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1.
Artigo em Inglês | MEDLINE | ID: mdl-31989135

RESUMO

Trichloroethylene (TCE), an industrial solvent and degreaser, is an environmental toxicant that contaminates over half of Superfund sites, is a known carcinogen, and is linked to congenital defects and neurodegenerative disease. The developmental toxicity of TCE near ecologically relevant levels needs further characterization in order to better assess health risks of exposure. In this study, the toxicodynamics of TCE in the zebrafish (Danio rerio) model was investigated through the establishment of a LC50 concentration and by monitoring the acute developmental toxicity of ecologically relevant concentrations (0, 5, 50, and 500 parts per billion; ppb) of TCE during two different exposure lengths (1-72 hours post fertilization (hpf) and 1-120 hpf). Acute developmental toxicity was assessed by monitoring survival and hatching, larval morphology, larval heart rate, and behavioral responses during an embryonic photomotor response test and a larval visual motor response test. Embryonic exposure to TCE was associated with decreased percent hatch at 48 hpf, altered larval morphology, increased heart rate, and altered behavioral responses during the photomotor response test and visual motor response test. Larval morphology and behavioral alterations were more pronounced in the 1-120 hpf exposure length trials. The observed alterations suggest developmental TCE toxicity is still a concern at regulatory concentrations and that timing of exposure influences developmental toxicity.

2.
Chem Res Toxicol ; 33(1): 95-118, 2020 Jan 21.
Artigo em Inglês | MEDLINE | ID: mdl-31625720

RESUMO

Unpredicted human safety events in clinical trials for new drugs are costly in terms of human health and money. The drug discovery industry attempts to minimize those events with diligent preclinical safety testing. Current standard practices are good at preventing toxic compounds from being tested in the clinic; however, false negative preclinical toxicity results are still a reality. Continual improvement must be pursued in the preclinical realm. Higher-quality therapies can be brought forward with more information about potential toxicities and associated mechanisms. The zebrafish model is a bridge between in vitro assays and mammalian in vivo studies. This model is powerful in its breadth of application and tractability for research. In the past two decades, our understanding of disease biology and drug toxicity has grown significantly owing to thousands of studies on this tiny vertebrate. This Review summarizes challenges and strengths of the model, discusses the 3Rs value that it can deliver, highlights translatable and untranslatable biology, and brings together reports from recent studies with zebrafish focusing on new drug discovery toxicology.

3.
Environ Pollut ; 258: 113712, 2019 Dec 13.
Artigo em Inglês | MEDLINE | ID: mdl-31875570

RESUMO

How environmental chemicals can affect and exert their toxic effect at a molecular level has gained significant interest in recent years, not only for understanding their immediate health implications over exposed individuals, but also for their subsequent progeny. Atrazine (ATZ) is a commonly used herbicide in the U.S. and a long-suspected endocrine disrupting chemical. The molecular mechanism conferring long-term adverse health outcomes, however, remain elusive. Here, we explored changes in epigenetic marks that arise after exposure to ATZ at selected doses using image-based analysis coupled with data clustering. Significant decreases in methylated CpG (meCpG) and histone 3 lysine 9 tri-methylated (H3K9me3) were observed in the selected human cell line with a clear spatial preference. Treating cells with ATZ leads to the loss of a subpopulation of cells with high meCpG levels as identified in our clustering and histogram analysis. A similar trend was observed in H3K9me3 potentially attributing to the cross-talking between meCpG and H3K9me3. Changes in meCpG are likely to be associated with alterations in epigenetic enzyme expression levels regulating meCpG and persist after the removal of ATZ source which collectively provide a plausible mechanism for long-term ATZ-induced toxicity.

4.
Birth Defects Res ; 111(16): 1234-1236, 2019 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-31385457
5.
Medicines (Basel) ; 6(2)2019 May 30.
Artigo em Inglês | MEDLINE | ID: mdl-31151179

RESUMO

Most neurodegenerative diseases are currently incurable, with large social and economic impacts. Recently, there has been renewed interest in investigating natural products in the modern drug discovery paradigm as novel, bioactive small molecules. Moreover, the discovery of potential therapies for neurological disorders is challenging and involves developing optimized animal models for drug screening. In contemporary biomedicine, the growing need to develop experimental models to obtain a detailed understanding of malady conditions and to portray pioneering treatments has resulted in the application of zebrafish to close the gap between in vitro and in vivo assays. Zebrafish in pharmacogenetics and neuropharmacology are rapidly becoming a widely used organism. Brain function, dysfunction, genetic, and pharmacological modulation considerations are enhanced by both larval and adult zebrafish. Bioassay-guided identification of natural products using zebrafish presents as an attractive strategy for generating new lead compounds. Here, we see evidence that the zebrafish's central nervous system is suitable for modeling human neurological disease and we review and evaluate natural product research using zebrafish as a vertebrate model platform to systematically identify bioactive natural products. Finally, we review recently developed zebrafish models of neurological disorders that have the potential to be applied in this field of research.

6.
Sci Transl Med ; 11(475)2019 01 16.
Artigo em Inglês | MEDLINE | ID: mdl-30651321

RESUMO

The therapeutic success of interventions targeting glucokinase (GK) activation for the treatment of type 2 diabetes has been limited by hypoglycemia, steatohepatitis, and loss of efficacy over time. The clinical characteristics of patients with GK-activating mutations or GK regulatory protein (GKRP) loss-of-function mutations suggest that a hepatoselective GK activator (GKA) that does not activate GK in ß cells or affect the GK-GKRP interaction may reduce hyperglycemia in patients with type 2 diabetes while limiting hypoglycemia and liver-associated adverse effects. Here, we review the rationale for TTP399, an oral hepatoselective GKA, and its progression from preclinical to clinical development, with an emphasis on the results of a randomized, double-blind, placebo- and active-controlled phase 2 study of TTP399 in patients with type 2 diabetes. In this 6-month study, TTP399 (800 mg/day) was associated with a clinically significant and sustained reduction in glycated hemoglobin, with a placebo-subtracted least squares mean HbA1c change from baseline of -0.9% (P < 0.01). Compared to placebo, TTP399 (800 mg/day) also increased high-density lipoprotein cholesterol (3.2 mg/dl; P < 0.05), decreased fasting plasma glucagon (-20 pg/ml; P < 0.05), and decreased weight in patients weighing ≥100 kg (-3.4 kg; P < 0.05). TTP399 did not cause hypoglycemia, had no detrimental effect on plasma lipids or liver enzymes, and did not increase blood pressure, highlighting the importance of tissue selectivity and preservation of physiological regulation when targeting key metabolic regulators such as GK.

7.
Bio Protoc ; 8(17)2018 Sep 05.
Artigo em Inglês | MEDLINE | ID: mdl-30547052

RESUMO

This protocol details a method to analyze two tissue samples at the transcriptomic level using microarray analysis, ingenuity pathway analysis (IPA) and gene set enrichment analysis (GSEA). Methods such as these provide insight into the mechanisms underlying biological differences across two samples and thus can be applied to interrogate a variety of processes across different tissue samples, conditions, and the like. The full method detailed below can be applied to determine the effects of muscle-specific Notch1 activation in the mdx mouse model and to analyze previously published microarray data of human liposarcoma cell lines.

8.
Toxics ; 6(4)2018 Nov 09.
Artigo em Inglês | MEDLINE | ID: mdl-30423906

RESUMO

Tungsten is a refractory metal that is used in a wide range of applications. It was initially perceived that tungsten was immobile in the environment, supporting tungsten as an alternative for lead and uranium in munition and military applications. Recent studies report movement and detection of tungsten in soil and potable water sources, increasing the risk of human exposure. In addition, experimental research studies observed adverse health effects associated with exposure to tungsten alloys, raising concerns on tungsten toxicity with questions surrounding the safety of exposure to tungsten alone or in mixtures with other metals. Tungsten is commonly used as an alloy with nickel and cobalt in many applications to adjust hardness and thermal and electrical conductivity. This review addresses the current state of knowledge in regard to the mechanisms of toxicity of tungsten in the absence or presence of other metals with a specific focus on mixtures containing nickel and cobalt, the most common components of tungsten alloy.

9.
J Proteomics ; 186: 71-82, 2018 08 30.
Artigo em Inglês | MEDLINE | ID: mdl-30012420

RESUMO

Atrazine (ATZ), the second most commonly used herbicide in the United States, is an endocrine disrupting chemical linked to cancer and a common drinking water contaminant. This study further investigates ATZ-related developmental toxicity by testing the following hypotheses in zebrafish: the effects of embryonic ATZ exposure are dependent on timing of exposure; embryonic ATZ exposure alters brain development and function; and embryonic ATZ exposure changes protein abundance in carcinogenesis-related pathways. After exposing embryos to 0, 0.3, 3, or 30 parts per billion (ppb) ATZ, we monitored the expression of cytochrome P450 family 17 subfamily A member 1 (cyp17a1), glyoxalase I (glo1), ring finger protein 14 (rnf14), salt inducible kinase 2 (sik2), tetratricopeptide domain 3 (ttc3), and tumor protein D52 like 1 (tpd52l1) at multiple embryonic time points to determine normal expression and if ATZ exposure altered expression. Only cyp17a1 had normal dynamic expression, but ttc3 and tpd52l1 had ATZ-related expression changes before 72 h. Larvae exposed to 0.3 ppb ATZ had increased brain length, while larvae exposed to 30 ppb ATZ were hypoactive. Proteomic analysis identified altered protein abundance in pathways related to cellular function, neurodevelopment, and genital-tract cancer. The results indicate embryonic ATZ toxicity involves interactions of multiple pathways. SIGNIFICANCE: This is the first report of proteomic alterations following embryonic exposure to atrazine, an environmentally persistent pesticide and common water contaminant. Although the transcriptomic alterations in larval zebrafish with embryonic atrazine exposure have been reported, neither the time at which gene expression changes occur nor the resulting proteomic changes have been investigated. This study seeks to address these knowledge gaps by evaluating atrazine's effect on gene expression through multiple time points during embryogenesis, and correlating changes in gene expression to pathological alterations in brain length and functional changes in behavior. Finally, pathway analysis of the proteomic alterations identifies connections between the molecular changes and functional outcomes associated with embryonic atrazine exposure.


Assuntos
Atrazina/farmacologia , Embrião não Mamífero/efeitos dos fármacos , Regulação da Expressão Gênica no Desenvolvimento/efeitos dos fármacos , Proteômica , Animais , Atrazina/toxicidade , Encéfalo/crescimento & desenvolvimento , Relação Dose-Resposta a Droga , Desenvolvimento Embrionário , Disruptores Endócrinos/farmacologia , Disruptores Endócrinos/toxicidade , Herbicidas/farmacologia , Herbicidas/toxicidade , Larva/efeitos dos fármacos , Proteínas/efeitos dos fármacos , Poluentes Químicos da Água/farmacologia , Peixe-Zebra/embriologia
10.
Metallomics ; 10(3): 463-473, 2018 03 01.
Artigo em Inglês | MEDLINE | ID: mdl-29485154

RESUMO

Coal mining is one of the economic activities with the greatest impact on environmental quality. At all stages contaminants are released as particulates such as coal dust. The first aim of this study was to obtain an aqueous coal dust extract and characterize its composition in terms of trace elements by ICP-MS. In addition, the developmental toxicity of the aqueous coal extract was evaluated using zebrafish (Danio rerio) after exposure to different concentrations (0-1000 ppm; µg mL-1) to establish acute toxicity, morphology and transcriptome changes. Trace elements within the aqueous coal dust extract present at the highest concentrations (>10 ppb) included Sr, Zn, Ba, As, Cu and Se. In addition, Cd and Pb were found in lower concentrations. No significant difference in mortality was observed (p > 0.05), but a delay in hatching was found at 0.1 and 1000 ppm (p < 0.05). No significant differences in morphological characteristics were observed in any of the treatment groups (p > 0.05). Transcriptomic results of zebrafish larvae revealed alterations in 77, 61 and 1376 genes in the 1, 10, and 100 ppm groups, respectively. Gene ontology analysis identified gene alterations associated with the development and function of connective tissue and the hematological system, as well as pathways associated with apoptosis, the cell cycle, transcription, and oxidative stress including the MAPK signaling pathway. In addition, altered genes were associated with cancer; connective tissue, muscular, and skeletal disorders; and immunological and inflammatory diseases. Overall, this is the first study to characterize gene expression alterations in response to developmental exposure to aqueous coal dust residue from coal mining with transcriptome results signifying functions and systems to target in future studies.


Assuntos
Carvão Mineral/toxicidade , Tecido Conjuntivo/patologia , Poeira/análise , Regulação da Expressão Gênica no Desenvolvimento , Doenças Hematológicas/patologia , Doenças do Sistema Imunitário/patologia , Inflamação/patologia , Peixe-Zebra/crescimento & desenvolvimento , Animais , Tecido Conjuntivo/efeitos dos fármacos , Tecido Conjuntivo/metabolismo , Poluentes Ambientais/toxicidade , Doenças Hematológicas/induzido quimicamente , Doenças Hematológicas/genética , Doenças do Sistema Imunitário/induzido quimicamente , Doenças do Sistema Imunitário/genética , Inflamação/induzido quimicamente , Inflamação/genética , Transcriptoma , Peixe-Zebra/genética
11.
Toxicol Sci ; 163(1): 5-12, 2018 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-29471431

RESUMO

The laboratory zebrafish (Danio rerio) is now an accepted model in toxicologic research. The zebrafish model fills a niche between in vitro models and mammalian biomedical models. The developmental characteristics of the small fish are strategically being used by scientists to study topics ranging from high-throughput toxicity screens to toxicity in multi- and transgenerational studies. High-throughput technology has increased the utility of zebrafish embryonic toxicity assays in screening of chemicals and drugs for toxicity or effect. Additionally, advances in behavioral characterization and experimental methodology allow for observation of recognizable phenotypic changes after xenobiotic exposure. Future directions in zebrafish research are predicted to take advantage of CRISPR-Cas9 genome editing methods in creating models of disease and interrogating mechanisms of action with fluorescent reporters or tagged proteins. Zebrafish can also model developmental origins of health and disease and multi- and transgenerational toxicity. The zebrafish has many advantages as a toxicologic model and new methodologies and areas of study continue to expand the usefulness and application of the zebrafish.


Assuntos
Modelos Animais , Toxicologia/métodos , Peixe-Zebra , Animais , Comportamento Animal/efeitos dos fármacos , Embrião não Mamífero/efeitos dos fármacos , Desenvolvimento Embrionário/efeitos dos fármacos , Ensaios de Triagem em Larga Escala , Larva/efeitos dos fármacos , Toxicologia/tendências , Xenobióticos/toxicidade , Peixe-Zebra/embriologia , Peixe-Zebra/genética , Peixe-Zebra/fisiologia
12.
Cell Rep ; 22(7): 1810-1823, 2018 02 13.
Artigo em Inglês | MEDLINE | ID: mdl-29444433

RESUMO

MicroRNA-223 is known as a myeloid-enriched anti-inflammatory microRNA that is dysregulated in numerous inflammatory conditions. Here, we report that neutrophilic inflammation (wound response) is augmented in miR-223-deficient zebrafish, due primarily to elevated activation of the canonical nuclear factor κB (NF-κB) pathway. NF-κB over-activation is restricted to the basal layer of the surface epithelium, although miR-223 is detected throughout the epithelium and in phagocytes. Not only phagocytes but also epithelial cells are involved in miR-223-mediated regulation of neutrophils' wound response and NF-κB activation. Cul1a/b, Traf6, and Tab1 are identified as direct targets of miR-223, and their levels rise in injured epithelium lacking miR-223. In addition, miR-223 is expressed in cultured human bronchial epithelial cells, where it also downregulates NF-κB signaling. Together, this direct connection between miR-223 and the canonical NF-κB pathway provides a mechanistic understanding of the multifaceted role of miR-223 and highlights the relevance of epithelial cells in dampening neutrophil activation.


Assuntos
Inflamação/patologia , Queratinócitos/metabolismo , MicroRNAs/metabolismo , NF-kappa B/metabolismo , Neutrófilos/patologia , Transdução de Sinais , Nadadeiras de Animais/fisiologia , Animais , Sequência de Bases , Brônquios/citologia , Embrião não Mamífero/metabolismo , Células Epiteliais/metabolismo , Células HEK293 , Humanos , Inflamação/metabolismo , MicroRNAs/genética , Neutrófilos/metabolismo , Fagócitos/metabolismo , Regeneração , Peixe-Zebra/embriologia , Peixe-Zebra/genética , Proteínas de Peixe-Zebra/metabolismo
13.
Neurotoxicol Teratol ; 65: 60-69, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29074346

RESUMO

Developmental lead (Pb) exposure is linked to neurological health issues. Results from non-human primate and rodent studies suggest detrimental effects of an early life Pb exposure, showing transcriptional disturbances and pathological evidence of Alzheimer's disease in the adult animal brain. To elucidate the impacts of an embryonic Pb exposure on the adult brain, transcriptomic analysis was completed on the brain of zebrafish aged 12months exposed to a control treatment or to an embryonic 100µg/L Pb exposure by sex. In the adult female zebrafish brain, significant changes in expression profiles occurred in a number of genes involved in neurological disease and nervous system development and function. On the other hand, in adult males, a number of genes with significant expression alterations were found to be associated with cancer and tumors. p38 mitogen-activated protein kinase (p38 MAPK) was also indicated as an upstream regulator of observed gene expression changes. Western blot analysis confirmed activation of p38 MAPK in the form of phosphorylated p38 MAPK in the male zebrafish brain. In addition, we compared transcriptomic changes observed in this study to a previous study with an embryonic exposure of 10µg/L Pb by sex, showing unique sets of genes dependent on Pb concentration. Overall, these results show sex-specific and concentration-dependent disturbances of global gene expression patterns in the brain of adult zebrafish exposed to Pb during embryogenesis.


Assuntos
Desenvolvimento Embrionário/efeitos dos fármacos , Regulação da Expressão Gênica no Desenvolvimento/efeitos dos fármacos , Chumbo/toxicidade , Neoplasias/genética , Doenças do Sistema Nervoso/genética , Caracteres Sexuais , Peixe-Zebra/crescimento & desenvolvimento , Animais , Encéfalo/efeitos dos fármacos , Encéfalo/embriologia , Encéfalo/crescimento & desenvolvimento , Embrião não Mamífero/efeitos dos fármacos , Desenvolvimento Embrionário/genética , Perfilação da Expressão Gênica , Neoplasias/embriologia , Doenças do Sistema Nervoso/embriologia , Peixe-Zebra/embriologia , Peixe-Zebra/genética
14.
Food Chem Toxicol ; 109(Pt 1): 727-734, 2017 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-28859886

RESUMO

Atrazine, a herbicide used on agricultural crops is widely applied in the Midwestern United States as well as other areas of the globe. Atrazine frequently contaminates potable water supplies and is a suspected endocrine disrupting chemical. Previous studies have reported morphological, hormonal, and molecular alterations due to developmental and adulthood atrazine exposure; however, studies examining epigenetic alterations are limited. In this study, the effects of atrazine exposure on DNA methyltransferase (DNMT) activity and kinetics were evaluated. Global DNA methylation levels and dnmt expression in zebrafish larvae exposed to 0, 3, or 30 parts per billion (ppb) atrazine throughout embryogenesis was then assessed. Results indicate that atrazine significantly decreased the activity of maintenance DNMTs and that the inhibition mechanism can be described using non-competitive Michaelis-Menten kinetics. Furthermore, results show that an embryonic atrazine exposure decreases global methylation levels and the expression of dnmt4 and dnmt5. These findings indicate that atrazine exposure can decrease the expression and activity of DNMTs, leading to decreased DNA methylation levels.


Assuntos
Atrazina/toxicidade , Metilação de DNA/efeitos dos fármacos , DNA-Citosina Metilases/genética , Herbicidas/toxicidade , Proteínas de Peixe-Zebra/genética , Peixe-Zebra/genética , Animais , DNA-Citosina Metilases/metabolismo , Embrião não Mamífero/efeitos dos fármacos , Embrião não Mamífero/metabolismo , Desenvolvimento Embrionário , Feminino , Regulação da Expressão Gênica no Desenvolvimento , Masculino , Peixe-Zebra/embriologia , Peixe-Zebra/metabolismo , Proteínas de Peixe-Zebra/metabolismo
15.
Chemosphere ; 188: 599-607, 2017 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-28917212

RESUMO

On January 9, 2014, a chemical mixture containing crude methylcyclohexanemethanol (MCHM) contaminated the water supply of Charleston, West Virginia. Although the mixture was later identified as a mix of crude MCHM and stripped propylene glycol phenyl ethers, initial risk assessment focused on 4-MCHM, the predominant component of crude MCHM. The mixture's exact composition and the toxicity differences between 4-MCHM, crude MCHM, and the tank mixture were unknown. We analyzed the chemical composition of crude MCHM and the tank mixture via GC/MS and, based on identified spectra, found that crude MCHM and the tank mixture differed in chemical composition. To evaluate acute developmental toxicity, zebrafish embryos were exposed to 0, 1, 6.25, 12.5, 25, 50, or 100 parts per million (ppm; mg/L) of 4-MCHM, crude MCHM, or the tank mixture. The percent mortality and percent hatch, larval morphology alterations, and larval visual motor response test were used to establish toxicity profiles for each of the chemicals or mixtures. The acute toxicity differed between 4-MCHM, crude MCHM and the tank mixture with significant differences in survival, hatching, morphology, and locomotion at levels as low as the short-term screening level of 1 ppm, suggesting a need for further research into human health risks. This study is the first to evaluate the developmental toxicity of the tank mixture and highlights that studies evaluating risk should not assume the effects of 4-MCHM or crude MCHM are representative of the Tank 396 mixture.


Assuntos
Vazamento de Resíduos Químicos , Cicloexanos/toxicidade , Rios/química , Poluentes Químicos da Água/análise , Animais , Cicloexanos/análise , Exposição Ambiental , Cromatografia Gasosa-Espectrometria de Massas , Éteres Fenílicos , Abastecimento de Água/normas , West Virginia , Peixe-Zebra/embriologia
16.
J Med Chem ; 60(13): 5364-5376, 2017 07 13.
Artigo em Inglês | MEDLINE | ID: mdl-28657311

RESUMO

The 7-azaindenoisoquinolines are cytotoxic topoisomerase I (Top1) inhibitors. Previously reported representatives bear a 3-nitro group. The present report documents the replacement of the potentially genotoxic 3-nitro group by 3-chloro and 3-fluoro substituents, resulting in compounds with high Top1 inhibitory activities and potent cytotoxicities in human cancer cell cultures and reduced lethality in an animal model. Some of the new Top1 inhibitors also possess moderate inhibitory activities against tyrosyl-DNA phosphodiesterase 1 (TDP1) and tyrosyl-DNA phosphodiesterase 2 (TDP2), two enzymes that are involved in DNA damage repair resulting from Top1 inhibitors, and they produce significantly more DNA damage in cancer cells than in normal cells. Eighteen of the new compounds had cytotoxicity mean-graph midpoint (MGM) GI50 values in the submicromolar (0.033-0.630 µM) range. Compounds 16b and 17b are the most potent in human cancer cell cultures with MGM GI50 values of 0.063 and 0.033 µM, respectively. Possible binding modes to Top1 and TDP1were investigated by molecular modeling.


Assuntos
Antineoplásicos/farmacologia , DNA Topoisomerases Tipo I/metabolismo , Desenho de Drogas , Isoquinolinas/farmacologia , Inibidores da Topoisomerase I/farmacologia , Animais , Antineoplásicos/síntese química , Antineoplásicos/química , Proliferação de Células/efeitos dos fármacos , Clivagem do DNA , Relação Dose-Resposta a Droga , Ensaios de Seleção de Medicamentos Antitumorais , Humanos , Isoquinolinas/síntese química , Isoquinolinas/química , Modelos Moleculares , Estrutura Molecular , Relação Estrutura-Atividade , Inibidores da Topoisomerase I/síntese química , Inibidores da Topoisomerase I/química , Células Tumorais Cultivadas , Peixe-Zebra
17.
Artigo em Inglês | MEDLINE | ID: mdl-28111253

RESUMO

Atrazine is an agricultural herbicide used throughout the Midwestern United States that frequently contaminates potable water supplies resulting in human exposure. Using the zebrafish model system, an embryonic atrazine exposure was previously reported to decrease spawning rates with an increase in progesterone and ovarian follicular atresia in adult females. In addition, alterations in genes associated with distinct molecular pathways of the endocrine system were observed in brain and gonad tissue of the adult females and males. Current hypotheses for mechanistic changes in the developmental origins of health and disease include genetic (e.g., copy number alterations) or epigenetic (e.g., DNA methylation) mechanisms. As such, in the current study we investigated whether an atrazine exposure would generate copy number alterations (CNAs) in the zebrafish genome. A zebrafish fibroblast cell line was used to limit detection to CNAs caused by the chemical exposure. First, cells were exposed to a range of atrazine concentrations and a crystal violet assay was completed, showing confluency decreased by ~60% at 46.3µM. Cells were then exposed to 0, 0.463, 4.63, or 46.3µM atrazine and array comparative genomic hybridization completed. Results showed 34, 21, and 44 CNAs in the 0.463, 4.63, and 46.3µM treatments, respectively. Furthermore, CNAs were associated with previously reported gene expression alterations in adult male and female zebrafish. This study demonstrates that atrazine exposure can generate CNAs that are linked to gene expression alterations observed in adult zebrafish exposed to atrazine during embryogenesis providing a mechanism of the developmental origins of atrazine endocrine disruption.


Assuntos
Atrazina/toxicidade , Dosagem de Genes/efeitos dos fármacos , Regulação da Expressão Gênica/efeitos dos fármacos , Genoma/efeitos dos fármacos , Herbicidas/toxicidade , Mutagênicos/toxicidade , Peixe-Zebra/fisiologia , Animais , Encéfalo/efeitos dos fármacos , Encéfalo/embriologia , Encéfalo/metabolismo , Linhagem Celular , Hibridização Genômica Comparativa , Desenvolvimento Embrionário/efeitos dos fármacos , Disruptores Endócrinos/toxicidade , Feminino , Regulação da Expressão Gênica no Desenvolvimento/efeitos dos fármacos , Masculino , Concentração Osmolar , Ovário/efeitos dos fármacos , Ovário/embriologia , Ovário/metabolismo , RNA Mensageiro/metabolismo , Testículo/efeitos dos fármacos , Testículo/embriologia , Testículo/metabolismo , Transcriptoma/efeitos dos fármacos , Poluentes Químicos da Água/toxicidade , Peixe-Zebra/embriologia
18.
J Appl Toxicol ; 37(4): 400-407, 2017 04.
Artigo em Inglês | MEDLINE | ID: mdl-27535807

RESUMO

Developmental lead (Pb) exposure is suggested in laboratory studies to be a trigger for neurodegenerative diseases such as Alzheimer's disease (AD). Sortilin-related receptor, L (DLR class) A repeats-containing (SORL1) is a recently identified AD genetic risk factor. SORL1 has limited characterization in vertebrate models in comparison to other AD genetic risk factors. To characterize SORL1 further, protein sequence homology between humans, mice and zebrafish was analyzed and showed conservation of functional repeats and domain orientation. Next, spatial expression of sorl1 in zebrafish larvae was completed and diffuse expression in neural tissue that was not restricted to the brain was observed. Influences of sex and age on quantitative expression of sorl1 in the brain of adult zebrafish were then assessed. Sex-specific alteration of sorl1 expression transpired during the aging process in females. The zebrafish was then utilized to investigate the impacts of a 100 ppb embryonic Pb exposure on sorl1 expression and other known AD genetic risk factors. Sex-specific quantitative gene expression analysis was completed with adult zebrafish brain to compare those developmentally exposed to Pb or a control treatment, but no significant difference in sorl1 expression or other AD genetic risk factors was observed. Overall, this study provided characterization of sorl1 with changes in brain expression during aging being female-specific. This finding is in agreement with females being more prone to the onset of AD, but analysis of additional AD genetic risk factors is needed to facilitate our understanding of the impact of a 100 ppb embryonic Pb exposure. Copyright © 2016 John Wiley & Sons, Ltd.


Assuntos
Doença de Alzheimer/induzido quimicamente , Doença de Alzheimer/genética , Embrião não Mamífero/patologia , Proteínas Relacionadas a Receptor de LDL/genética , Intoxicação do Sistema Nervoso por Chumbo/genética , Proteínas de Peixe-Zebra/genética , Envelhecimento/patologia , Doença de Alzheimer/patologia , Animais , Encéfalo/patologia , Feminino , Intoxicação do Sistema Nervoso por Chumbo/patologia , Masculino , Sinais Direcionadores de Proteínas/genética , Fatores de Risco , Caracteres Sexuais , Peixe-Zebra
19.
Metallomics ; 9(2): 149-160, 2017 02 22.
Artigo em Inglês | MEDLINE | ID: mdl-27934997

RESUMO

Low-dose exposure to lead (Pb) is connected to developmental neurological alterations by inducing molecular changes, such as aberrant gene expression patterns. The attributing molecular mechanism, however, is not well-elucidated. In this study, we revealed epigenetic features and mechanisms that can alter gene expression patterns by identifying changes in DNA methyltransferase (DNMT) activity, expression pattern and DNA methylation level using moelcular studies and a zebrafish animal model. We characterized the effects of Pb on the activities of various DNMTs in vitro and determined the molecular role of Pb in modulating DNMT activity via kinetic experiments. An exposure of 100 or 500 ppb of Pb was found to significantly lower the activity of maintenance DNMTs. The inhibition mechanism can be described using non-competitive Michaelis-Menten kinetics. A zebrafish animal model was then used to assess the biological significance of our findings. An embryonic exposure to 100 or 500 ppb Pb resulted in a significant change in global methylation levels consistent with previous studies using human and rodent model. Our study also suggests that Pb exposure in zebrafish alters the expression patterns of dnmt3 and dnmt4 which are human DNMT3b orthologs. The knowledge from this study suggests that Pb exposure can affect the activity of maintenance DNMTs via non-competitive inhibition, which has not been reported previously. Meanwhile, the expression pattern of de novo methyltransferases can also be altered. Collectively, they result in a reduction in global DNA methylation level in Pb-exposed zebrafish model, which can be compared to findings in human and rodent studies.


Assuntos
Metilação de DNA/efeitos dos fármacos , Embrião não Mamífero/metabolismo , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Chumbo/toxicidade , Metiltransferases/antagonistas & inibidores , Proteínas de Peixe-Zebra/genética , Peixe-Zebra/metabolismo , Animais , Embrião não Mamífero/efeitos dos fármacos , Regulação da Expressão Gênica no Desenvolvimento/efeitos dos fármacos , Metiltransferases/genética , Regiões Promotoras Genéticas , Elementos de Resposta , Peixe-Zebra/genética , Proteínas de Peixe-Zebra/metabolismo
20.
Oncotarget ; 7(49): 81698-81714, 2016 Dec 06.
Artigo em Inglês | MEDLINE | ID: mdl-27835581

RESUMO

In this article we inspect the roles and functions of the methyl-CpG-binding domain protein 3 (MBD3) in human malignant glioma, to assess its potential as an epigenetic biomarker for prognosis. The regulatory effects of MBD3 on glioma transcriptome were first profiled by high-throughput microarray. Our results indicate that MBD3 is involved in both transcriptional activation and repression. Furthermore, MBD3 fine-controls a spectrum of proteins critical for cellular metabolism and proliferation, thereby contributing to an exquisite anti-glioma network. Specifically, the expression of MHC class II molecules was found to positively correlate with MBD3, which provides new insight into the immune escape of gliomagenesis. In addition, MBD3 participates in constraining a number of oncogenic non-coding RNAs whose over-activation could drive cells into excessive growth and higher malignancy. Having followed up a pilot cohort, we noted that the survival of malignant glioma patients was proportional to the content of MBD3 and 5-hydroxymethylcytosine (5hmC) in their tumor cells. The progression-free survival (PFS) and overall survival (OS) were relatively poor for patients with lower amount of MBD3 and 5hmC in the tissue biopsies. Taken together, this work enriches our understanding of the mechanistic involvement of MBD3 in malignant glioma.


Assuntos
Neoplasias Encefálicas/genética , Proteínas de Ligação a DNA/genética , Regulação Neoplásica da Expressão Gênica , Glioma/genética , 5-Metilcitosina/análogos & derivados , 5-Metilcitosina/metabolismo , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/mortalidade , Neoplasias Encefálicas/patologia , Linhagem Celular Tumoral , Proteínas de Ligação a DNA/metabolismo , Intervalo Livre de Doença , Perfilação da Expressão Gênica/métodos , Glioma/metabolismo , Glioma/mortalidade , Glioma/patologia , Antígenos HLA-D/metabolismo , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , MicroRNAs/genética , MicroRNAs/metabolismo , Análise de Sequência com Séries de Oligonucleotídeos , Projetos Piloto , Interferência de RNA , RNA Longo não Codificante/genética , RNA Longo não Codificante/metabolismo , RNA não Traduzido/genética , RNA não Traduzido/metabolismo , Transdução de Sinais , Fatores de Tempo , Transcrição Genética , Transfecção
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