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Record Count: 12
To sort columns alphabetically or numerically, click on the column
header (Title, Principal Investigator, Institution, City, ST, Award Code, or
Pubs).
DESCRIPTION (provided by applicant): Arsenic contamination of drinking water is a global health concern since exposure to arsenic has been linked with increased rates of cancer of skin, bladder, lung, liver and kidney. Arsenic is a known carcinogen; however the molecular mechanisms by which arsenic induces carcinogenesis remain undefined. Recent studies have shown that arsenite, an inorganic trivalent species of arsenic, induces oxidative stress when administered at clinically relevant concentrations in cell culture models. Selenium, a required micronutrient for mammalian cell growth and survival, is a component of several critical enzymes (thioredoxin reductase and glutathione peroxidases) with a role in defense against reactive oxygen species (ROS). Our laboratory has recently discovered that arsenite is a competitive inhibitor of selenophosphate synthetase, the first committed step in the pathway for insertion of selenium into selenoproteins. We have also determined that clinically relevant concentrations of arsenite inhibited the incorporation of selenium into selenoproteins in both keratinocytes (HaCat) and HeLa cells. Based on these observations, our hypothesis is that the increase in ROS observed in cells treated with arsenicals is due to inhibition of SPS and subsequent decreased levels of thioredoxin reductase (TrxR) and glutathione peroxidases (Gpx). We will test this hypothesis by: 1) Determining the effect of trivalent and pentavalent arsenicals on the incorporation of selenium into selenoproteins, 2.) Determining the level of thioredoxin reductase and glutathione peroxidase activity in HaCat cells treated with trivalent or pentavalent arsenicals while altering the nutritional source of selenium, and 3) Determining whether trivalent or pentavalent arsenicals inhibit either of the two human selenophosphate synthetase enzymes (SPS1 and SPS2) in vitro. The proposed experiments are designed to identify the arsenical(s) which affect selenoprotein synthesis and the molecular mechanism(s) involved. Relevance to Public Health: Arsenic is a known carcinogen, and selenium is an important micronutrient. We have evidence that certain arsenic compounds can block the synthesis of selenoproteins, which are important in the defense against oxidative damage that can lead to cancer. We believe arsenic acts to promote cancer by blocking the cell's ability to make selenoproteins. We will test this hypothesis and in doing so perhaps find selenium compounds that can counteract the negative effects of arsenic exposure.
Crisp Terms/Key Words: enzyme induction /repression, enzyme inhibitor, enzyme biosynthesis, mutant, growth media, arsenic, selenium, environment related neoplasm /cancer, nutrition related neoplasm /cancer, chemical carcinogenesis, dietary trace element, nutrition related tag, free radical oxygen, glutathione peroxidase, phosphotransferase, affinity chromatography, protein biosynthesis, protein purification, radiotracer, keratinocyte, environmental toxicology, NAD(P)H oxidoreductase, cell line, oxidative stress, selenoprotein
DESCRIPTION (provided by applicant): The Ah-receptor (AHR) is a ligand activated transcription factor that belongs to the family of basic-helix-loop-helix/PER-ARNT-SIM (bHLH/PAS) proteins. AHR-mediated signaling has been extensively investigated in the C57BL/6J mouse model system in an attempt to define the components of the pathway, understand protein and DNA interactions and define specific changes in gene expression that likely impact human health. There has been less emphasis placed on the degradation of AHR protein especially as it relates turning the AHR signaling pathway off. Unfortunately, the proteins that modulate the degradation of the AHR and the domains of the AHR that are necessary for degradation remain to be identified. Thus, multifaceted approaches must be initiated to define candidate target enzymes involved in the degradation process and obtain "gain of function" mutants of the AHR. Therefore, the central hypothesis of this proposal is that defects in protein degradation will be manifest as "gain of function" mutants of the AHR. Three sets of studies are proposed to address this hypothesis and isolate the first ligand- activated gain of function mutants in the AHR pathway. First, studies will be carried out that utilize a bacterial two-hybrid screen to identify proteins interacting with the COOH-terminal domain of the AHR. Second, cell lines expressing a stably integrated GFP tagged AHR will be used to carry out a genetic screen for cells that do not degrade the AHR. Finally, yeast strains will be generated that express a functional AHR signaling pathway. Specific genes involved in protein degradation will be deleted from these strains to assess their role in AHR degradation. Thus, the proposed experiments utilize candidate gene approaches as well as genetic screens and employ mammalian cells, yeast and bacteria to address this hypothesis. The mechanism involved in turning off AHR-mediated signaling and the regulation of this pathway are especially critical with respect to halogenated aromatic compounds that are not readily metabolized or cleared from an organism. In addition, since modeling of AHR signaling and risk assessment in humans relies heavily on understanding the complete signaling pathway, it is essential to determine how the AHR pathway is attenuated. Statement of Relevance: Human health can be adversely affected by exposure to environmental contaminants. For chemicals that act through association with endogenous proteins and modulation of gene regulation, it is essential to understand how the pathways are regulated. Understanding how the Ah receptor protein is degraded is critical to risk assessment of chemicals typified by 2,3,7,8, tetrachlorodibezo-p-dioxin (Dioxin), and will be applicable to other toxicologically relevant receptors (i.e. steroid hormone receptors).
DESCRIPTION (provided by applicant): Dichloroacetate (DCA) is an environmentally important xenobiotics, widely distributed in our biosphere. It is also employed as an investigational drug for treatment of congenital forms of lactic acidosis (CLA). The cardinal manifestations of chronic DCA exposure are reversible hepatotoxicity and peripheral neuropathy (PN) in humans and hepatic neoplasia and reversible neurotoxicity in rodents. The etiologies of these adverse effects are obscure as in the relevance of the toxicological findings in animals to human risk assessment. DCA is dehalogenated by a glutathione transferase that is identical to maleylacetoacetate isomerase (MAAI), a key enzyme in tyrosine catabolism. Inhibition of MAAI by DCA causes accumulation of certain tyrosine catabolites, such as maleylacetone (MA), and heme precursors, such as ((-aminolevulinate ((-ALA) that may be responsible for its toxicity. The principal objective of this competitive renewal application is to elucidate the molecular mechanisms of DCA toxicity in rodents and humans by addressing the following specific aims and hypotheses: Specific Aim 1: Quantify the changes in tyrosine and heme metabolism in the urine of humans following exposure to DCA alone or in combination with pharmacologic inhibition of tyrosine catabolism. This aim tests the postulate that the chemical NTBC, which inhibits an early step in tyrosine catabolism, will prevent or mitigate the clinical toxicity of DCA. Specific Aim 2: Determine the mechanism of DCA neurotoxicity in peripheral nerves of dosed humans undergoing treatment with DCA. These studies test the hypotheses 1) that the PN from DCA exposure is primarily due to selected disruption of neuronal cell metabolism, as a result of a direct effect of DCA and from accumulation of catabolites and (-ALA, and 2) that inhibition of this latter effect by NTBC mitigates neurologic damage. Specific Aim 3: Determine the mechanism of DCA neurotoxicity in peripheral nerves of dosed rats and in cultures of neuronal cells from rodents. This aim will address the postulate that DCA induces neuropathology in rats by the same basic mechanisms as occurs in humans. It will also test the hypothesis that susceptibility to DCA PN is age-dependent, being increased in young animals in whom myelination of peripheral nerves is an active, ongoing process.
DESCRIPTION (provided by applicant): For several years we have researched the kinetics, metabolism and human toxicology of dichloroacetate (DCA), a potentially harmful metabolite of trichloroethylene (TCE). This new proposal continues this research and also integrates it with new human studies of chloral hydrate (CH), which is both a primary metabolite of TCE and a precursor of DCA. Indeed, recent advances in our understanding of the mechanisms of DCA and CH biotransformation in vivo provide new insight into the possible causes of the adverse effects of chronic DCA and CH exposure in humans. Specifically, we have discovered that 1) children metabolize CH to DCA, 2) CH and DCA alter each other's clearance in vivo, 3) DCA is dechlorinated to glyoxylate, 4) dechlorination is mediated by hepatic maleylacetoacetate isomerase (MAAI), an enzyme of tyrosine catabolism, 5) DCA inhibits MAAI expression in animals causing, buildup of potentially toxic tyrosine intermediates, and 6) these metabolites may be responsible for DCA (and CH) toxicity. Humans exhibit polymorphisms of MAAI that may possess different kinetic properties toward DCA. In turn, human haplotype variability may influence DCA's kinetics and toxicology. These hypotheses will be tested by accomplishing the following Specific Aims: Specific Aim 1: Quantify the influence of DCA, at exposure levels ranging from environmental (mu g/kg/d) to clinical (mg/kg/d), on human liver MAAI and tyrosine catabolism. This aim tests the hypotheses that 1) there is a dose-dependent effect of DCA on human hepatocellular tyrosine metabolism in general and on the accumulation of potentially hepatotoxic tyrosine intermediates in particular, 2) DCA inhibits hepatic MAAI expression in vivo in humans in a dose-dependent manner and 3) DCA, tyrosine and/or their metabolites form adducts with MAAI. Specific Aim 2: Establish the relationship between human MAAI haplotype and DCA and tyrosine metabolism. This aim tests the postulates that MAAI haplotype determines, and thus can predict, 1) dose-dependent DCA kinetics and biotransformation and 2) DCA's effects on tyrosine metabolism. Specific Aim 3: Determine the in vivo kinetics and biotransformation of CH in healthy adults and the influence of CH and DCA on each other's metabolism. This specific aim will examine three postulates 1) CH is-metabolized in adults to DCA, 2) CH, via DCA formation, inhibits its own metabolism and that of tyrosine and 3) these effects are dependent upon exposure level but not upon gender.
Crisp Terms/Key Words: environmental exposure, liquid chromatography mass spectrometry, clinical research, enzyme activity, glutathione transferase, toxicology, protein structure function, intermolecular interaction, biotransformation, liver pharmacology, liver metabolism, human subject, trichloroethylene, dichloroacetate, chloral hydrate, genetic polymorphism, pharmacogenetics, pharmacokinetics, dosage, tyrosine, gas chromatography mass spectrometry, chemical kinetics, adduct, aminoacid metabolism, adult human (21+)
DESCRIPTION (provided by applicant): Cyanobacteria, or "blue-green algae," particularly in freshwater habitats, are recognized to produce an array of potently toxic compounds. Growing evidence indicates that toxigenic cyanobacteria represent a serious concern to human and environmental health. Development of novel approaches will be key to understanding both current and emerging roles of cyanobacterial toxins in environmental health. The proposed research will utilize the zebrafish (Danio rerio) embryo as a toxicological model to investigate toxic or otherwise bioactive metabolites from cyanobacteria. This research will isolate and culture cyanobacteria from two, ecologically unique and distinct freshwater microbial communities, namely the Florida Everglades (and associated waterways in South Florida) and the northern Great Lakes. Cyanobacterial isolates will be screened using the zebrafish embryo as a model of vertebrate development specifically to identify metabolites which interfere with developmental pathways (i.e. developmental toxins). The bioassay will be further utilized to guide initial purification of identified toxins toward the goal of chemical and biological characterization of potentially novel compounds. In addition, the zebrafish model will be used specifically to investigate a ubiquitous class of bacterial toxins, the lipopolysaccharides (LPSs), from cyanobacteria. Though LPSs (or "endotoxins") from Gram-negative heterotophic eubacteria are well described, the same compounds from cyanobacteria have been largely unexplored. However, emerging evidence indicates that cyanobacterial LPSs may have specific effects on certain "detoxifying enzymes," and a subsequent interactive effects with other toxins. Accordingly, the proposed research will further utilize the zebrafish embryo model to test several hypothesis related to the effects of LPSs, specifically from isolated strains of Microcystis, on heavy metal toxicity and accumulation. The proposed research will contribute significantly to our growing understanding of the role of freshwater cyanobacterial toxins in environmental health.
Transient and permanent reproductive dysfunction in fish has been linked to chemicals that disrupt the
endocrine system, but the mechanisms involved are unclear. Many endocrine-disrupting chemicals (EDCs)
interact with sex hormone receptors by acting as agonists to induce gene expression at inappropriate times
or act as antagonists to prevent the normal functioning of the receptors. Other EDCs may act indirectly by
altering the processes involved in regulating sex steroid synthesis and metabolism. Poor reproduction and
altered plasma hormone levels have been observed in largemouth bass living in ecosystems polluted with
organochlorine pesticides (OCPs), including the Superfund site at Lake Apopka and its surrounding muck
farms, suggesting that OCP exposure has adversely affected their reproduction.
In past research, we have identified three estrogen receptors (a, pi, and (32) in largemouth bass, which
exhibit tissue specific expression and respond differently to 17-p-estradiol at the message and activity level.
These differences are important to control normal reproductive function. We have evidence that EDCs can
alter their normal expression and activity patterns, possibly disrupting reproduction. In addition, we have
preliminary evidence that expression of enzymes involved in the synthesis and metabolism of EDCs is
affected by the OCPs. The proposed studies will test the central hypothesis that exposure of largemouth bass
to concentrations of OCPs found in the Lake Apopka region disrupts endocrine system function by sex
hormone receptor mediated and sex hormone receptor independent mechanisms. In this proposal, we have
designed a set of interrelated experiments from the organismal level where pleiotropic effects of OCPs can
be measured to the cellular level where specific molecular mechanisms of action can be assessed. We are
exposing largemouth bass to methoxychlor (and its metabolites), p,p' DDE, dieldrin and toxaphene. Our
specific aims include 1: Develop biomarkers of exposure to organochlorinated pesticides in vivo, via the use
of microarrays and novel proteomics methodologies; 2: Determine the effect of in vivo OCP exposure on LME
steroid synthesis and metabolism; and 3: Evaluate the effects of OCPs on the molecular mechanisms of
action of the three estrogen receptors. We plan on using exposures to compounds with known modes of
action as controls to determine mode of action specific gene expression patterns against which we will
compare the patterns of gene expression changes induced by the OCPs. We will check these pattern
changes against reproductive endpoints including the ability of eggs to mature (germinal vesicle breakdown)
and sperm function (sperm motility). Fish are useful as sentinels of environmental quality and are perfect
models to monitor adverse effects in reproduction caused by contaminants in superfund sites. By
extrapolation fish can report on the potential of OCPs to harm human health.
DESCRIPTION (provided by applicant): Development of the ovarian follicle is a complex process integral to female fertility. The objective of this proposal is to expand understanding of the mechanism(s) causing the formation of aberrant multi-oocytic follicles (MOFs) during vertebrate folliculogenesis using a novel study species, the American alligator. Multioocytic follicles (MOFs, also called polyovular follicles) have been associated with decreased success rates of in vitro fertilization and increased embryonic loss. MOFs have been studied in laboratory animals, however, a natural population of alligators has been identified in which 100% of neonatal females exhibit this pathology apparently due to environmental contaminant exposure. In comparison, reference alligator populations exhibit no MOFs or a very low frequency. Due to the conserved nature of folliculogenesis, we propose this comparative approach to address the mechanism(s) associated with the development of MOFs. Laboratory research has identified both estrogens and the transforming growth factor inhibin as factors that can induce MOF formation. The central hypothesis of this proposal is that the disruption of the ovarian hormonal milieu by exogenous factors alter ovarian follicle development leading to MOFs. The studies proposed will test this hypothesis by investigating 3 related specific aims. Aim 1 will examine the gene expression patterns for selective components of the estrogen and inhibin/activin signaling pathways in cellular compartments of normal and multioocytic ovarian follicles using laser capture microdissection and quantitative RT-PCR. Aim 2 will examine estrogen-induced perturbation of the inhibin A/Activin A signaling pathway and their potential role in altered granulosa cell proliferation or apoptosis. This will be tested by embryonic exposure to estradiol followed by FSH challenge during 2 life stages. Aim 3 will examine environment estrogen or pharmaceutical anti-estrogen-induced perturbation of the inhibin A/Activin A signaling pathway and their role in altered granulosa cell proliferation or apoptosis to firmly establish if an environmental perturbation leads to altered inhibin/activin signaling. In total, the studies described in this application provide a novel approach to provide new insights into normal follicle development, establish mechanisms by which normal development could be disrupted leading to aberrant ovarian pathologies such as multi-oocytic follicles, and offer new insights into the regulatory functions of inhibin/activins and estrogens in this important reproductive process. Understanding and documenting the conserved mechanisms of induction of MOFs among vertebrates will strengthen the causal and mechanistic relationships, allowing a better understanding of this phenomenon in human populations.
DESCRIPTION (provided by applicant): Microarrays are a powerful way to measure the impact of contaminants in the environment, and sheepshead minnows (Cyprinodon variegatus) are the most commonly used species in salt water environmental testing. In this proposal, we explain how we will develop and validate a large microarray (5,000+ genes) in sheepshead minnows. We will conduct a number of short and long term exposures on sheepshead minnows using several anthropogenic compounds (pyrene, copper, cadmium, and bisphenol A), then use the microarrays to measure the gene expression signatures for these compounds. This data, along with a variety of physiological endpoints, will be the basis for a relational database. We will analyze the data and decipher the patterns resulting from the various exposures to identify the unique fingerprints for each compound. Microarrays in sheepshead minnows will round out EcoArray's offering of microarrays for environmentally significant aquatic species. Measurement and analysis of environmental contaminants is very important to the EPA in its Superfund monitoring activities, and the sheepshead minnow is an important species that is routinely used for the monitoring of coastal superfund sites. In addition, diagnosis of ecotoxological effects at the gene level in sentinel species like sheepshead minnow offer the promise of future ability to tie ecotoxicology to human health, a goal of the National Center for Toxicogenomics. Experiments by our research group and others have shown that microarrays can be used to detect changes in gene expression caused by exposure to contaminants, and it is clear that contaminants have unique genetic signatures. Because many contaminants act at the gene level to induce or repress gene expression through both receptor-mediated and non-receptor mediated pathways, microarrays can help to elucidate signaling pathways that are affected. In general, microarrays offer a direct, effective way to provide detailed data about the biological effects of the environment on animals. In addition, analysis using microarrays is generally considerably less expensive than current testing methods. With the successful completion of this grant, we will incorporate the sheepshead minnow microarray and its database into our existing product line and sell it to the EPA, USGS, researchers in academia, as well as industrial concerns interested in compound screening and environmental monitoring/remediation.
This project will result in a fully developed microarray for ecotoxicology testing in salt water. This microarray, together with the database this project will begin, can provide detailed, gene-level data on the biological impact of a chemical or an environment. In so doing, it can lead to assessment not only of water quality, but also of the implications of chemicals and environments for human health.
DESCRIPTION (provided by applicant): The overall emphasis of this research is to discover patterns of gene expression that affect the health of individuals subjected to chronic chemical exposure. This provides a framework to understand basic biological responses to environmental stress and toxicants and identifies genes that are markers of chemical exposure. The challenges are to identify those genes that respond to toxicant exposure, to discover novel gene interactions and to improve knowledge of complex regulatory networks and cross-communication between different pathways in both health and disease. Concern about the adverse effects of environmental pollutants results from evidence that these chemical pollutants are affecting the health of individuals. However, not all individuals are affected equally: within populations, individuals show variable susceptibility to chemical exposure. Often, the molecular basis for this difference is unknown. This research will use microarray analyses of natural populations to 1) identify genes and patterns of gene expression that are affected by environmental chemicals and 2) examine individual differences in toxic responses. Elucidating the molecular mechanisms underlying biological effects of environmental chemical exposure is important both to understand the responses of animals to chronic chemical exposure and to identify molecular markers of susceptibility associated with increased risk in populations of animals, including humans. Analyzing a variety of different individuals within natural populations is important to identify factors associated with variation in susceptibility. Importantly, non-random patterns of gene expression within and between the polluted populations will be identified. These patterns will provide indices of pollutant exposure in other polluted populations.
Crisp Terms/Key Words: bioaccumulation, environmental exposure, microarray technology, genetic susceptibility, Osteichthyes, DNA damage, environmental toxicology, pesticide resistance, hepatotoxin, animal population genetics, population genetics, gene expression, genetic marker, environmental contamination
DESCRIPTION (provided by applicant): Polycyclic aromatic hydrocarbons (PAHs) are environmental and occupational carcinogens that are produced by the incomplete combustion of organic material, such as that from tobacco, coal, and petroleum. In addition to causing lung cancer, exposure to high levels of PAHs is hypothesized to contribute to atherosclerosis and lead to increased rates of cardiovascular disease. Considering the number of deaths attributable to tobacco smoke exposure, ambient air pollution, and occupational hazards, PAHs may be a significant contributor to the high prevalence and mortality rate of cardiovascular disease. However, a clear exposure-response relationship between PAH and cardiovascular disease has not been demonstrated. While PAH exposure has been shown to be associated with indicators of cardiovascular disease in animal experiments, this relationship has not been studied adequately in large human populations. Using data from the National Health and Nurition Examination Survey (NHANES) collected since 1999, this proposed study will investigate associations between urinary metabolites of PAHs and self-reported and laboratory measures of cardiovascular disease. Information regarding individuals' occupation, exposure to tobacco smoke and residence in areas with clean indoor air laws will be included to investigate factors associated with PAH exposure. The specific aims are: 1. To determine if exposure to polycyclic aromatic hydrocarbons (PAHs) is associated with clinical and self- reported indicators of cardiovascular disease in children and adults in the U.S. 2. To determine if exposure to environmental sources of PAHs at home or in the workplace is associated with elevated levels of urinary metabolites of PAHs. 3. To determine if exposure to high levels of ambient air pollution is associated with elevated levels of urinary metabolites of PAHs when controlled for smoking status and exposure to secondhand smoke. Cardiovascular disease remains the single largest cause of death in the United States. Understanding the contribution of PAH exposure to the development of cardiovascular disease will provide important insight into new strategies to prevent cardiovascular disease. Results of this proposed study can be used to influence policy decisions and resource allocation for the prevention of cardiovascular disease.
DESCRIPTION (provided by applicant): Environmental arsenic is a known toxin that has detrimental effects on a variety of organ systems including the male urogenital tract. Increased exposure to environmental arsenic is associated with male infertility. The manifestations of infertility in these men include production of abnormal sperm, decreased sperm counts and decreased sperm mobility. While there is evidence to support the relationship between arsenic and male infertility the exact mechanism is unclear. Androgen receptors (AR) are ligand-activated transcription factors that regulate genes involved in development of the male urogenital tract, secondary sexual characteristics, and sperm production. Since the androgen receptor plays a critical role in male fertility it may be a target of arsenic toxicity in the urogenital tract. We have found that sodium arsenite causes a dose dependent inhibition of AR transcriptional activity. This proposal seeks to assess the effect of sodium arsenite on AR-regulated gene transcription. Three different models of androgen target tissues, TM4 (sertoli), PNT2 (normal prostate), and PCS (prostate cancer) cells, will be used for these studies. Our long- term goal is to elucidate the mechanism by which arsenic may regulate AR activity leading to male infertility.
DESCRIPTION (provided by applicant):
This proposal is for the competitive renewal of our ARCH program at Florida International University (an MSI). Our partner (RIU) institution is the University of Miami which hosts an NIEHS MFBS Center and an NIEHS/ NSF OHH Center. A two fold approach which includes faculty development and infrastructure development is being used. The MFBS Center encompasses two principle research themes; Marine Toxins and dietary risk and Marine Models of human disease. The OHH Center focuses on algal toxins and genomics and pathogenic microbes. For this ARCH program, we will continue to pursue our research theme of Algal Toxins and will expand our research into Trace Metals in the environment. This program will be a collaborative effort between eight faculty from four departments at FIU and six faculty from four departments at the University of Miami. An Administrative and Planning Core will provide administrative and clerical support to ARCH investigators at FIU and will coordinate activities with UM. In addition to an administrative assistant, who will assist with day to day operations, this core will be composed of three committees to direct and oversee the progress of the program; the IAC, the Executive Committee and the EAC. Program Evaluation will be provided through this core by an independent evaluator. The principal goal of the ARCH program is to establish externally funded, independent research programs in environmental health sciences which are relevant to the NIEHS mission at FIU. The Administrative and planning core will promote these goals through seminars and workshops, and will track the progress of ARCH faculty and provide this data to the independent evaluator. The Research Program Development Core will be composed of two core facilities and research and pilot projects. We propose the continuation of the Toxic Algae Culture Facility to support the research and two pilot projects. This core provides materials and services to ARCH researchers. In addition, we propose to add a Trace Metal Core Facility, which will support three of the proposed pilot projects. This facility will provide analytical support in trace metal analysis to ARCH researchers. Pilot projects will permit ARCH faculty at FIU to acquire preliminary data for research program grant applications.
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