R.J. Handa

Colorado State University and CIIT Centers for Health Research

35 Potential Activation of Human Estrogen Receptors in HepG2 Cells by Genistein, Genistein-glucuronide, and Genistein-sulfate

S.J. Borghoff, H.D. Parkinson, S.M. Ross, K. Gaido, and M. Sochaski


36 A Multi-endpoint Strategy for Determining Mechanism of Action of Thyroid Active Chemicals

S.J. Degitz and J.E. Tiegte U.S. Environmental Protection Agency

37 Development of a Reptile Egg Screening Assay for Endocrine-Disrupting Chemicals

L.G. Talent and D.M. Janz Oklahoma State University and the University of Saskatchewan


Poster 1 Analysis of North Atlantic Aircraft Data on Oxygenated Intermediate Species Using an Adapted Regional Chemistry-Transport Model R. Chatfield1, R. Mathur2, K. Alapaty2, A. Hanna2, F. Binkowski2, H. Guan1,3, and R. Esswein1,3 1NASA Ames Research Center 2University of North Carolina at Chapel Hill 3Bay Area Environmental Research Institute Our study is on the interaction of nitrogen oxides with organics as they are exported from their complex sources in Eastern North America. Both urban and specific industrial emissions contribute the nitrogen of the C-H-O-N compounds that affect the global atmosphere, helping determine both ozone and the self-cleaning radical chemistry of the troposphere mediated by the OH radical. Different industrial sources, urban, and natural emissions contribute the organic C. Peroxyacetyl nitrate, CH3C(=O)OONO2 is the most interesting compound for which we can measure the outflow to the full depth of the Atlantic troposphere. As we adapt the 3-d chemical model to describe outflow for specific periods with sufficient accuracy, we are analyzing some valuable information in the NARE-97 complete airborne dataset (NARE: North Atlantic Regional Experiment). Ames researchers find that there are substantial puzzles in the ratios of PAN/NO2. Peroxy acetyl nitrate provides one of the major long-distance export pathways for active nitrogen from Eastern North America. It should be closely linked with NOx (defined as the sum NO + NO2) by simple thermal association and decomposition reactions, at least when the ambient temperature is substantially above 5° C. Over the course of many hours, the ratio, [PAN]/[CH3C(=O)OO][NO2] = kdissoc/kassoc, should be maintained. The ratios observed below ~4 km (>5° C) in NARE are significantly variable. This variability may imply signficant variation in the concentration of the important smog radical CH3C(=O)OO, peroxy acetyl, and this may provide important information about variations in the radical activity that go beyond current theory. We report progress in understanding these variations.

Observations of such unexplained great radical activity in the upper troposphere (SONEX, the Subsonic (A)ssessment Ozone and NOx EXperiment. also a 1997 field mission with complimentary information) have never been completely explained. Implications for the reactive radical chemistry of the free troposphere will be outlined. Figure 1. Values of log10 ([PAN]/[NO2]) plotted along airplane flight path during NARE-97. Significant mid-altitude variations suggest CH3C(=O)OO radical variability and are discussed in text.


Poster 2 A Diagnostic Approach for Identifying Biological Impairment and Dominant Stressors G. A. Burton1, W. Clements2, and J. Oris3 1Wright State University 2Colorado State University 3Miami University The primary objective of this research is to demonstrate a novel, systematic, and accurate approach for identifying the dominant stressors at stream and river sites. Sub-objectives include showing: (1) how this approach adapts to widely differing ecoregions and freshwater systems; (2) how significant biological impairment can be identified with a high degree of certainty; and (3) how the multiple lines of evidence utilized in the approach identify differing stressor classes where impairment is identified in a tiered assessment approach. This project describes a novel, weight-of-evidence approach that is superior to traditional assessment methods because the exposures to stressors are more realistic and less subject to sampling and testing artifacts. The assessment methods employed range from sensitive molecular biomarkers to community level endpoints. In addition, the identification of dominant stressors is possible through a proven, effective, tiered approach that separates out stressor sources (e.g., surface water, suspended sediments, pore water, surficial sediment, deep sediment, groundwater upwelling, low flow, high flow) and stressor types (e.g., dissolved oxygen, flow and suspended solids, substrate type, persistent and non-persistent organic pollutants, ammonia, photo-toxicants, endocrine disruptors, and metals). Biological impairment is verified via assessments of benthic macroinvertebrate communities where impairment is based on realistic assessments of optimal watershed conditions considering hydrological characteristics and land uses.


Poster 3 On the Use of TRI and NPRI Data in Air Dispersion Modeling for Ecological Exposure Assessment R. E. Clinkenbeard, D. L. Johnson, N. A. Esmen, T. A. Hall, and D. V. Shinde Department of Occupational and Environmental Health, University of Oklahoma Health Sciences Center, Oklahoma City, OK Dispersion modeling of industrial source air emissions serves as a primary basis for ecological risk estimation and regulatory decision-making concerning source permits and emission control requirements. Numerous air quality models have been developed over the past 25 years to estimate pollutant concentrations expected at a specific location downwind of an emission source. It is critical that model users be aware of the constraints and limitations of the modeling process, and of the appropriate and inappropriate uses of modeling results. This is particularly true regarding current EPA efforts to utilize Toxics Release Inventory (TRI) data to characterize trends in pollution prevention via such tools as the TRI Relative Risk-Based Environmental Indicators Methodology. The methodology employs a dispersion model to calculate indicators, based on TRI data, intended to track changes in human health and environmental impacts. The purpose of this work was to compare TRI-derived annual receptor exposure estimates with annual averages calculated from observed concentrations measured hourly at the target locations. A coal-fired power plant and a refinery operating in flat rural terrain and emitting SO2 were selected and the AERMOD and ISCST3 models were used to predict receptor exposures at locations corresponding to nearby State and Local Air Monitoring Stations (SLAMS). The SLAMS were approximately 2-3 km from the facilities and provided hourly SO2 measurements that were compared with the hourly predicted exposures. There was good correlation between the model predictions (Pearson product moment correlation = 0.86), with no apparent seasonal effect. There was essentially no correlation between the predicted and observed hourly results. Annual average receptor exposures calculated from the predicted and measured hourly values were within plus or minus a factor of three of one another; however, for one facility the predicted value was one-third of that observed, while for the other facility the predicted value was three times that observed. Although close to the "factor of two" accuracy generally considered good for dispersion modeling, these differences could be in either direction from the true value. The differences did not appear to be sensitive to errors in the TRI data; rather, they appeared likely to be due to the temporal de-coupling of emissions and meteorological data and the model assumption of constant emission rate. Therefore, application of current air dispersion models for determining ecological exposures and projecting human health risks using annual average emission rates, especially as derived from TRI- or NPRI data, should be tempered with an understanding of how the models work and their limitations in predicting quantitative exposure values.


Poster 4 Integration of Habitat Models Database with a Spatially Explicit Wildlife Foraging Model L. A. Kapustka1, I. Linkov2, M. McVey2, A. J. Grebenkov2, A. G. Trifonov2, H. Galbraith3, M. Luxon4, and J. Yocum1

1ecological planning and toxicology, inc. 2ICF Consulting 3Galbraith Environmental Sciences 4Windward Environmental, LLC Habitat Suitability Index (HSI) values, derived from comparatively routine measures of landscape features, describe the quality of habitat for particular wildlife species. The values represent estimates of potential carrying capacity of a specific area and provide indications of relative use patterns by the species. Previously, we encoded the information to enable calculation of the HSIs for approximately 90 wildlife species. These values are assigned to polygons or for an entire site. The values can then be used to modify the exposure estimates of Ecological Risk Assessments. In this project, we are developing and adapting software to link the calculations of HSI values to a series of modules (known collectively as Risk Trace) that calculate spatially explicit probabilistic estimates of foraging activities and hence exposure estimates. The modules use Microsoft applications and functions (e.g., Access, Excel, Visual Basic) for calculations, data processing, tracking user decisions, and reporting. Libraries of species-specific information on life histories, foraging range, food preferences, ingestion rates, foraging behaviors, etc. are incorporated into the calculation of exposure. Output will include Geographic Information System (GIS) and tabular displays that will provide probabilistic summaries of receptors, calculates exposures based on ingestion rates, and generates Hazard Quotients (HQs) for each contaminant. More sophisticated risk estimates (i.e., concentration-response relationships not just HQs) will be built into the software package where possible.


Poster 5 Innovative Experimental Techniques to Help Understand Exposure to Volatile Organic Air Toxics H. E. Jeffries1, K. G. Sexton1, I. Jaspers2 1School of Public Health, University of North Carolina, Chapel Hill, NC 2School of Medicine, University of North Carolina, Chapel Hill, NC The overall goal of this project is to develop and demonstrate new experimental techniques and methodologies drawn from human health effects and atmospheric chemistry practices that can be used to advance and prioritize the study of atmospheric chemical reactions of realistic mixtures including volatile organic air toxics and their transformation products, a significant subset of hazardous air pollutants (HAPs), while simultaneously, testing for their true toxicological effects on cultured human lung cells. By creating and manipulating inorganic and organic gaseous mixtures in environmental (“smog”) chambers that closely resemble those of urban-like photochemically reacting polluted air and by conducting simultaneous in vitro lung cell exposures to a suite of such conditions, followed by determining pro-inflammatory mediator expression and cytotoxicity, and applying advanced gas-phase organic species analytical methodologies, we can generate valuable mechanistic information on the chemical fate and true toxic potential of air pollutants needed for risk assessment calculations and simulations. We have shown the advantage of combining two experimental methods, to investigate what gaseous mixtures, components, and reaction products within these mixtures, are most potent in causing toxic effects. We have shown in this project, that photochemical products of primary pollutant mixtures, representing atmospheric transformations, can be more toxic than the precursor pollutants. In addition, small differences in molecular structure, between similar compounds such as 1,3-butadiene and 2-methyl-1,3-butadiene (isoprene), can result in surprising differences in relative cytotoxicity and inflammation, in both the original compounds and their photochemical products.


Poster 6 Biomarkers of Exposure to Hexamethylene Diisocyanate

L. A. Nylander-French, K. Jayaraj, D. Klapper, L. M. Ball, and A. Gold University of North Carolina, Chapel Hill, NC Diisocyanates are potent skin sensitizers and a leading cause of occupationally induced asthma. To determine the relative importance of dermal versus inhalation exposure in developing a systemic response to 1,6-hexamethylene diisocyanate (HDI) we will analyze samples of tape-stripped epidermis and sera from a population of exposed spray painters for tissue specific HDI-protein adducts using an enzyme-linked immunosorbent assay (ELISA). To develop the ELISA, 6-[(N-t-BOC)aminohexamethylene]isocyanate and N α-(FMOC)cysteine were conjugated to yield the protected hapten S-(6-[t-BOC-amino]hexyl)aminocarbonylcysteine-N α-FMOC. This putative monofunctional HDI adduct is suitably protected for conjugation with carrier protein or automated peptide synthesis. Conjugation with bovine serum albumin (BSA) has been accomplished and polyclonal antibodies (PCA) are currently in production. Hexamethylene-1,6-bisthiocarbamyl-(2-[FMOC-amino])propanoic acid has also been synthesized for analogous conjugation with BSA and production of PCA to detect bifunctional HDI adducts. These compounds were structurally characterized by mass spectrometry and 1H NMR spectrometry. Elemental compositions were determined by accurate mass measurement and purity established by 1H NMR spectrometry. The identification of adducts is critical and will provide biomarkers of diisocyanate exposure. Antibodies specific for dermal and for respiratory epithelium protein adducts, will allow determination of the relative importance of dermal versus inhalation exposure to HDI. The results obtained should yield insights into the correlation of dermal exposure both with dermal dose and with potential dermal and systemic effects.


Poster 7 Mechanisms of Adaptive and Adverse Responses in the Respiratory Tract Following Low-Level Exposure to Inhaled Reactive Gases J. S. Kimbell, D. C. Dorman, P. M. Schlosser, A. M. Jarabek, and M. E. Andersen CIIT Centers for Health Research, Research Triangle Park, NC There is a growing emphasis on subchronic, low-level, animal exposure-response studies for estimating human health risks from inhaled reactive gases. Interpreting less overt endpoints resulting from these studies, such as changes in the subcellular and genome domains (e.g., cytokine mediators, cell signaling and genetic profiling), in the context of adverse health effects already established as outcome measures is therefore becoming increasingly important. An understanding of the mechanisms of adaptive and adverse respiratory responses long the pathogenesis continuum that result from exposures of different durations to inhaled reactive gases is needed to guide risk assessors and managers in making use of these diverse data for human health risk estimates. To better understand mechanisms underlying these and other effects, an integrated program focused on chlorine and hydrogen sulfide research with a core dosimetry modeling component has been developed to characterize effects as adverse and adaptive. Experiments and computer simulations for these gases are coordinated to facilitate results interpretation, resource leveraging, and project synergy. The program will compare and contrast focused studies on the acute and low-level effects of chlorine and hydrogen sulfide and interpret the results in the context of previous experience with other reactive gases such as formaldehyde and ozone. This approach will provide information and context for the integration of these effects in human health risk assessment.


Poster 8 Correlation of Nasal Surface-Area-to-Volume Ratio with Predicted Inhaled Gas Uptake Efficiency in Humans R. A. Segal1, G. M. Kepler2, D. L. Kalisak1, R. B. Richardson1, and J. S. Kimbell1 1CIIT Centers for Health Research, Research Triangle Park, NC 2University of North Carolina at Chapel Hill, Chapel Hill, NC Variances in nasal anatomy, airflow, and patterns of inhaled gas uptake among human individuals may cause significant differences in the regional dose of inhaled gases or vapors in the nasal passages and subsequently to the lung. Computational fluid dynamics (CFD) models can quantify the contribution of anatomical variation to risk assessment uncertainty and reduce the reliance on default uncertainty values. For water-soluble, reactive gases, nasal surface area and volume may be predictors of nasal scrubbing efficiency. To test this hypothesis, nasal surface area and volume from the nostrils to the posterior end of the nasal septum were calculated from digitized MRI scans of the head for a number of adult individuals provided to us by Dr. Ray Guilmette and the Lovelace Respiratory Research Institute. Four subjects were selected whose ratio of nasal surface area to volume (SAVR) represented a range of values: 1.07, 1.03, 0.94, and 0.85 mm-1 for Subjects A (male), 12 (female), 14 (male) and 18 (female), respectively. Anatomically-realistic CFD models for the nasal passages of these individuals were constructed by a semi-automated process that provided input to a commercial mesh generator (Gambit, Fluent, Inc., Lebanon, NH). Allometrically-scaled minute volume flow rates were calculated to be 16.1, 12.9, 16.2, and 13.9 L/min for Subjects A, 12, 14, and 18, respectively. Steady-state inspiratory airflow at individualized flow rates and inhaled gas uptake were simulated for a reactive, water-soluble, low-molecular-weight gas using commercial CFD software (FIDAP, Fluent, Inc., Lebanon, NH). Total nasal uptake was estimated to be 90, 89, 85, and 84% for Subjects A, 12, 14, and 18, respectively, suggesting a correlation with SAVR. These studies and their extension to additional individuals form a basis for developing an empirical means of characterizing populations that may be at a higher risk for developing respiratory tract effects than the general population.


Poster 9 Statistical Methods for Evaluating Exposure-Biomarker Relationships S. M. Rappaport1, L. L. Kupper2, D. J. Taylor2, B. A. Johnson2, and Y. Lin1 1Department of Environmental Sciences & Engineering, School of Public Health, University of North Carolina, Chapel Hill, NC 2Department of Biostatistics, School of Public Health, University of North Carolina, Chapel Hill, NC It has long been anticipated that studies of biomarkers in persons exposed to toxic chemicals would reduce uncertainties inherent in extrapolation from animal assays to predict human risks of disease. Yet, applications of human biomarker data for risk assessment have been extremely limited due to the following inherent problems: 1) True subject-specific exposure and biomarker levels are unobservable and only error-prone surrogate

measurements are available for each human subject; 2) The numbers of available exposure and biomarker measurements per subject are quite small (e.g.,

many human subjects in a typical data set have only one surrogate exposure measurement and/or only one surrogate biomarker measurement);

3) There is often considerable within-subject and between-subject variability in (and correlation between) exposure and biomarker measurements, as well as for numerous important toxicokinetic parameters;

4) Required models relating changes in biomarker levels to changes in exposure levels are often non-linear due to saturation of toxicokinetic processes involving uptake, bioactivation, and detoxification;

5) Background levels of biomarkers often arise, both from unsampled exposures to the toxicant of interest and from other unrelated sources; and,

6) Measured exposure and biomarker levels often fall below analytical detection limits. In this project, we couple toxicokinetic theory with innovative statistical methods to develop hybrid models to quantify exposure-biomarker relationships in human populations. The models will be particularly suitable for evaluating the range of toxicokinetic linearity and sources of intra-individual and inter-individual variability. The models rely upon detailed simulation studies to identify appropriate mathematical forms and employ an extensive database of human exposures and biomarkers for validation.


Poster 10

Statistical Issues in Modeling Pregnancy Outcome Data

G. M. Buck1, V. M. Dukic2, P. J. Heagerty3, T. A. Louis4, C. D. Lynch1, L. M. Ryan5, E. F. Schisterman1, A. Trumble1, and the Pregnancy Modeling Working Group 1National Institute of Child Health & Human Development, Department of Health and Human Services 2University of Chicago 3University of Washington 4Johns Hopkins Bloomberg School of Public Health 5Harvard University Risk of repeating an adverse pregnancy outcome is approximately two-fold, underscoring the need for statistical designs and analyses that accommodate the dependent (correlated) nature of pregnancy outcomes. Failure to do so may mask effects, result in inaccurate variance estimators or produce biased or inefficient exposure estimates. Past analytic approaches include ignoring history, treating it as a covariate or side-stepping it by analyzing only one pregnancy per women. To address how best to model a sequence of pregnancy outcomes, we utilized data from the U.S. Collaborative Perinatal Project (CPP) to identify determinants of infant birth weight and small-for-gestational age (SGA). The CPP Study enrolled approximately 48,197 pregnant women at one of 12 clinical centers in the United States between 1959-1964. For study purposes, we restricted our sample to 2,717 mothers with 2+ consecutively born infants with complete information on study covariates (i.e., clinical site, maternal age, race, pre-pregnancy weight, cigarette smoking, family income, infant sex). Modeling strategies included generalized estimating equations (GEE) and mixed models with a variety of correlation structures. The various approaches resulted in clinically important differences in estimated effect size for birth weight (slopes) and robustly estimated standard errors (SE) and odds ratios (OR) and 95% confidence intervals (CI) for SGA for known biologic determinants of fetal growth. Pregnancy Outcome

Determinants of Fetal Growth

Independence GEE

Prior History as Covariate plus

GEE

One Random Pregnancy per

Woman

Random Intercept

(Mixed Model)

Birth weight (g) Slope(SE) Slope(SE) Slope(SE) Slope(SE) Smoking 1+ppd -259(28) -199 (22) -220(34) -217 (25)

Black race -234(34) -180(26) -199(35) -224 (31)

Age >30 years -141(49) -118(39) -163(63) -146(55) SGA OR (95% CI) OR (95% CI) OR (95% CI) OR (95% CI)

Smoking 1+ppd 2.9(2.2,3.8) 2.6(2.0,3.4) 2.2(1.5,3.2) 2.9(2.1,4.0) Black race 0.8(0.6,1.2) 0.8(0.6,1.1) 0.8(0.5,1.2) 0.7(0.5,1.1) Age >30 years 1.1(0.6,2.2) 1.1(0.6,2.2) 1.1(0.5,2.8) 1.3(0.6,2.9)

Depending on the purpose of the analysis, we recommend the use of mixed models or GEE with robust SE estimates to minimize statistical inefficiency and to provide the most appropriate effect estimation.


Poster 11 Dosimetry of Inhaled Particles in Humans

B. Asgharian, J. Kimbell, B. Wong, and O. Moss CIIT Centers for Health Research, Research Triangle Park, NC Risk assessments for respiratory toxicants are prone to specific uncertainties regarding inter- and intraspecies variability, high-to-low-dose extrapolation, and susceptible subpopulations. Computational modeling techniques are particularly useful in reducing the uncertainties by identifying and studying various physical and biological parameters involved in the exposure-dose-response paradigm. The goal in this project is to develop methods to improve risk assessment from exposure to inhaled material. Predictions of particle deposition in the nasal molds of human, monkey and rat were measured. A clear pattern of deposition with species emerged. In addition, deposition of hygroscopic particle in adult human lungs was calculated. Deposition characteristic of hygroscopic particles were found to be drastically different from that of insoluble ones. In another study, airflow distribution among the five lobes of adult human and children lungs was computed and used to obtain predictions of particle deposition. Children had fairly uniform ventilation that changed with age with more air going to the basal lobes for adults.


Poster 12 Fetal Origins of Adult Disease: Do In Utero Exposures to Environmental Toxicants Play a Role? W. G. Foster Department of Obstetrics & Gynecology, McMaster University Increasingly fetal insult has been linked with neonatal and adult diseases such as obesity, type-II diabetes, and cardiovascular disease. Reports in the literature of an increase in the prevalence of developmental abnormalities of the male reproductive tract, hormone dependent cancers, and asthma have given rise to concerns that in utero exposure to environmental contaminants may be associated with adverse health outcomes in children and adults. However, numerous limitations of contemporary studies preclude establishment of a link between exposure to any environmental contaminant or group of contaminants and adverse health outcomes in the human population. A major limitation of studies to date is accurate assessment of in utero exposure to environmental contaminants. Furthermore, exposure studies have tended to measure chemical contaminants without consideration of potential mechanism of action for the adverse outcome of concern. We hypothesize that exposure to estrogenic agents in the environment are too low to induce adverse health outcomes in the human population but exposure to aryl hydrocarbon receptor agonists could potentially change maternal thyroid function and contribute to developmental abnormalities in the fetus and neonate. To address the hypothesis 310 pregnant women attending a prenatal diagnosis clinic at McMaster University Medical Centre were invited to participate in this study from which 150 women recruited. The study was conducted in accordance with the McMaster University ethics committee approval and all participants completed signed consent, before completing intake, food frequency, and obstetrical history questionnaires. The mean age of women enrolling in this study was 38.0 ± 0.2 years (range 34-44 years). A single 20 ml blood sample was collected during the second trimester prenatal visit from each woman and allowed to clot at 4 C, serum collected, and aliquoted for subsequent analyses. Serum samples were subsequently analyzed for aryl hydrocarbon receptor activity using an established chemically activated luciferase expression assay (CALUX). Maternal thyroid function was assessed by commercially available radioimmunoassays for thyroid stimulating hormone (TSH) and thyroxine (T4) as well as the presence of thyroid peroxidase antibodies (TPO). Birth weight was recorded for each neonate as were clinical assessments of neonatal development. Preliminary CALUX results reveal mean (± SEM) serum levels of 0.34 ± 0.01 and range of 0.19 to 0.73 TEQ pg 2,3,7,8 tetrachlorodibenzo-p-dioxin (TCDD)/g serum lipid. Of this study cohort 6 of 150 women (4%) were clinically hypothyroid (TSH > 7.0 µIU/ml and T4 < 61.7 nmol/L) and 12/150 women (8%) were classified with sub-clinical hypothyroidism (TSH > 4.5 µIU/ml and T4 < 57.9 nmol/L). Five neonates were classified as low birth weight babies (less than 2500 g/birth). Multiple regression analysis of our preliminary results failed to demonstrate a statistically significant relationship between maternal serum TCDD TEQ level, thyroid hormone function, and birth weight after controlling for potential confounding factors. Therefore our preliminary results fail to support the hypothesis and we conclude that in utero exposure to AhR agonists fail to affect maternal thyroid function or induce adverse health effects in neonates. It should be noted however that these are preliminary results and the study is ongoing. Furthermore, infants continue to be followed and markers of immune function during the first postnatal year are being recorded to assess both immune system development.


Poster 13

A Computer Simulation Model of Low-Dose-Response Relations for VOC Risks

L. A. Cox Cox Associates and University of Colorado, Denver, CO, Pharmacodynamic models representing interactions among chemicals and cells can help to clarify how time patterns of administered dose affect risks of adverse health outcomes. This paper presents a model that predicts the effects of myelotoxic chemicals on the hematopoietic (blood-forming) system. It consists of a set of physiological compartments representing hematopoietic progenitor cell, granulocyte-macrophage (GM)-committed stem cells, and more mature blood cells. These compartments are linked by nonlinear feedback control loops and are susceptible to first-order cell-killing kinetics from cytotoxic metabolites. To clarify the shapes of low-dose dose-response relations for benzene and VOC mixtures, we developed an integrated pharmacokinetic-pharmacodynamic (PBPK/PD) model of effects of benzene and VOC mixtures on hematopoietic stem cell populations (e.g., CFU-GM) involved in chemically-induced s-AML. The model, including a partly experimentally validated feedback-control sub-model of suppression and compensating proliferation of hematopoietic stem cells in response to hematotoxins, identifies combinations of PBPK and PD factors by which the timing of VOC administration affects hematotoxicity and risk at physiologically relevant levels. A striking prediction is that low (linear PBPK) concentrations of benzene-containing VOC mixtures can reduce stem cell proliferation, premature recruitment of early stem cells, and resulting risk of s-MDS and s-AML. The prediction of a U-shaped dose-response relation is robust to model uncertainties, but occurs only below the concentrations used in most experiments. The model successfully explains some past puzzles in published data (e.g., how and why smaller total doses of inhaled benzene can have larger hematotoxic effects if administered as a relatively short, concentrated dose) and may help to interpret the epidemiology of low exposures to benzene and hematotoxic VOC mixtures. The hematotoxicity portion of the model has been validated by testing its predictions against experimental and clinical data for blood cell counts following administration of cyclophosphamide to mice, dogs, and humans. It successfully explains apparent anomalies and patterns in previously published data, including the fact that smaller cumulative doses can cause larger hematotoxic responses. An intriguing prediction from the model is that certain combinations of concentrations and spacing between consecutive exposures can greatly change (e.g., by 7-fold) the predicted carcinogenic risk from the same total AUC of administered dose or exposure.


Poster 14 Interaction Analysis of Synthetic Chemicals and Phytoestrogens In Vitro E. W. Carney1, G. D. Charles1, C. Gennings2, B. B. Gollapudi1 and T. R. Zacharewski3 1The Dow Chemical Company, Midland, MI 2Department of Biostatistics, Virginia Commonwealth University, Richmond, VA 3Department of Biochemistry and the National Food Safety & Toxicology Center Michigan State University, East Lansing, MI Humans are potentially exposed to many different estrogenic chemicals including synthetic chemicals (SC’s), typically at very low levels, and phytoestrogens (PE’s), often present in diets at high levels. In this study we assessed interactions between SC’s and PE’s using an estrogen receptor-α reporter gene assay. Individual concentration-response curves were first generated for each of six SC’s [methoxyclor, o,p-DDT, bisphenol A, β-hexachlorocyclohexane, 2,3-bis (4-hydroxyphenyl)-propionitrile), octylphenol] and for a PE preparation (genistein:daidzein, 2:1). A fixed-ratio mixture of all six SC’s was then devised with the proportion of each chemical in the mixture based on its no-effect concentration. The SC mixture was tested at total concentrations of 0.02, 0.2, 1.0, 2.0, or 3.0 µM, with 2.0 µM being the total dose at which each individual chemical in the mixture was at its own response threshold. We then asked whether or not the threshold and slope of the PE concentration-response curve (0, 0.01, 0.025, 0.05, 0.1, 0.15, 0.5 µM) were shifted when the PE’s were combined with various concentrations of the SC mixture. In general, the SC mixture significantly increased response thresholds relative to the PE’s alone, an effect most likely due to the lower potency of the SC’s and their competition with the more potent PE’s. Once these thresholds were exceeded, slopes of the PE+SC mixture responses were significantly steeper than that of the PE’s alone. Finally, analysis of the SC mixture without PE’s indicated an antagonistic interaction between the SC mixture components at the ratio tested. These results highlight the dose-dependency of chemical interactions and also exemplify a novel experimental and statistical approach for assessing complex chemical mixtures at low concentrations.


Poster 15 Interaction Analysis of Synthetic Chemicals and Phytoestrogens In Vivo S. D. Seidel1, D. R. Boverhof3, E. W. Carney1, G. D. Charles1, C. Gennings2, B. B. Gollapudi1 and T. R. Zacharewski3 1The Dow Chemical Company, Midland, MI 2Department of Biostatistics, Virginia Commonwealth Univ., Richmond, VA 3Department of Biochemistry and Molecular Biology, National Food Safety & Toxicology Center, Michigan State Univ., East Lansing, MI Humans are potentially exposed to many different estrogenic chemicals including synthetic chemicals (SCs), typically at very low levels, and phytoestrogens (PEs), often present in the diet at high levels. In this study we assessed interactions between SCs and PEs using the immature rat uterotrophic assay (UA) coupled with expression analysis of estrogen-responsive genes. All test compounds were administered by gavage once daily for 3 days. 24 h after the last dose, uteri were harvested, wet weights recorded and tissues were snap frozen for real-time RT-PCR analysis. Individual dose-response curves in the UA were first generated for each of six SCs [methoxyclor, o,p-DDT, octylphenol, bisphenol A, β-hexachloro-cyclohexane, 2,3-bis(4-hydroxyphenyl)-propionitrile)] and for a PE preparation (genistein:daidzein, 2:1) with ethynylestradiol (EE) (10 µg/kg/day) as a positive control. A fixed-ratio mixture of all six SCs was then devised with each chemical in the mixture present at its individual response threshold dose (i.e., a dose causing a 20% increase in uterine wet weight). We then compared the dose-response for the PE preparation alone (total doses of 0, 10, 30 or 120 mg/kg/day) with that of the PEs combined with the SC mixture. Statistical analysis of the UA data indicated that the PEs and SCs at the tested dose levels produced an additive response in the uterotrophic assay. Expression of two genes induced by EE, complement component 3 and intestinal calcium-binding protein, also appeared to increase in a dose-responsive manner following treatment with the PE’s alone and with the PE+SC mixtures. Studies to further evaluate the estrogenic effects of mixtures of PEs with lower levels of SCs are currently in progress.


Poster 16 Investigating How Age and Gender Affect Internal Doses of Chemicals

H. Clewell ENVIRON Health Sciences Institute, Ruston, LA The objective of this research was to identify age- and gender-specific differences in physiological and biochemical processes that affect tissue dosimetry and integrate them into a predictive pharmacokinetic (PK) framework. Three case studies were conducted based on information available in the literature. The first case study focused on physiological and biochemical determinants of systemic toxicity, and tested the hypothesis that many of the pharmacokinetic differences observed across age and gender may be attributed to fundamental changes in physiological and biochemical processes. The second case study focused on the impact of age- and gender-specific differences in lung morphology and ventilation rate on both local and systemic inhalation toxicity as a function of the properties of the chemical. The third case study focused on the perinatal period, using physiologically based PK models to evaluate in utero exposure via placental transfer versus infant exposure via lactational transfer to demonstrate critical periods of exposure from a pharmacokinetic perspective. These studies were exercised for multiple compounds to provide information on pharmacokinetic behaviors as a function of the physicochemical properties of a compound. In general, variations in pharmacokinetic dose metrics for a chemical over a lifetime were within a factor of two; however, exceptions were observed in the neonatal period where the dose metrics varied by as much as an order of magnitude from adult values. Perinatal simulations demonstrated the importance of in utero, as compared to neonatal, exposure, even in the case of chemicals with a high potential for lactation transfer.


Poster 17 Modeling Inter-Individual Variation in Physiological Factors Used in PBPK Models of Humans P. S. Price1, R. B. Conolly2, C. F. Chaisson1, J. S. Young3, D. R. Tedder1, and E. T. Mathis1 1The LifeLine Group Inc. 2CIIT Centers for Health Research 3Hebrew University Modeling interindividual variation in internal doses in humans using PBPK models requires data on the variation in the physiological parameters across the population of interest. These data should also capture the correlations between the values in each person. In this project, a software program Physiological Parameters for PBPK Modeling (P3M) is developed to provide such data. P3M provides a source of data for human physiological parameters where (1) the parameter values for an individual are correlated with one another, and (2) values of parameters vary according to interindividual variation in the general population, by gender, race, and age. The parameters investigated in this project include: (1) volumes of selected organs and tissues; (2) blood flows for the organs and tissues; and (3) the total cardiac output under resting conditions and average daily inhalation rates. These parameters are expressed as records of correlated values for the approximately 30,000 individuals evaluated in the NHANES III survey. The software program, P3M, allows data on selected parameters to be retrieved randomly from the database with specification of constraints on, age, sex, and ethnicity. The software provides a convenient tool for parameterization of human PBPK models for the study of interindividual variation. In addition, the data in P3M provide a useful source of information on the variation in physiological parameters in adults and children.


Poster 18 Developmental Neurotoxicity Testing with Methyl Mercury and Methylazoxy Methanol: Neuropathology Endpoints Reconsidered D. M. G. de Groot1, M. H. M. Bos-Kuijpers1, H. J. G. Gundersen2, W. S. H. Kaufmann3, J. H. C. M. Lammers1, B. Pakkenberg4, J. P. O’Callaghan5, and D. H. Waalkens-Berendsen1 1TNO Nutrition and Food Research, Zeist, NL 2University of Aarhus, DK 3BASF, Ludwigshafen, FRG 4Research Laboratory for Stereology & Neuroscience, Copenhagen, DK 5National Institute for Occupational Health and Safety, Morgantown, WV A Developmental Neurotoxicity Test (EPA, OPPTS 6300 and 8600) with extensive neuropathology and limited behavioural testing was carried out with methylmercury (MeHg) chloride (dosing period: GD6-PN10) and methylazoxy methanol (MAM) acetate (dosing period: GD13-15) in rats (5 dose levels each, including vehicle control). The main goal of the research was to study existing and new neuropathology endpoints for potential use in regulatory Developmental Neurotoxicity Testing (American Chemistry Council RfP 01-01; Ref.no.1847).

MeHg caused significant maternal (body weight) and developmental toxicity (number of pups delivered, number of stillborn pups, pup weight and hydronephrosis at necropsy) in the 1 mg MeHg group. MAM caused significant and dose-related maternal toxicity (mainly body weight) in the 2.5, 5 and 7.5 mg MAM groups and developmental toxicity (pup weight and motor activity) in the 7.5 mg MAM group. Effects of MAM on time patterns of motor activity during development were observed, suggesting a developmental neurotoxic effect of MAM. Various macroscopical brain size measures indicated significant effects of MAM (5 and 7.5 mg) but not of MeHg. Linear measures of morphometry showed a dose-related reduction in size of certain brain regions (hypoplasia), significant even in the 2.5 mg MAM dose group. Effects of MeHg were not observed.

Estimation of neuron numbers and neuron layer volume by stereological means is on-going to evaluate absolute neuron numbers as potential new neuropathology endpoint for use in regulatory Developmental Neurotoxicity Testing. Neurons in the hippocampal CA1 pyramidal layer (MAM) and in the cerebellar granular layer (MeHg) are investigated.


Poster 19 - 20


K. Kidd, V. Palace, B. Evans, K. Mills, P. Blanchfield, C. Podemski, M. Paterson, A. Salki, and D. Findlay Fisheries and Oceans Canada, Freshwater Institute, Winnipeg, MB, Canada The potent estrogen mimic used in birth control pills has been added continuously over three summers (2001-03) to a lake at the Experimental Lakes Area in northwestern Ontario. Effects on the microbial through fish populations have been assessed in this lake during additions, and contrasted to pre-addition and reference lake data. Male minnows exposed to the environmentally-relevant concentrations of 17α-ethynylestradiol (EE2; 5-6 ng/L) produced high concentrations of the egg protein precursor vitellogenin and have developed gonadal abnormalities (including intersex). Female fish are being impacted at the biochemical through tissue level with increased production of vitellogenin and delayed development of oocytes. Population monitoring indicates that the fathead minnow have not successfully spawned in the treated lake in the last two years resulting in a loss of the smaller size classes of this species; no changes in the abundances of the other three species (lake trout, white sucker, pearl dace) have yet been observed. Some impacts on lower-trophic-level organisms have also been found. Female zooplankton of some species produced fewer eggs in both 2001 and 2002, and total zooplankton abundances were higher in 2003 likely due to decreased predation by minnows. Intersex was also observed in mink frog tadpoles caged in the experimental lake but not in the reference systems. Low concentrations of a potent estrogen mimic can have adverse impacts on both vertebrates and invertebrates in aquatic ecosystems either through direct or indirect effects.


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Poster 21

Comparison of Fetal vs. Adult-induced Immunotoxicity with Cyclosporin-A

I. Hussain, M. Piepenbrink, K. Fitch, J. Marsh, and R. Dietert Department of Microbiology and Immunology, College of Veterinary Medicine, Cornell University, ] Ithaca, NY While Cyclosporin A (CYP-A) is a widely-used immunosuppressant drug, information on the long-term impact of fetal exposure is scarce. CYP-A effects on reproductive and immunologic parameters were examined in rat CD-strain female offspring exposed in utero at doses of 0, 0.2, 2, 10, or 20 mg/kg/day (from gestational day 6 to 21) and compared against identically-dosed adults. CYP-A was acutely immunotoxic (thymus morphology) in adults but with minimum long-term effects. In offspring (two lowest doses examined, postnatally), CYP-A increased ano-genital distance at one but not three weeks of age. At 5-weeks, the DTH response and splenic B cell number were both decreased (2 mg dose level). IL-4 level was reduced and blood monocytes were increased at both doses. All other parameters were unchanged (cell populations, cytokines). In the adult offspring (13 wk-old), no difference was seen in either the DTH response or B cell number, but IL-4 was increased at 2 mg/kg/day, and anti-KLH IgG titer decreased at both low doses examined. In exposed adults (four doses), changes were minimal following the 13-wk recovery period. Blood neutrophils were increased at all doses and modest perturbations in CD4+CD8+ cells, macrophages, and B-cells (decreased at 10 mg but increased at 20 mg dose) were seen. All other functional and cytokine parameters were unchanged. In conclusion, the adult rodent immune system largely recovered from CYP-A exposure given sufficient time. However, fetal exposure produced a series of immune perturbations during postnatal maturation including functional impairment.


Poster 22 Investigation of Chemical Mixtures in the Upper Ocklawaha River Basin: Reproduction, Development and Endocrine Status in Alligators, Fish, and Mussels C. J. Borgert1,3 and T. S. Gross2,3

1Applied Pharmacology and Toxicology, Inc., Gainesville, FL 2United States Geological Survey Biological Resources Division, Gainesville, FL 3Department of Physiological Sciences, College of Veterinary Medicine, University of Florida, Gainesville, FL The specific aims of this project are to (1) extend development and validation of assays for endocrine sensitive endpoints in alligators, fish, and freshwater mussels for use in interaction experiments and whole mixture studies; (2) determination of an appropriate combined action model for predicting the effects of binary and ternary mixtures of endocrine-active compounds; and (3) test the utility of binary interactions for predicting the toxicity of more complex mixtures, such as those found in the Ocklawaha River Basin in Florida where putative endocrine disruptive effects have been reported. Studies designed to validate a microinjection technique for delivery of OCPs into alligator eggs found a high incidence of neonate mortality. Attempts to validate topical treatment techniques for delivery of OCPs into alligator eggs revealed that little, if any, of the applied dose is absorbed into the egg yolk. These results, and the lack of published validation studies for the topical treatment technique in reptilian eggs, calls into question the validity of in ovo endocrine disruption studies in reptiles. Therefore, we have abandoned further efforts to use alligator eggs as a laboratory model for mixtures. Single doses of p,p’-DDE and dieldrin to the freshwater mussel Elliptio buckleyi (environmental exposure) and the Florida largemouth bass (Micropterus salmoides floridanus) (dietary exposure) produced effects on circulating sex-steroid hormone and vitellogenin levels (assayed in mussels only), but with mixed results. These studies are being used to determine appropriate doses for binary and ternary interaction studies and studies with defined whole mixtures scheduled for spring and summer, 2004.


Poster 23 Endocrine-Active Compounds (EACs) and Development of Sexually Dimorphic Forebrain Circuits E. K. Polston CIIT Centers for Health Research, Research Triangle Park, NC Perinatal estradiol and synthetic estrogens act on the developing mammalian brain to sexually differentiate forebrain circuits, allowing for the expression of sex-specific neuroendocrine and behavioral profiles during adulthood. There is a possibility that estrogen-like EACs in the environment may perturb sex-specific developmental processes in the brain. In animal models, perinatal exposure to high-doses of estrogenic EACs such as genistein (GEN) and bisphenol-A (BIS) result in reproductive deficits in adults, suggesting that the exposures of the developing brain can lead to persistent, permanent effects. However, the extent to which these and other EACs are capable of reaching hormone responsive brain areas during development and the potential for them to permanently disrupt sexually dimorphic brain circuits at lower doses remains largely unexplored, making it difficult to assess the neurological risks associated with early EAC exposure. To begin to address these questions, we are utilizing neuroanatomical methods to investigate the effects of a natural phytoestrogen, GEN, and a synthetic EAC, BIS, on the development of a limbic-hypothalamic circuit that regulates gonadotropin release in adults. While both compounds are able to reach the brain and potentially affect reproductive development, they have differential binding affinities for the estrogen receptor, making them useful in experiments that examine estrogen receptor-mediated events. Initial dose-response studies will utilize wide dose ranges of estrogen, GEN, and BIS to determine the extent to which estrogen receptors in the neonatal rat brain are bound by each compound and whether perinatal exposure to physiologically relevant doses can alter the development of the forebrain circuits that regulate reproduction. Ultimately, these methods, coupled with mechanistic studies of estrogen action in the brain, will further the goal of reducing risk assessment uncertainties regarding effects of these EACs on brain development and function.


Poster 24 Field-Deployable Methods for Evaluating Exposure and Effects of Endocrine-Active Substances in Wild Bird Populations L. Brewer1, S. Tank1, N. Murray1, and K. Hunt2

1 Springborn Smithers Laboratories, Wareham, MA 2 Center for Conservation Biology, University of Washington, Seattle, WA We tested the use of digital, time-lapse videography and radio-immunoassay (RIA) to compare the levels of excreted estrogen of wild birds exposed to exogenous estrogen verses non-exposed birds. Uniquely designed feeding station equipped with digital video equipment and feces collection devices were deployed for repeated collections of fecal-urate-urine (FUU) samples from local populations of songbirds. Mist netting was conducted at the feeding stations to capture and implant time-release estradiol tablets in a portion of the local birds. Implanted birds at some stations were color-marked on the head for identification on video and subsequent separation of FUU samples from treated verses non-treated birds. At some stations a proportion of the birds were implanted with estrogen, but were not otherwise marked. FUU samples were identified to species and exposure scenario with the video. We categorized FUU samples into three source categories: (1) estrogenized birds, (2) non-estrogenized birds, and (3) mixed sample from unmarked birds. FUU samples were analyzed for estrogen content and compared for mean differences using analysis of variance. Mean estrogen level in FUU from known estrogenized birds was significantly greater than that of non-estrogenized birds and the mixed-sample birds. Also, the mean level of excreted estrogen in the mixed-sample birds was statistically greater than that of the non-estrogenized birds. These results suggest that field exposure of birds to exogenous estrogen can be evaluated by simultaneous collection of FUU samples from an area of suspected contamination and the estrogen content compared to that of FUU samples collected from a non-contaminated area.


Poster 25 Methods and Applications of Functional Genomics to Health Effects Research R. S. Thomas Functional Genomics Research Program, CIIT Centers for Health Research, Research Triangle Park, NC “Functional genomics” is a new catchphrase that has a number of commonly used definitions. A broad definition of the term is that it is the branch of genomics that studies the genome in a cellular and functional context and is usually characterized by high-throughput, large-scale experimental methods. The overarching goals of the Functional Genomics Research Program at CIIT Centers for Health Research are to develop tools that delineate the cellular signaling pathways involved in toxicity and identify the cause-and-effect relationships among altered genes. To accomplish this objective, the research in this project will focus on three primary areas: Area 1: Develop functional genomic screens to dissect upstream cellular signaling pathways. Dissecting the upstream cellular signaling pathway is important for understanding the potential targets of toxicity, identifying key nodal points in the signaling pathway, and potential crosstalk among pathways known to be involved in other biological endpoints (e.g., development, inflammation). Area 2: Perform functional genomic screens to identify genes involved in susceptibility. Applying gain-of-function and loss-of-function screens in cells exposed to a marginally toxic or sub-toxic dose can identify potential modifiers of susceptibility. Area 3: Use a combination of gene expression analysis and functional genomic tools to map primary, secondary, and tertiary gene expression responses following toxicant exposure. In the cascade model of gene regulation, genes altered early in the process are the primary regulators, while secondary and tertiary changes in expression are more directly involved in the biochemical and structural response to the stimulus. Biologically, the sum of the connections that link the primary regulators with the secondary subnetworks determines the physiological response of an organism or cell to a particular stimulus. Identifying these connections will be an important part of investigating the toxicological mechanisms. Through the application of functional genomic tools, a novel approach to toxicology will be developed to provide critical mechanistic understanding of toxic responses that is essential for predicting the shape of the dose-response curves and supporting risk assessments.


Poster 26 Physiologically Based Pharmacokinetic (PBPK) Modeling of Genistein in Rats P. M. Schlosser1, S. J. Borghoff 1, N. G. Coldham2, H. T. Tran3, and M. G. Zager1,3 1CIIT Centers for Health Research, Research Triangle Park, NC, USA 2Department of Bacterial Diseases, Veterinary Laboratories Agency, Surrey, UK 3Center for Research in Scientific Computation, NCSU, Raleigh, NC, USA Genistein is a phytoestrogen naturally occurring in soy products. While a high-soy diet is generally considered beneficial for adults, high-level exposures during development have been shown to cause adverse effects in reproductive tissues of rats, underscoring concerns about exposure of human infants and fetuses. To help interpret these findings we are developing a PBPK model of genistein in rats to quantify the amount of genistein reaching target tissues. Such a model can later be extrapolated to humans. In particular the model should accurately reproduce data for the parent compound, genistein, and total radiolabel activity in blood plasma (including genistein and all metabolites), along with total radiolabel activity in a wide range of tissues. In order to accurately simulate all of these data, additional mechanistic information must be incorporated into the model description. Binding of genistein and its conjugates to rat plasma protein has been observed experimentally. Moreover, much higher concentrations of genistein found in gastrointestinal (GI) tissue after oral dosing compared to other richly perfused tissues suggest that the GI tissue was not completely separated from the GI contents. Changing the model structure to include plasma protein binding of genistein and its metabolites, and to describe contamination of the GI tissue by GI lumen contents, greatly improved the ability of the model to reproduce the experimental results. But these results also show the value of coordinating computational modeling with experimental studies, since experiments done in parallel would allow us to validate or refute these proposed changes in model structure.


Poster 27 The 2004 Long-Range Research Initiative (LRI) Portfolio at the CIIT Centers for Health Research: An Overview F. J. Miller CIIT Centers for Health Research, Research Triangle Park, NC The goal of CIIT’s LRI-funded research program is to develop and implement interdisciplinary research programs of the highest scientific merit that are aligned with and responsive to the human health issues of highest priority to the ACC and its members as well as to society at large. Some areas of concern to industry are defined very broadly, such as children’s health, while other areas and questions are much narrower in scope. Our objective for the LRI research effort is to have a mixed portfolio of program projects that address broad issues, individual projects that address important topics more narrow in scope, and projects that leverage the capabilities of CIIT centers of excellence. In developing the LRI research program, the CIIT Research Leadership Group uses various criteria to select the projects that collectively comprise an integrated, interdisciplinary program of high relevance to the ACC and its members. Broadly stated, the basic criteria that are used address the scientific value and relevancy of the project to the chemical industry, the value of the research in protecting public health, the potential for impact on a specific research field, the feasibility of the research, and whether sufficient resources (staff, supplies, equipment, and facilities) are available to conduct the research. This poster provides an overview of the various projects comprising the 2004 LRI-funded program at CIIT and how these projects link to the focus areas established in the LRI Research Strategy document.


Poster 28 Topical Sensitization and Intranasal Challenge to Trimellitic Anhydride Induces an Allergic Rhinitis Similar to that Induced by Intranasal Sensitization and Challenge in A/J Mice A. K. Farraj1,2, J. R. Harkema2, and N. E. Kaminski1

1Department of Pharmacology and Toxicology, Michigan State University, East Lansing, MI 2Pathobiology and Diagnostic Investigation and Center for Integrative Toxicology, Michigan State University, East Lansing, MI Allergic airway diseases caused by low molecular weight chemicals, including trimellitic anhydride (TMA), have been linked to Th2 cytokine production and are characterized by mucus hypersecretion and the infiltration of lymphocytes and eosinophils into the airways. The most common route of human exposure to chemical allergens is inhalation. Most murine models, however, use topical exposure to sensitize mice. The present study tests the hypothesis that topical sensitization on the ears of mice with TMA will induce a qualitatively similar immunologic and pathologic response in the nasal airways after intranasal challenge to that induced after intranasal sensitization and challenge. A/J mice were topically or intranasally sensitized and then intranasally challenged with TMA in an ethyl acetate/olive oil vehicle. Intranasal challenge with TMA in mice that were either topically or intranasally sensitized with TMA caused a marked allergic rhinitis characterized by an influx of eosinophils and lymphocytes of similar severity. Both the topical and intranasal routes of sensitization also caused significant increases in total serum IgE after intranasal challenge with TMA. In addition, both the topical and intranasal routes of sensitization caused significant increases in the mRNA expression of the Th2 cytokines IL-4, IL-5, and IL-13. The data suggest that topical application is effective in sensitizing mice to TMA and induces a nasal airway lesion and associated immune response after intranasal challenge that is qualitatively similar to that induced by intranasal sensitization and challenge. Skin exposure may be a potential route of sensitization of the human respiratory tract to chemical allergens.


Poster 29 Dose-Response Analysis: Bridging from the Laboratory to Science-Based Risk Assessment

R. B. Conolly Center for Computational Biology & Extrapolation Modeling, CIIT Centers for Health Research, Research Triangle Park, NC Laboratory testing often identifies toxicological hazards while providing little or no useful information about actual risks to human health. If the stringency of chemical regulation is to align with actual risks, then an understanding is needed, not just of hazard, but also of dose-response. Knowledge of the shape of the dose-response curve, combined with information on exposure, is the key to accurate risk prediction. To address this problem, we study dose-response behaviors of chemicals for which hazards are well characterized but for which actual health risks are uncertain. Two examples are covered in this presentation: (1) Chloroform is a high dose carcinogen in rats and mice while people are exposed to much lower

levels usually associated with water disinfection. Our work has established that the experimental conditions necessary for tumor formation in rodents are irrelevant to human environmental exposures. The key finding is that tumor formation is uniformly associated with chloroform exposures at sufficiently high levels to cause frank cytolethality. Chloroform levels associated with water disinfection are generally well below the levels associated with cytolethal effects. A computational model is being developed to allow prediction of the human dose-response for chloroform cytolethality.

(2) Anticholinesterase pesticides include the organophosphates (OPs) and the N-methyl carbamates

(NMCs). The Food Quality Protection Act of 1996 mandated that pesticides with a common mechanism of action, such the OPs and NMCs, be regulated together rather than individually. U.S. EPA has already developed a cumulative assessment for the OPs and the cumulative assessment for NMCs is underway. While the OP cumulative assessment was based on empirical dose-response modeling, physiologically based pharmacokinetic (PBPK) modeling is being used for the NMC assessment. We are working with the U.S. EPA to implement the PBPK models for NMCs and to assist in development of risk evaluations that take advantage of the PBPK modeling. This work will help to ensure the accuracy of dose-response predictions for acetylcholinesterase inhibition used in the NMC cumulative assessment.

Central to both of these examples is the development of computational models that integrate key physiological and chemical-specific data. The models, when sufficiently mature, can be used for prediction of dose-response behaviors that are a central component of risk assessment. The models thereby serve as bridges between the laboratory and risk assessment. To summarize, the main points of this presentation are that (a) hazard should not be equated with risk and, (b) appropriate combinations of laboratory studies and computational modeling can provide significant capabilities for improved prediction of the dose-response behaviors that underlie human health risks.


Poster 30 Comparison of Empirical and Mechanistic Models to Identify Relevant Toxicodynamic and Toxicokinetic Interactions Between Chemicals in a Mixture A. P. Freidig1, W. H. M. Heijne1, R. H. Stierum1, H. M. Wortelboer1, M. W. Schut1, H. A. El-Masri2, D. Moffett2, and J. P. Groten1 1TNO Nutrition and Food Research, Zeist, The Netherlands 2Agency for Toxic Substances and Disease Registry, Atlanta, GA The goal of this research project is to improve risk assessment of chemical mixtures with different modes of action using mechanistic models to describe interactions between mixture components. In a three-year project, mechanistic models are being established for both, toxicokinetic and toxicodynamic interactions within a quaternary mixture of lead acetate, methyl mercury, benzene and trichloroethylene (TCE), four chemicals which are expected to act by different modes of action. In a pilot study a methodology of toxicogenomic fingerprinting and multivariate statistics was applied to micro-array data of rat liver samples exposed to single compounds and mixtures. Thereby, a TCE-specific fingerprint was identified, which indicates an independent mode of action for TCE liver toxicity. Furthermore, in vitro experiments were performed in rat hepatocyte sandwich cultures to study the impact of each of the four compounds in the study on liver enzyme activities of CYP2E1 and GST, both of which are known to be relevant for the metabolism of benzene and trichloroethylene. Computational PBTK (physiologically based toxicokinetic) models were used to connect in-vitro with in vivo experiments. Estimates for thresholds of toxicokinetic interaction were made for the four tested compounds.


Poster 32 An Analysis of the Need for an Additional Toxicokinetic Uncertainty Factor for Neonates N. V. Corea and A. G. Renwick Clinical Pharmacology Group, University of Southampton, UK The objective of this study was to evaluate the interspecies and age-related differences in the internal dose (toxicokinetics) of chemicals that are cleared from the body by the major routes of elimination in humans. The in vivo pharmacokinetics of compounds, which are probe substrates for different pathways of adult human xenobiotic metabolism, were evaluated experimentally in immature and adult rats, and compared with the equivalent published pharmacokinetic values for the same compounds in human neonates, infants, children and adults. The data provided a scientific assessment of the appropriateness of the default interspecies uncertainty factor when the reference dose (RfD) value is based on effects seen in young rats. Several therapeutic drugs have been identified from published studies, for which data are available in young and adult humans, that are probe substrates for the metabolic (Phase I and Phase II metabolism) and renal elimination of chemicals. Plasma kinetic profiles were defined in groups of adult and 10-day-old rats administered a particular probe substrate. Animals received a single intraperitoneal dose of 5 mg/kg caffeine, 50 mg/kg caffeine or 50 mg/kg theophylline (metabolized by CYP1A2), 5 mg/kg midazolam (metabolized by CYP3A4), 200 mg/kg chloramphenicol (metabolized by glucuronyltransferase), or 500 mg/kg amoxicillin (eliminated by renal clearance). Clearance values (ml/min/kg) were compared to equivalent published data in human neonates, infants, children, and adults. The ratios of the clearance in rats to the clearance in age-equivalent humans for each substrate were then compared to the default inter-species toxicokinetic factor of 4. Elimination was generally more efficient in rats compared to age-equivalent humans, and the default factor was slightly exceeded for human neonates for chloramphenicol and caffeine (at both doses). However, the clearance of midazolam was much higher in immature rats and the default interspecies factor of 4.0 was exceeded by 7.3-fold for neonates, although this may have been a consequence of first pass metabolism following intraperitoneal administration in the test animal. Overall an additional 10-fold factor for infants and children as proposed by the Food Quality Protection Act (FQPA) would be excessive in relation to toxicokinetic differences between young rats and human neonates, infants, and children.


Poster 33 Dose-Response Modeling with Monoalkylphthalates: Assessing Cellular Targets for Mechanisms-Based Risk Assessment M. E. Andersen and R. A. Clewell Division of Computational Biology, CIIT Centers for Health Research, Research Triangle Park, NC Some dialkylphthalate plasticizers, including di-n-butylphthalate (DBP) and diethylhexylphthate (DEHP) cause testicular toxicity. These toxic responses are associated with monoalkylester metabolites (MBP and MEHP, respectively). In adults, these monoesters target Sertoli cells. During fetal development, MBP causes reduced testicular testosterone production by fetal Leydig cells in male fetuses. The molecular target(s) for the phthalate monoesters, i.e., the initial interaction of monoesters with molecular targets that initiate steps leading to toxicity, in these two cell types is unknown. This uncertainty in mechanism of action is the most significant impediment to accurate risk assessment with these phthalates. We have developed pharmacokinetic (PK) models for synthesis and clearance of fetal testosterone (T) and adapted a physiologically based pharmacokinetic (PBPK) model for distribution of phthalates in the adult rat to estimate blood concentrations of monoalkylphthalates associated with biological responses. The biological responses were reduced fetal T and induction of peroxisomal enzymes in rat liver. This induction occurs through activation of peroxisomal proliferation activating receptor (PPAR)-α by the monoesters. Dose-response modeling for peroxisomal enzyme induction, based on tissue phthalates predicted from our PBPK model, indicated a half-maximal induction with daily doses that produced a peak blood concentration of 87 µM MBP. Half-maximal inhibition of T synthesis occurred at daily doses to the dam that produced a peak blood concentration in the adult rat of ~ 10 µM. Prior research in the literature found that MBP inhibits cytoplasmic phospholipase A2 (cPLA2) in platelets in vitro with a much higher half-maximal inhibition concentration, 340 µM. cPLA2 hydrolyzes phospholipid substrates, releasing free arachidonic acid (AA). The biology of the PPAR and cPLA2 signaling pathways and our dose-response modeling suggests that the molecular targets for phthalates in adults and in fetal tissues may be AA-related signaling networks in which the monophthalates act through protein sensors that usually recognize AA and its metabolites. These sensors would include PPAR-α and cPLA2. These phthalate monoesters may block cPLA2 activation in the cascades initiated by gonadotropin stimulation of Sertoli and Leydig cells, accounting for both fetal and adult responses. Our current research focuses on refining these PK models for dose-response assessments in adult and fetal stages. Future efforts will directly evaluate AA metabolism in cells or cell-lines from testes to develop models for AA-related signaling processes in this tissue. The goal of our work, in combination with other anti-androgen projects at CIIT Centers for Health Research, is to understand the mechanistic basis of the relationship between phthalate concentrations and reduced T in order to reduce uncertainties in the human risk assessment for phthalates.


Poster 34

Non-Linear Dose-Response Relationships for Developmental Responses: An Example with Defeminization by Estrogenic Xenobiotics W. C. J. Chung1, M. E. Andersen2, and R. J. Handa1

1Department of Biomedical Sciences, Colorado State University, Fort Collins, CO 2CIIT Center for Health Research, Research Triangle Park, NC We are developing data sets and modeling tools to assess risks of exposures to low doses of estrogenic compounds on brain development. Early postnatal exposure to gonadal steroid hormones have rapid and marked effects on the morphological and neurochemical phenotype of the vertebrate brain. However, these exposures used gonadal steroid hormone dosages from milligrams to micrograms and may not reflect a true physiological response of the developing brain to lower doses of gonadal steroid hormones. To address the question of linearity in responses to estradiol during development, we have examined the dose-response relationship of dosages of 17β-estradiol (E) in the nanogram range given to neonates at birth (postnatal day (PN) 0) on the progestin receptor immunoreactivity in male and female rat brain on PN 4. Our results confirmed earlier studies describing more PR immunoreactive (IR) cells in sham/vehicle-treated males than in vehicle-treated females on PN 4. No sex difference in the number of PR-IR was found between castrated/vehicle-treated males and vehicle-treated females. The overall total number of PR-IR cells in the medial preoptic nucleus (MPN) significantly changed with increasing levels of estradiol in males and females. Moreover, total number of PR-IR MPN cells significantly changed with sex and there was an significant interaction between E and sex. A dose of 100 ng E significantly induced PR immunoreactivity in neurons of castrated males, while a dose of a 1000 ng E was required to significantly induce PR immunoreactivity in females. In addition to the studies with E, we have also evaluated the effects of synthetic estrogenic compounds on estrogen-dependent gene expression in the neonatal rat brain (i.e., the presence of PR-IR cells in the PN 4 male and female rat MPN). The compounds tested included propylpryazole-triol (i.e., estrogen receptor (ER) α selective agonist), diarylpropionitrole (i.e., ER β selective agonist) and 2,2 bis (p-hydroxyphenyl) -1,1,1-trichloroethane (i.e., ER α agonist/ER β antagonist) a metabolite of the pesticide and endocrine disrupter methoxychlor.


Poster 35 Potential Activation of Human Estrogen Receptors in HepG2 Cells by Genistein, Genistein-Glucuronide, and Genistein-Sulfate S.J. Borghoff, H. D. Parkinson, S. M. Ross, K. Gaido, and M. Sochaski. CIIT Centers for Health Research, Research Triangle Park, NC Genistein (Gen) is a phytoestrogen present in soy and virtually all rodent diets. It has been shown to activate both estrogen receptor (ER) α and β in vitro. Genistein has also been reported to elicit estrogenic effects in developing rodents. Following oral administration, genistein is found in plasma predominantly conjugated to glucuronic acid (GA) and to a lesser degree to sulphate (S). However, the major metabolite in fetal plasma following Gen exposure in utero is Gen-S. Because both Gen-S and Gen-GA are the major metabolites of Gen in vivo, these metabolites should not be overlooked as possible contributors to estrogenic effects of Gen. The objective of this study was to determine if conjugates of Gen activate ERα or ERβ as does free Gen. HepG2 human hepatoma cells were transfected with various concentrations of receptor plasmids encoding human ERα or ERβ, and complement 3-luciferase and pCMV β-galactosidase plasmids. Transfected cells were treated with various concentrations of either estradiol, Gen, Gen-GA (isolated from bile of Gen-treated rats), Gen-S (chemically synthesized), or DMSO (control vehicle). Following 24-hr incubation, cell lysates were assayed for luciferase activity (receptor activation) and β-galactosidase activity (transfection control). Estradiol and Gen activated both ER α and β, however, neither Gen-GA nor Gen-S activated either receptor or antagonized the ability of estradiol to activate these receptors. These data demonstrate that conjugation eliminates the estrogenic activity of Gen and that it is important to determine the ratio of free Gen to conjugated forms when estimating estrogenic potency.


Poster 36 A Multi-endpoint Strategy for Determining Mechanism of Action of Thyroid Active Chemicals S. J. Degitz and J. E. Tiegte U.S. Environmental Protection Agency To facilitate the process of identifying the mechanisms by which xenobiotics perturb the thyroid pathway, we scientists have adopted a multiple endpoint strategy using the amphibian model, Xenopus laevis. The research approach differs somewhat from that used for the estrogen/androgen pathway in that we have placed little emphases on examining xenbiotic-receptor interaction. A recent workshop on thyroid hormone disruption concluded that there is very little evidence that thyroid pathway disruption occurs through direct action on the thyroid hormone receptor and that disruption generally occurs elsewhere in the pathway. Theoretically, normal thyroid hormone homeostasis and action can be disrupted at several sites in the pathway, including: the hypothalamohyphophysial-thyroid axis, thyroid hormone synthesis, thyroid hormone transport, metabolic conversion of thyroid hormone to active and inactive forms, receptor-mediated transcriptional effects of T3, and nonspecific effects. Xenobiotic chemicals which are known to interfere with normal thyroid hormone homeostasis and action have been shown to act primarily by (1) up-regulation of catabolism of thyroid hormone, (2) inhibition of iodide uptake, (3) inhibition of T4 synthesis, or (4) inhibition and/or up-regulation of deiodinases. Several chemicals are also known to associate with the carrier protein, tranthyretin, with affinities sufficiently high to competitively displace T4, thereby altering circulating T4 concentrations. To this point, the specific actions of xenobiotics have been identified on a case-by-case basis. Given the fact that the xenobotics may have their action at different points in the thyroid pathway, it is advantageous to develop a suite of endpoints that allows one to ascertain where in the pathway impact is occurring. Our strategy makes use of recently developed molecular approaches in combination with classical in vivo methods. These approaches include analysis of thyroid pathway specific gene expression, analysis of differential protein expression, and advanced biochemical analysis of the endogenous hormone, the related synthetic precursors and degradation products. These endpoints are being assessed following challenges with thyroid active chemicals with known mechanisms of action to determine if the responses have characteristic patterns which can be used as signatures of the given mechanism. Once characteristic patterns have been elucidated for this suite of model chemicals, efforts will focus on testing chemicals whose mechanism of action have yet to be defined.


Poster 37

Development of a Reptile Egg Screening Assay for Endocrine-Disrupting Chemicals

L. G. Talent1, and D. M. Janz2 1Oklahoma State University, Stillwater, OK 2University of Saskatchewan, Saskatoon, Saskatchewan, Canada The purpose of our research is to evaluate the effects of endocrine disrupting chemicals on one of the most common and widespread groups of lizards in North America and to develop a reptile-egg screening assay for detecting endocrine disrupting chemicals. We evaluated different populations of fence lizards from across the United States and identified a population of western fence lizard, Sceloporus occidentalis, from California as a good candidate for a laboratory reptile model. Fence lizard embryos are sensitive to estrogenic and thyroid inhibiting chemicals. Post-anal scale development, a secondary sex characteristic of males, is inhibited by an estrogenic chemical (17α-ethinylestradiol). Therefore, post-anal scale development appears to be a good endpoint for evaluating exposure to estrogenic chemicals. In addition, both male and female embryos develop copulatory organs, i.e., hemipenal structures, during early development. Later in development, females resorb their hemipenal structures. Exposure to ethinylestradiol also causes males to reduce the size or resorb their hemipenal structures, indicating that hemipenal development and size is another endpoint that is useful for detecting estrogenic chemicals. In addition, the length of incubation is extended and the hatching process is inhibited in fence lizard embryos that are exposed to perchlorate, a thyroid-affecting chemical.

AUTHOR INDEX

2004 LRI Annual Science Meeting 63 Author Index

Alapaty, A. Poster No. 1 p. 27 Galbraith, H. Poster No. 4 p. 30Andersen, M.E. Presentation p. 16 Gennings, B.B. Poster No. 14 p. 40 Poster No. 7 p. 33 Poster No. 15 p. 41 Poster No. 33 p. 58 Gerhart, J. Presentation p. 9 Poster No. 34 p. 59 Gold, A. Poster No. 6 p. 32Asgharian, B. Poster No. 11 p. 37 Gollapudi, B.B. Poster No. 14 p. 40Ball, L.M. Poster No. 6 p. 32 Poster No. 15 p. 41Binkowski, F. Poster No. 1 p. 27 Grebendov, A.J. Poster No. 4 p. 30Blanchfield, P. Poster No. 19/20 p. 45 Gross, T.S. Poster No. 22 p. 48Borgert, C.J. Poster No. 22 p. 48 Groten, J.P. Poster No. 30 p. 56Borgoff, S.J. Poster No. 26 p. 52 Guan, H. Poster No. 1 p. 27 Poster No. 35 p. 60 Gundersen, H-J.G. Poster No. 18 p. 44Bos-Kuijpers, M.H.M. Poster No. 18 p. 44 Hall, T.A. Poster No. 3 p. 29Boverhof, D.R. Poster No. 15 p. 41 Handa, R.J. Poster No. 34 p. 59Brewer, L. Poster No. 24 p. 50 Hanna, A. Poster No. 1 p. 27Buck, G.M. Poster No. 10 p. 36 Harkema, J.R. Poster No. 28 p. 54Burton, G.A. Poster No. 2 p. 28 Heagerty, P.J. Poster No. 10 p. 36Carney, E.W. Poster No. 14 p. 40 Heijne, W.H.M. Poster No. 30 p. 56 Poster No. 15 p. 41 Hunt, K. Poster No. 24 p. 50Chaisson, C.F. Poster No. 17 p. 43 Hussain, I. Poster No. 21 p. 47Charles, G.D. Poster No. 14 p. 40 Janz, D.M. Poster No. 37 p. 62 Poster No. 15 p. 41 Jarabek, A.M. Poster No. 7 p. 33Chatfield, R. Poster No. 1 p. 27 Jaspers, I. Poster No. 5 p. 31Chung, W.C.J. Poster No. 34 p. 59 Jayaraj, K. Poster No. 6 p. 32Clements, W. Poster No. 2 p. 28 Jeffries, H.E. Poster No. 5 p. 31Clewell, H.J. Presentation p. 18 Johnson, B.A. Poster No. 9 p. 35 Poster No. 16 p. 42 Johnson, D.L. Poster No. 3 p. 29Clewell, R.A. Poster No. 33 p. 58 Kalisak, D.L. Poster No. 8 p. 34Clinkenbeard, R.E. Poster No. 3 p. 29 Kaminski, N.E. Poster No. 28 p. 54Coldham, N.G. Poster No. 26 p. 52 Kapustka, L.A. Poster No. 4 p. 30Conolly, R.B. Poster No. 17 p. 43 Kaufmann, W.S.H. Poster No. 18 p. 44 Poster No. 29 p. 55 Kepler, G.M. Poster No. 8 p. 34Corea, N.V. Poster No. 32 p. 57 Kidd, K. Poster No. 19/20 p. 45Cox, L.A. Poster No. 13 p. 39 Kimbell, J.S. Poster No. 7 p. 33de Groot, D.M.G. Poster No. 18 p. 44 Poster No. 8 p. 34Degitz, S.J. Poster No. 36 p. 61 Poster No. 11 p. 37Dietert, R. Poster No. 21 p. 47 Klapper, D. Poster No. 6 p. 32Dorman, D.C. Poster No. 7 p. 33 Kupper, L.L. Poster No. 9 p. 35Dukic, V.M. Poster No. 10 p. 36 Lammers, J.H.C.M. Poster No. 18 p. 44El-Masri, H.A. Poster No. 30 p. 56 Lin, Y. Poster No. 9 p. 35Esmen, N.E. Poster No. 3 p. 29 Linkov, I. Poster No. 4 p. 30Esswein, R. Poster No. 1 p. 27 Louis, T.A. Poster No. 10 p. 36Evans, B. Poster No. 19/20 p. 45 Luxon, M. Poster No. 4 p. 30Farraj, A. Poster No. 28 p. 54 Lynch, C.D. Poster No. 10 p. 36Findlay, D. Poster No. 19/20 p. 45 Marsh, J. Poster No. 21 p. 47Fitch, K. Poster No. 21 p. 47 Mathis, E.T. Poster No. 17 p. 43Foster, W.G. Presentation p. 14 Mathur, R. Poster No. 1 p. 27 Poster No. 12 p. 38 McVey, M. Poster No. 4 p. 30Freidig, A.P. Poster No. 30 p. 56 Miller, F.J. Poster No. 27 p. 53Gaido, K. Poster No. 35 p. 60 Mills, K. Poster No. 19/20 p. 45

AUTHOR INDEX

Author Index 64 2004 LRI Annual Science Meeting

Moffett, D. Poster No. 30 p. 56 Moss, O. Poster No. 11 p. 37 Murray, N. Poster No. 24 p. 50 Nylander-French, L.A. Poster No. 6 p. 32 O'Callaghan, J.P. Poster No. 18 p. 44 Oris, J. Poster No. 2 p. 28 Pakkenberg, B. Poster No. 18 p. 44 Palace, V. Presentation p. 15 Poster No. 19/20 p. 45 Parkinson, H.D. Poster No. 35 p. 60 Paterson, M. Poster No. 19/20 p. 45 Piepenbrink, M. Poster No. 21 p. 47 Podenmski, C. Poster No. 19/20 p. 45 Polston, E.K. Presentation p. 13 Poster No. 23 p. 49 Price, P.S. Poster No. 17 p. 43 Rappaport, S.M. Poster No. 9 p. 35 Renwick, A.G. Poster No. 32 p. 57 Richardson, R.B. Poster No. 8 p. 34 Ross, S.M. Poster No. 35 p. 60 Ryan, L.M. Poster No. 10 p. 36 Salki, A. Poster No. 19/20 p. 45 Schisterman, E.F. Poster No. 10 p. 36 Schlosser, P.M. Poster No. 7 p. 33 Poster No. 26 p. 52 Schut, M.W. Poster No. 30 p. 56 Segal, R.A. Poster No. 8 p. 34 Seidel, S.D. Poster No. 15 p. 41 Sexton, K.G. Poster No. 5 p. 31 Shinde, D.V. Poster No. 3 p. 29 Sochaski, M. Poster No. 35 p. 60 Stierum, R.H. Poster No. 30 p. 56 Talent, L.G. Poster No. 37 p. 62 Tank, S. Poster No. 24 p. 50 Taylor, D.J. Poster No. 9 p. 35 Tedder, D.R. Poster No. 17 p. 43 Thomas, R.S. Presentation p. 17 Poster No. 25 p. 51 Tiegte, J.E. Poster No. 36 p. 61 Tran, H.T. Poster No. 26 p. 52 Trifonov, A.G. Poster No. 4 p. 30 Trumble, A. Poster No. 10 p. 36 Waalkens-Berendsen, D. Poster No. 18 p. 44 Wong, B. Poster No. 11 p. 37 Wortelboer, H.M. Poster No. 30 p. 56 Yocum, J. Poster No. 4 p. 30 Young, J.S. Poster No. 17 p. 43 Zacharewski, T.R. Poster No. 14 p. 40 Poster No. 15 p. 41 Zager, M.G. Poster No. 26 p. 52

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