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1.
Braz J Biol ; 83: e242942, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34468508

RESUMO

Thimerosal is an organomercurial compound, which is used in the preparation of intramuscular immunoglobulin, antivenoms, tattoo inks, skin test antigens, nasal products, ophthalmic drops, and vaccines as a preservative. In most of animal species and humans, the kidney is one of the main sites for mercurial compounds deposition and target organs for toxicity. So, the current research was intended to assess the thimerosal induced nephrotoxicity in male rats. Twenty-four adult male albino rats were categorized into four groups. The first group was a control group. Rats of Group-II, Group-III, and Group-IV were administered with 0.5µg/kg, 10µg/kg, and 50µg/kg of thimerosal once a day, respectively. Thimerosal administration significantly decreased the activities of catalase (CAT), superoxide dismutase (SOD), peroxidase (POD), glutathione reductase (GR), glutathione (GSH), and protein content while increased the thiobarbituric acid reactive substances (TBARS) and hydrogen peroxide (H2O2) levels dose-dependently. Blood urea nitrogen (BUN), creatinine, urobilinogen, urinary proteins, kidney injury molecule-1 (KIM-1), and neutrophil gelatinase-associated lipocalin (NGAL) levels were substantially increased. In contrast, urinary albumin and creatinine clearance was reduced dose-dependently in thimerosal treated groups. The results demonstrated that thimerosal significantly increased the inflammation indicators including nuclear factor kappaB (NF-κB), tumor necrosis factor-α (TNF-α), Interleukin-1ß (IL-1ß), Interleukin-6 (IL-6) levels and cyclooxygenase-2 (COX-2) activities, DNA and histopathological damages dose-dependently. So, the present findings ascertained that thimerosal exerted nephrotoxicity in male albino rats.


Assuntos
Estresse Oxidativo , Timerosal , Animais , Peróxido de Hidrogênio/metabolismo , Rim , Masculino , Ratos , Superóxido Dismutase/metabolismo , Timerosal/metabolismo , Timerosal/toxicidade
2.
Environ Res ; 188: 109734, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32544722

RESUMO

Aluminum and mercury are environmentally ubiquitous. Individually they are both neurotoxic elements with shared neuro-pathogenic pathways: oxidative stress, altered neurotransmission, and disruption of the neuroendocrine and immune systems. In the infant, Al and Hg differ in type of exposure, absorption, distribution (brain access), and metabolism. In environmentally associated exposure (breast milk and infant formulas) their co-occurrences fluctuate randomly, but in Thimerosal-containing vaccines (TCVs) they occur combined in a proprietary ratio; in these cases, low-doses of Thimerosal-ethylmercury (EtHg) and adjuvant-Al present the most widespread binary mixture in less developed countries. Although experimental studies at low doses of the binary Hg and Al mixture are rare, when studied individually they have been shown to affect neurological outcomes negatively. In invitro systems, comparative neurotoxicity between Al and Hg varies in relation to the measured parameters but seems less for Al than for Hg. While neurotoxicity of environmental Hg (mainly fish methyl-Hg, MeHg) is associated with neurobehavioral outcomes in children, environmental Al is not associated, except in certain clinical conditions. Therefore, the issues of their neurotoxic effects (singly or combined) are discussed. In the infant (up to six months) the organic-Hg and Al body burdens from a full TCV schedule are estimated to reach levels higher than that originating from breastfeeding or from high aluminum soy-based formulas. Despite worldwide exposure to both Al and Hg (inorganic Hg, MeHg, and Thimerosal/EtHg), our knowledge on this combined exposure is insufficient to predict their combined neurotoxic effects (and with other co-occurring neurotoxicants).


Assuntos
Mercúrio , Compostos de Metilmercúrio , Vacinas , Alumínio/toxicidade , Animais , Carga Corporal (Radioterapia) , Criança , Feminino , Humanos , Lactente , Mercúrio/toxicidade , Leite Humano , Síndromes Neurotóxicas/epidemiologia , Timerosal/toxicidade
3.
Int J Biol Macromol ; 154: 661-671, 2020 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-32198046

RESUMO

Thimerosal (TH), an organomercurial compound, is used as a preservative in vaccines and cosmetics. Its interaction with human hemoglobin (Hb) was investigated under physiological conditions using biophysical and biological assays, aiming to evaluate hazardous effects. TH interacts spontaneously with Hb (stoichiometry 2:1, ligand-protein), preferably by electrostatic forces, with a binding constant of 1.41 × 106 M-1. Spectroscopic data allows to proposing that TH induces structural changes in Hg, through ethylmercury transfer to human Hb-Cys93 residues, forming thiosalicylic acid, which, in turn, interacts with the positive side of the amino acid in the Hb-HgEt adduct chain. As a consequence, inhibition of Hb-O2 binding capacity up to 72% (human Hb), and 50% (human erythrocytes), was verified. Dose-dependent induction of TH forming advanced glycation end products (AGE) and protein aggregates (amyloids) was additionally observed. Finally, these results highlight the toxic potential of the use of TH in biological systems, with a consequent risk to human health.


Assuntos
Peptídeos beta-Amiloides/metabolismo , Produtos Finais de Glicação Avançada/metabolismo , Hemoglobinas/metabolismo , Oxigênio/metabolismo , Conservantes Farmacêuticos/toxicidade , Timerosal/toxicidade , Humanos
4.
Neurotoxicology ; 71: 6-15, 2019 03.
Artigo em Inglês | MEDLINE | ID: mdl-30503815

RESUMO

In humans, mutation of glycine 93 to alanine of Cu++/Zn++ superoxide dismutase type-1 (SOD1-G93 A) has been associated to some familial cases of Amyotrophic Lateral Sclerosis (ALS). Several evidence proposed the involvement of environmental pollutants that like mercury could accelerate ALS symptoms. SH-SY5Y cells stably transfected with SOD1 and G93 A mutant of SOD1 constructs were exposed to non-toxic concentrations (0.01 µM) of ethylmercury thiosalicylate (thimerosal) for 24 h. Interestingly, we found that thimerosal, in SOD1-G93 A cells, but not in SOD1 cells, reduced cell survival. Furthermore, thimerosal-induced cell death occurred in a concentration dependent-manner and was prevented by the Sirtuin 1 (SIRT1) activator Resveratrol (RSV). Moreover, thimerosal decreased the protein expression of transcription factor Downstream Regulatory Element Antagonist Modulator (DREAM), but not DREAM gene. Interestingly, DREAM reduction was blocked by co-treatment with RSV, suggesting the participation of SIRT1 in determining this effect. Immunoprecipitation experiments in SOD1-G93 A cells exposed to thimerosal demonstrated that RSV increased DREAM deacetylation and reduced its polyubiquitination. In addition, RSV counteracted thimerosal-enhanced prodynorphin (PDYN) mRNA, a DREAM target gene. Furthermore, cortical neurons transiently transfected with SOD1-G93 A construct and exposed to thimerosal (0.5 µM/24 h) showed a reduction of DREAM and an up-regulation of the prodynorphin gene. Importantly, both the treatment with RSV or the transfection of siRNA against prodynorphin significantly reduced thimerosal-induced neurotoxicity, while DREAM knocking-down potentiated thimerosal-reduced cell survival. These results demonstrate the particular vulnerability of SOD1-G93 A neuronal cells to thimerosal and that RSV via SIRT1 counteracts the neurodetrimental effect of this toxicant by preventing DREAM reduction and prodynorphin up-regulation.


Assuntos
Córtex Cerebral/efeitos dos fármacos , Córtex Cerebral/metabolismo , Neurônios/efeitos dos fármacos , Neurônios/metabolismo , Resveratrol/administração & dosagem , Transdução de Sinais , Superóxido Dismutase/metabolismo , Timerosal/toxicidade , Animais , Morte Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Encefalinas/metabolismo , Humanos , Proteínas Interatuantes com Canais de Kv/metabolismo , Precursores de Proteínas/metabolismo , Ratos Wistar , Proteínas Repressoras/metabolismo , Sirtuína 1/metabolismo , Superóxido Dismutase-1/metabolismo
5.
Biol Trace Elem Res ; 184(1): 7-15, 2018 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-28967039

RESUMO

Infant exposure to neurotoxic elements is a public health issue that needs monitoring with regard to breast milk composition. We studied six neurotoxic elements in breast milk samples at different stages of lactation in mothers from Porto Velho, Brazil. We used a flow-injection mercury system (FIMS) to determine total Hg concentrations and an inductively coupled plasma optical emission spectrometer (ICP-OES) to determine the concentrations of Al, As, Cd, Pb, and Mn in 106 donors of a human milk bank. Association rules analyses were applied to determine the pattern of binary and ternary mixtures of the measured exposants. The metal concentration was mostly below the limit of detection (LOD) for Cd (99%), Pb (84%), and Hg (72%), and it was above the LOD for As (53%), Mn (60%), and Al (82%), respectively. Median concentrations (dry weight) of Al, As, Hg, Mn, and Pb were 1.81 µg/g, 13.8 ng/g, 7.1 ng/g, 51.1 ng/g, and 0.43 µg/g, respectively. Al is singly the most frequent element to which infants are exposed. Occurring binary combination (> LOD) was 56% for Al-Mn, 41% for Al-As, 22% for Al-Hg, and 13% for Al-Pb. In 100% of neonates, exposure to Al-ethylmercury (EtHg) occurred through immunization with thimerosal-containing vaccines (TCV). Association rules analysis revealed that Al was present in all of the multilevel combinations and hierarchical levels and that it showed a strong link with other neurotoxic elements (especially with Mn, As, and Hg). (a) Nursing infants are exposed to combinations of neurotoxicants by different routes, dosages, and at different stages of development; (b) In breastfed infants, the binary exposures to Al and total Hg can occur through breast milk and additionally through TCV (EtHg and Al);


Assuntos
Aleitamento Materno/efeitos adversos , Exposição Materna/efeitos adversos , Metais Pesados/toxicidade , Alumínio/toxicidade , Cádmio/toxicidade , Compostos de Etilmercúrio/toxicidade , Feminino , Humanos , Chumbo/toxicidade , Manganês/toxicidade , Leite Humano , Mães , Timerosal/toxicidade
6.
J Int Med Res ; 45(2): 407-438, 2017 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-28415925

RESUMO

The wide range of factors associated with the induction of autism is invariably linked with either inflammation or oxidative stress, and sometimes both. The use of acetaminophen in babies and young children may be much more strongly associated with autism than its use during pregnancy, perhaps because of well-known deficiencies in the metabolic breakdown of pharmaceuticals during early development. Thus, one explanation for the increased prevalence of autism is that increased exposure to acetaminophen, exacerbated by inflammation and oxidative stress, is neurotoxic in babies and small children. This view mandates extreme urgency in probing the long-term effects of acetaminophen use in babies and the possibility that many cases of infantile autism may actually be induced by acetaminophen exposure shortly after birth.


Assuntos
Acetaminofen/efeitos adversos , Analgésicos não Narcóticos/efeitos adversos , Transtorno Autístico/etiologia , Transtorno Autístico/fisiopatologia , Estresse Oxidativo , Aspartame/administração & dosagem , Aspartame/metabolismo , Aspartame/toxicidade , Transtorno Autístico/diagnóstico , Criança , Pré-Escolar , Feminino , Ácido Fólico/efeitos adversos , Humanos , Hiperbilirrubinemia/complicações , Hiperbilirrubinemia/fisiopatologia , Lactente , Inflamação , Masculino , Metais Pesados/toxicidade , Organofosfatos/toxicidade , Gravidez , Fatores de Risco , Timerosal/toxicidade , Vitamina B 12/efeitos adversos
7.
Toxicology ; 370: 86-93, 2016 Aug 31.
Artigo em Inglês | MEDLINE | ID: mdl-27693314

RESUMO

A presynaptic protein SNAP-25 belonging to SNARE complex which is instrumental in intracellular vesicular trafficking and exocytosis, has been implicated in hyperactivity and cognitive abilities in some neuropsychiatric disorders. The unclear etiology of the behavior disrupting neurodevelopmental disabilities in addition to genetic causes most likely involves environmental factors. The aim of this in vitro study was to test if various suspected developmental neurotoxins can alter SNAP-25 mRNA and protein expression in neurons. Real-time PCR and Western blotting analyses were used to assess SNAP-25 mRNA and protein levels in primary cultures of rat cerebellar granule cells (CGCs). The test substances: tetrabromobisphenol-A (TBBPA), thimerosal (TH), silver nanoparticles (NAg), valproic acid (VPA) and thalidomide (THAL), were administered to CGC cultures at subtoxic concentrations for 24h. The results demonstrated that SNAP-25 mRNA levels were increased by 49 and 66% by TBBPA and THAL, respectively, whereas VPA and NAg reduced these levels to 48 and 64% of the control, respectively. The SNAP-25 protein content in CGCs was increased by 79% by TBBPA, 25% by THAL and 21% by NAg; VPA and TH reduced these levels to 73 and 69% of the control, respectively. The variety of changes in SNAP-25 expression on mRNA and protein level suggests the diversity of the mechanism of action of the test substances. This initial study provided no data on concentration-effect relations and on functional changes in CGCs. However it is the first to demonstrate the effect of different compounds that are suspected of causing neurodevelopmental disabilities on SNAP-25 expression. These results suggest that this protein may be a common target for not only inherited but also environmental modifications linked to behavioral deficits in neurodevelopmental disabilities.


Assuntos
Nanopartículas Metálicas/toxicidade , Bifenil Polibromatos/toxicidade , Prata/toxicidade , Proteína 25 Associada a Sinaptossoma/metabolismo , Talidomida/toxicidade , Timerosal/toxicidade , Ácido Valproico/toxicidade , Animais , Células Cultivadas , Cerebelo/citologia , Cerebelo/efeitos dos fármacos , Cerebelo/metabolismo , Exocitose/efeitos dos fármacos , Regulação da Expressão Gênica , Transtornos do Neurodesenvolvimento/induzido quimicamente , Transtornos do Neurodesenvolvimento/genética , Neurônios/citologia , Neurônios/efeitos dos fármacos , Neurônios/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Ratos , Ratos Wistar , Proteína 25 Associada a Sinaptossoma/genética , Testes de Toxicidade
8.
Toxicol Sci ; 154(2): 227-240, 2016 12.
Artigo em Inglês | MEDLINE | ID: mdl-27660204

RESUMO

Ethylmercury thiosalicylate (thimerosal) is an organic mercury-based compound commonly used as an antimicrobial preservative that has been found to be neurotoxic. In contrast, histone deacetylases (HDACs) inhibition has been found to be neuroprotective against several environmental contaminants, such as polychlorinated biphenyls, di-2-ethylhexyl phthalate, and methylmercury. The aim of this study was to investigate the effect of HDAC inhibition on thimerosal-induced neurotoxicity in neuroblastoma cells and cortical neurons. Interestingly, we found that thimerosal, at 0.5 µM in SH-SY5Y cells and at 1 µM in neurons, caused cell death by activation of apoptosis, which was prevented by the HDAC class IIA inhibitor MC1568 but not the class I inhibitor MS275. Furthermore, thimerosal specifically increased HDAC4 protein expression but not that of HDACs 5, 6, 7, and 9. Western blot analysis revealed that MC1568 prevented thimerosal-induced HDAC4 increase. In addition, both HDAC4 knocking-down and MC1568 inhibited thimerosal-induced cell death in SH-SY5Y cells and cortical neurons. Importantly, intramuscular injection of 12 µg/kg thimerosal on postnatal days 7, 9, 11, and 15 increased HDAC4 levels in the prefrontal cortex (PFC), which decreased histone H4 acetylation in infant male rats, in parallel increased motor activity changes. In addition, coadministration of 40 mg/kg MC1568 (intraperitoneal injection) moderated the HDAC4 increase which reduced histone H4 deacetylation and caspase-3 cleavage in the PFC. Finally, open-field testing showed that thimerosal-induced motor activity changes are reduced by MC1568. These findings indicate that HDAC4 regulates thimerosal-induced cell death in neurons and that treatment with MC1568 prevents thimerosal-induced activation of caspase-3 in the rat PFC.


Assuntos
Apoptose/efeitos dos fármacos , Inibidores de Histona Desacetilases/farmacologia , Histona Desacetilases/metabolismo , Ácidos Hidroxâmicos/farmacologia , Neurônios/efeitos dos fármacos , Fármacos Neuroprotetores/farmacologia , Córtex Pré-Frontal/efeitos dos fármacos , Pirróis/farmacologia , Proteínas Repressoras/antagonistas & inibidores , Timerosal/toxicidade , Animais , Comportamento Animal/efeitos dos fármacos , Caspase 3/metabolismo , Linhagem Celular Tumoral , Citoproteção , Relação Dose-Resposta a Droga , Histona Desacetilases/genética , Humanos , Masculino , Atividade Motora/efeitos dos fármacos , Neurônios/enzimologia , Neurônios/patologia , Córtex Pré-Frontal/enzimologia , Córtex Pré-Frontal/patologia , Córtex Pré-Frontal/fisiopatologia , Interferência de RNA , Ratos Wistar , Proteínas Repressoras/genética , Proteínas Repressoras/metabolismo , Proteína SUMO-1/genética , Proteína SUMO-1/metabolismo , Transdução de Sinais/efeitos dos fármacos , Transfecção , Regulação para Cima
9.
J Toxicol Environ Health A ; 79(12): 502-12, 2016.
Artigo em Inglês | MEDLINE | ID: mdl-27294299

RESUMO

Exposure to organomercurials has been associated with harmful effects on the central nervous system (CNS). However, the mechanisms underlying organomercurial-mediated neurotoxic effects need to be elucidated. Exposure to toxic elements may promote cellular modifications such as alterations in protein synthesis in an attempt to protect tissues and organs from damage. In this context, the use of a "proteomic profile" is an important tool to identify potential early biomarkers or targets indicative of neurotoxicity. The aim of this study was to investigate potential modifications in rat cerebral cell proteome following exposure to methylmercury (MeHg) or ethylmercury (EtHg). For MeHg exposure, animals were administered by gavage daily 140 µg/kg/d of Hg (as MeHg) for 60 d and sacrificed 24 h after the last treatment. For EtHg exposure, 800 µg/kg/d of Hg (as EtHg) was given intramuscularly (im) in a single dose and rats were sacrificed after 4 h. Control groups received saline either by gavage or im. After extraction of proteins from whole brain samples and separation by two-dimensional electrophoresis (2-DE), 26 differentially expressed proteins were identified from exposed animals by matrix-assisted laser desorption ionization-time of flight (MALDI-TOF/TOF). Both MeHg and EtHg exposure induced an overexpression of calbindin, a protein that acts as a neuroprotective agent by (1) adjusting the concentration of Ca(2+) within cells and preventing neurodegenerative diseases and (2) decreasing expression of glutamine synthetase, a crucial protein involved in regulation of glutamate concentration in synaptic cleft. In contrast, expression of superoxide dismutase (SOD), a protein involved in antioxidant defense, was elevated in brain of MeHg-exposed animals. Taken together, our data provide new valuable information on the possible molecular mechanisms associated with MeHg- and EtHg-mediated toxicity in cerebral tissue. These observed protein alterations may be considered as biomarkers candidates for biological monitoring of organomercurial poisoning.


Assuntos
Encéfalo/efeitos dos fármacos , Compostos de Metilmercúrio/toxicidade , Proteoma/efeitos dos fármacos , Timerosal/toxicidade , Animais , Poluentes Ambientais/toxicidade , Masculino , Distribuição Aleatória , Ratos , Ratos Wistar , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz
10.
Toxicology ; 347-349: 1-5, 2016 Mar 10.
Artigo em Inglês | MEDLINE | ID: mdl-26945727

RESUMO

A latency period preceding neurotoxicity is a common characteristic in the dose-response relationship induced by organic mercury. Latency periods have typically been observed with genotoxicants in carcinogenesis, with cancer being manifested a long time after the initiating event. These observations indicate that even a very small dose may cause extensive adverse effects later in life, so the toxicity of the genotoxic compound is dose and time-dependent. In children, methylmercury exposure during pregnancy (in utero) has been associated with delays in reaching developmental milestones (e.g., age at first walking) and decreases in intelligence, increasing in severity with increasing exposure. Ethylmercury exposure from thimerosal in some vaccines has been associated, in some studies, with autism and other neurological disorders in children. In this paper, we have examined whether dose-response data from in vitro and in vivo organic mercury toxicity studies fit the Druckrey-Küpfmüller equation c·t(n)=constant (c=exposure concentration, t=latency period), first established for genotoxic carcinogens, and whether or not irreversible effects are enhanced by time of exposure (n≥1), or else toxic effects are dose-dependent while time has only minor influence on the adverse outcome (n<1). The mode of action underlying time-dependent toxicity is irreversible binding to critical receptors causing adverse and cumulative effects. The results indicate that the Druckrey-Küpfmüller equation describes well the dose-response characteristics of organic mercury induced neurotoxic effects. This amounts to a paradigm shift in chemical risk assessment of mercurial compounds and highlights that it is vital to perform toxicity testing geared to investigate time-dependent effects.


Assuntos
Encéfalo/efeitos dos fármacos , Encéfalo/crescimento & desenvolvimento , Compostos de Metilmercúrio/toxicidade , Síndromes Neurotóxicas , Animais , Encéfalo/metabolismo , Relação Dose-Resposta a Droga , Feminino , Humanos , Mercúrio/administração & dosagem , Mercúrio/metabolismo , Mercúrio/toxicidade , Compostos de Metilmercúrio/administração & dosagem , Compostos de Metilmercúrio/metabolismo , Síndromes Neurotóxicas/metabolismo , Síndromes Neurotóxicas/patologia , Gravidez , Timerosal/administração & dosagem , Timerosal/metabolismo , Timerosal/toxicidade , Testes de Toxicidade/métodos , Testes de Toxicidade/normas
11.
Arch Toxicol ; 90(3): 543-50, 2016 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-25701957

RESUMO

Humans are exposed to different mercurial compounds from various sources, most frequently from dental fillings, preservatives in vaccines, or consumption of fish. Among other toxic effects, these substances interact with the immune system. In high doses, mercurials are immunosuppressive. However, lower doses of some mercurials stimulate the immune system, inducing different forms of autoimmunity, autoantibodies, and glomerulonephritis in rodents. Furthermore, some studies suggest a connection between mercury exposure and the occurrence of autoantibodies against nuclear components and granulocyte cytoplasmic proteins in humans. Still, the underlying mechanisms need to be clarified. The present study investigates the formation of neutrophil extracellular traps (NETs) in response to thimerosal and its metabolites ethyl mercury (EtHg), thiosalicylic acid, and mercuric ions (Hg(2+)). Only EtHg and Hg(2+) triggered NETosis. It was independent of PKC, ERK1/2, p38, and zinc signals and not affected by the NADPH oxidase inhibitor DPI. Instead, EtHg and Hg(2+) triggered NADPH oxidase-independent production of ROS, which are likely to be involved in mercurial-induced NET formation. This finding might help understanding the autoimmune potential of mercurial compounds. Some diseases, to which a connection with mercurials has been shown, such as Wegener's granulomatosis and systemic lupus erythematosus, are characterized by high prevalence of autoantibodies against neutrophil-specific auto-antigens. Externalization in the form of NETs may be a source for exposure to these self-antigens. In genetically susceptible individuals, this could be one step in the series of events leading to autoimmunity.


Assuntos
Compostos de Etilmercúrio/toxicidade , Armadilhas Extracelulares/efeitos dos fármacos , Mercúrio/toxicidade , Neutrófilos/efeitos dos fármacos , Células Cultivadas , Granulócitos/efeitos dos fármacos , Humanos , Leucócitos/efeitos dos fármacos , NADPH Oxidases/metabolismo , Neutrófilos/metabolismo , Neutrófilos/patologia , Fosforilação/efeitos dos fármacos , Proteínas Quinases/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Salicilatos/toxicidade , Compostos de Sulfidrila/toxicidade , Timerosal/toxicidade , Zinco/metabolismo
12.
J Trace Elem Med Biol ; 32: 200-8, 2015 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-26302930

RESUMO

Organic mercury (Hg) species exert their toxicity primarily in the central nervous system. The food relevant Hg species methylmercury (MeHg) has been frequently studied regarding its neurotoxic effects in vitro and in vivo. Neurotoxicity of thiomersal, which is used as a preservative in medical preparations, is to date less characterised. Due to dealkylation of organic Hg or oxidation of elemental Hg, inorganic Hg is present in the brain albeit these species are not able to readily cross the blood brain barrier. This study compared for the first time toxic effects of organic MeHg chloride (MeHgCl) and thiomersal as well as inorganic mercury chloride (HgCl2) in differentiated human neurons (LUHMES) and human astrocytes (CCF-STTG1). The three Hg species differ in their degree and mechanism of toxicity in those two types of brain cells. Generally, neurons are more susceptible to Hg species induced cytotoxicity as compared to astrocytes. This might be due to the massive cellular mercury uptake in the differentiated neurons. The organic compounds exerted stronger cytotoxic effects as compared to inorganic HgCl2. In contrast to HgCl2 exposure, organic Hg compounds seem to induce the apoptotic cascade in neurons following low-level exposure. No indicators for apoptosis were identified for both inorganic and organic mercury species in astrocytes. Our studies clearly demonstrate species-specific toxic mechanisms. A mixed exposure towards all Hg species in the brain can be assumed. Thus, prospectively coexposure studies as well as cocultures of neurons and astrocytes could provide additional information in the investigation of Hg induced neurotoxicity.


Assuntos
Astrócitos/citologia , Diferenciação Celular/efeitos dos fármacos , Cloreto de Mercúrio/toxicidade , Compostos de Metilmercúrio/toxicidade , Neurônios/citologia , Timerosal/toxicidade , Apoptose/efeitos dos fármacos , Astrócitos/efeitos dos fármacos , Disponibilidade Biológica , Caspase 3/metabolismo , Morte Celular/efeitos dos fármacos , Células Cultivadas , Ativação Enzimática/efeitos dos fármacos , Humanos , Lisossomos/efeitos dos fármacos , Lisossomos/metabolismo , Microscopia de Fluorescência , Neurônios/efeitos dos fármacos
13.
Toxicol Appl Pharmacol ; 286(3): 216-23, 2015 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-25981166

RESUMO

Mercury (Hg) is a strong toxicant affecting mainly the central nervous, renal, cardiovascular and immune systems. Thiomersal (TM) is still in use in medical practice as a topical antiseptic and as a preservative in multiple dose vaccines, routinely given to young children in some developing countries, while other forms of mercury such as methylmercury represent an environmental and food hazard. The aim of the present study was to determine the effects of thiomersal (TM) and its breakdown product ethylmercury (EtHg) on the thioredoxin system and NADP(+)-dependent dehydrogenases of the pentose phosphate pathway. Results show that TM and EtHg inhibited the thioredoxin system enzymes in purified suspensions, being EtHg comparable to methylmercury (MeHg). Also, treatment of neuroblastoma and liver cells with TM or EtHg decreased cell viability (GI50: 1.5 to 20µM) and caused a significant (p<0.05) decrease in the overall activities of thioredoxin (Trx) and thioredoxin reductase (TrxR) in a concentration- and time-dependent manner in cell lysates. Compared to control, the activities of Trx and TrxR in neuroblastoma cells after EtHg incubation were reduced up to 60% and 80% respectively, whereas in hepatoma cells the reduction was almost 100%. In addition, the activities of glucose-6-phosphate dehydrogenase and 6-phosphogluconate dehydrogenase were also significantly inhibited by all mercurials, with inhibition intensity of Hg(2+)>MeHg≈EtHg>TM (p<0.05). Cell incubation with sodium selenite alleviated the inhibitory effects on TrxR and glucose-6-phosphate dehydrogenase. Thus, the molecular mechanism of toxicity of TM and especially of its metabolite EtHg encompasses the blockage of the electrons from NADPH via the thioredoxin system.


Assuntos
Compostos de Etilmercúrio/toxicidade , NADPH Desidrogenase/antagonistas & inibidores , Via de Pentose Fosfato/efeitos dos fármacos , Timerosal/toxicidade , Tiorredoxinas/antagonistas & inibidores , Sobrevivência Celular , Relação Dose-Resposta a Droga , Células Hep G2 , Humanos , NADPH Desidrogenase/metabolismo , Via de Pentose Fosfato/fisiologia , Tiorredoxinas/metabolismo
14.
Environ Toxicol ; 30(12): 1423-33, 2015 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-24942245

RESUMO

Thimerosal is an ethyl mercury-containing compound used mainly in vaccines as a bactericide. Although the kidney is a key target for mercury toxicity, thimerosal nephrotoxicity has not received the same attention as other mercury species. The aim of this study was to determine the potential cytotoxic mechanisms of thimerosal on human kidney cells. Human kidney proximal tubular epithelial (HK2) cells were exposed for 24 h to thimerosal (0-2 µM), and assessed for cell viability, apoptosis, and cell proliferation; expression of proteins Bax, nuclear factor-κB subunits, and transforming growth factor-ß1 (TGFß1); mitochondrial health (JC-1, MitoTracker Red CMXRos); and fibronectin levels (enzyme-linked immunosorbent assay). Thimerosal diminished HK2 cell viability and mitosis, promoted apoptosis, impaired the mitochondrial permeability transition, enhanced Bax and TGFß1 expression, and augmented fibronectin secretion. This is the first report about kidney cell death and pro-fibrotic mechanisms promoted by thimerosal. Collectively, these in vitro results demonstrate that (1) thimerosal induces kidney epithelial cell apoptosis via upregulating Bax and the mitochondrial apoptotic pathway, and (2) thimerosal is a potential pro-fibrotic agent in human kidney cells. We suggest that new evidence on toxicity as well as continuous surveillance in terms of fibrogenesis is required concerning thimerosal use.


Assuntos
Apoptose/efeitos dos fármacos , Timerosal/toxicidade , Proteínas Reguladoras de Apoptose/metabolismo , Linhagem Celular , Proliferação de Células/efeitos dos fármacos , Ensaio de Imunoadsorção Enzimática , Células Epiteliais/citologia , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/metabolismo , Fibronectinas/análise , Fibrose , Humanos , Túbulos Renais/citologia , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Mitose/efeitos dos fármacos , NF-kappa B/metabolismo , Fator de Crescimento Transformador beta1/metabolismo , Proteína X Associada a bcl-2/metabolismo
15.
Toxicol Sci ; 144(1): 105-13, 2015 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-25490951

RESUMO

The effect of mercury compounds has been tested on the organic cation transporter, hOCTN1. MeHg(+), Hg(2+), or Cd(2+) caused strong inhibition of transport. 1,4-Dithioerythritol (DTE), cysteine (Cys), and N-acetyl-l-cysteine reversed (NAC) the inhibition at different extents. 2-Aminoethyl methanethiosulfonate hydrobromide (MTSEA), a prototype SH reagent, exerted inhibition of transport similar to that observed for the mercurial agents. To investigate the mechanism of action of mercurials, mutants of hOCTN1 in which each of the Cys residues was substituted by Ala have been constructed, over-expressed in Escherichia coli, and purified. Tetraethylammonium chloride (TEA) uptake mediated by each mutant in proteoliposomes was comparable to that of wild type (WT). IC50 values of the WT and mutants for the mercury compounds were derived from dose-response analyses. The mutants C50A and C136A showed significant increase of IC50 indicating that the 2 Cys residues were involved in the interaction with the mercury compounds and inhibition of the transporter. The double mutant C50A/C136A was constructed; the lack of inhibition confirmed that the 2 Cys residues are the targets of mercury compounds. MTSEA showed similar behavior with respect to the mercurial reagents with the difference that increased IC50 was observed also in the C81A mutant. Similar results were obtained when transport was measured as acetylcholine uptake. Ethyl mercury (Thimerosal) inhibited hOCTN1 as well. C50A, C50A/C136A and, at very lower extent, C136A showed increased IC50 indicating that C50 was the major target of this mercury compound. The homology model of hOCTN1 was built using as template PiPT and validated by the experimental data on mutant proteins.


Assuntos
Cloreto de Mercúrio/toxicidade , Compostos de Metilmercúrio/toxicidade , Proteínas de Transporte de Cátions Orgânicos/antagonistas & inibidores , Timerosal/toxicidade , Acetilcolina/metabolismo , Transporte Biológico , Cloreto de Cádmio/toxicidade , Cisteína , Relação Dose-Resposta a Droga , Genótipo , Humanos , Cinética , Modelos Moleculares , Mutação , Proteínas de Transporte de Cátions Orgânicos/química , Proteínas de Transporte de Cátions Orgânicos/genética , Proteínas de Transporte de Cátions Orgânicos/metabolismo , Fenótipo , Conformação Proteica , Proteínas Recombinantes/efeitos dos fármacos , Proteínas Recombinantes/metabolismo , Relação Estrutura-Atividade , Simportadores , Tetraetilamônio/metabolismo
16.
Environ Res ; 134: 218-27, 2014 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-25173055

RESUMO

Thimerosal (TM) is an ethylmercury (etHg)-containing preservative used in some vaccines despite very limited knowledge on the kinetics and direct interaction/effects in mammals׳ tissues after exposure. Thus, this study aimed to evaluate the kinetics of Hg species in mice in a time course analysis after intramuscular injection of TM, by estimating Hg half-lives in blood and tissues. Mice were exposed to one single intramuscular dose of 20 µg of Hg as TM. Blood, brain, heart, kidney and liver were collected at 0.5 hour (h), 1 h, 8 h, 16 h, 144 h, 720 h and 1980 h after TM exposure (n=4). Hg species in animal tissues were identified and quantified by speciation analysis via liquid chromatography hyphenated with inductively coupled mass spectrometry (LC-ICP-MS). It was found that the transport of etHg from muscle to tissues and its conversion to inorganic Hg (inoHg) occur rapidly. Moreover, the conversion extent is modulated in part by the partitioning between EtHg in plasma and in whole blood, since etHg is rapidly converted in red cells but not in a plasma compartment. Furthermore, the dealkylation mechanism in red cells appears to be mediated by the Fenton reaction (hydroxyl radical formation). Interestingly, after 0.5 h of TM exposure, the highest levels of both etHg and inoHg were found in kidneys (accounting for more than 70% of the total Hg in the animal body), whereas the brain contributed least to the Hg body burden (accounts for <1.0% of total body Hg). Thirty days after TM exposure, most Hg had been excreted while the liver presented the majority of the remaining Hg. Estimated half-lives (in days) were 8.8 for blood, 10.7 for brain, 7.8 for heart, 7.7 for liver and 45.2 for kidney. Taken together, our findings demonstrated that TM (etHg) kinetics more closely approximates Hg(2+) than methylmercury (meHg) while the kidney must be considered a potential target for etHg toxicity.


Assuntos
Mercúrio/farmacocinética , Timerosal/toxicidade , Animais , Cromatografia Líquida , Meia-Vida , Espectrometria de Massas , Mercúrio/sangue , Camundongos , Distribuição Tecidual
17.
PLoS One ; 9(4): e92705, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-24690681

RESUMO

Thimerosal is a preservative used widely in vaccine formulations to prevent bacterial and fungal contamination in multidose vials of vaccine. Thimerosal was included in the multidose non-adjuvanted pandemic 2009 H1N1 vaccine Panenza. In the context of the analysis of the ex-vivo T cell responses directed against influenza vaccine, we discovered the in vitro toxicity Panenza, due to its content in thimerosal. Because thimerosal may skew the immune response to vaccines, we investigated in detail the ex-vivo effects of thimerosal on the fate and functions of T cells in response to TCR ligation. We report that ex-vivo exposure of quiescent or TCR-activated primary human T cells to thimerosal induced a dose-dependent apoptotic cell death associated with depolarization of mitochondrial membrane, generation of reactive oxygen species, cytochrome c release from the mitochondria and caspase-3 activation. Moreover, exposure to non-toxic concentrations of thimerosal induced cell cycle arrest in G0/G1 phase of TCR-activated T cells, and inhibition of the release of proinflammatory cytokines such as IFN gamma, IL-1 beta, TNF alpha, IL-2, as well as the chemokine MCP1. No shift towards Th2 or Th17 cells was detected. Overall these results underline the proapoptotic effect of thimerosal on primary human lymphocytes at concentrations 100 times less to those contained in the multidose vaccine, and they reveal the inhibitory effect of this preservative on T-cell proliferation and functions at nanomolar concentrations.


Assuntos
Apoptose/efeitos dos fármacos , Linfócitos T CD4-Positivos/imunologia , Memória Imunológica/efeitos dos fármacos , Vacinas contra Influenza/imunologia , Timerosal/toxicidade , Adulto , Caspase 3/metabolismo , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Quimiocinas/metabolismo , Citocromos c/metabolismo , Relação Dose-Resposta Imunológica , Ativação Enzimática/efeitos dos fármacos , Humanos , Vírus da Influenza A Subtipo H1N1/efeitos dos fármacos , Vírus da Influenza A Subtipo H1N1/imunologia , Ativação Linfocitária/efeitos dos fármacos , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Proteínas de Transporte da Membrana Mitocondrial/efeitos dos fármacos , Proteínas de Transporte da Membrana Mitocondrial/metabolismo , Poro de Transição de Permeabilidade Mitocondrial , Espécies Reativas de Oxigênio/metabolismo , Receptores de Antígenos de Linfócitos T/metabolismo
18.
Toxicol Sci ; 139(2): 452-65, 2014 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-24675092

RESUMO

Thimerosal is a vaccine antimicrobial preservative which has long been suspected an iatrogenic factor possibly contributing to neurodevelopmental disorders including autism. The association between infant vaccine thimerosal exposure and autism remains an open question. Although thimerosal has been removed from mandatory childhood vaccines in the United States, thimerosal-preserved vaccines are still widely used outside of the United States especially in developing countries. Notably, thimerosal-containing vaccines are being given to the newborns within the first 12-24 h after birth in some countries. To examine the possible neurotoxic effects of early neonatal exposure to a higher level of thimerosal, FVB mice were subcutaneously injected with thimerosal-mercury at a dose which is 20× higher than that used for regular Chinese infant immunization during the first 4 months of life. Thimerosal-treated mice exhibited neural development delay, social interaction deficiency, and inclination of depression. Apparent neuropathological changes were also observed in adult mice neonatally treated with thimerosal. High-throughput RNA sequencing of autistic-behaved mice brains revealed the alternation of a number of canonical pathways involving neuronal development, neuronal synaptic function, and the dysregulation of endocrine system. Intriguingly, the elevation of anterior pituitary secreting hormones occurred exclusively in male but not in female thimerosal-treated mice, demonstrating for the first time the gender bias of thimerosal-mercury toxicity with regard to endocrine system. Our results indicate that higher dose of neonatal thimerosal-mercury (20× higher than that used in human) is capable of inducing long-lasting substantial dysregulation of neurodevelopment, synaptic function, and endocrine system, which could be the causal involvements of autistic-like behavior in mice.


Assuntos
Comportamento Animal/efeitos dos fármacos , Encéfalo/efeitos dos fármacos , Síndromes Neurotóxicas/etiologia , Conservantes Farmacêuticos/toxicidade , Timerosal/toxicidade , Transcriptoma/efeitos dos fármacos , Animais , Animais Recém-Nascidos , Encéfalo/crescimento & desenvolvimento , Encéfalo/patologia , Relação Dose-Resposta a Droga , Feminino , Perfilação da Expressão Gênica , Masculino , Camundongos Endogâmicos , Síndromes Neurotóxicas/patologia , Síndromes Neurotóxicas/psicologia , Conservantes Farmacêuticos/administração & dosagem , Caracteres Sexuais , Timerosal/administração & dosagem
19.
Metallomics ; 6(3): 662-71, 2014 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-24549367

RESUMO

The toxicologically most relevant mercury (Hg) species for human exposure is methylmercury (MeHg). Thiomersal is a common preservative used in some vaccine formulations. The aim of this study is to get further mechanistic insight into the yet not fully understood neurotoxic modes of action of organic Hg species. Mercury species investigated include MeHgCl and thiomersal. Additionally HgCl2 was studied, since in the brain mercuric Hg can be formed by dealkylation of the organic species. As a cellular system astrocytes were used. In vivo astrocytes provide the environment necessary for neuronal function. In the present study, cytotoxic effects of the respective mercuricals increased with rising alkylation level and correlated with their cellular bioavailability. Further experiments revealed for all species at subcytotoxic concentrations no induction of DNA strand breaks, whereas all species massively increased H2O2-induced DNA strand breaks. This co-genotoxic effect is likely due to a disturbance of the cellular DNA damage response. Thus, at nanomolar, sub-cytotoxic concentrations, all three mercury species strongly disturbed poly(ADP-ribosyl)ation, a signalling reaction induced by DNA strand breaks. Interestingly, the molecular mechanism behind this inhibition seems to be different for the species. Since chronic PARP-1 inhibition is also discussed to sacrifice neurogenesis and learning abilities, further experiments on neurons and in vivo studies could be helpful to clarify whether the inhibition of poly(ADP-ribosyl)ation contributes to organic Hg induced neurotoxicity.


Assuntos
Astrócitos/efeitos dos fármacos , Compostos de Metilmercúrio/toxicidade , Mutagênicos/toxicidade , Timerosal/toxicidade , Astrócitos/metabolismo , Linhagem Celular , Quebras de DNA/efeitos dos fármacos , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Peróxido de Hidrogênio/metabolismo , Poli(ADP-Ribose) Polimerase-1 , Poli Adenosina Difosfato Ribose/análise , Poli Adenosina Difosfato Ribose/metabolismo , Poli(ADP-Ribose) Polimerases/metabolismo
20.
Graefes Arch Clin Exp Ophthalmol ; 252(2): 275-84, 2014 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-24384799

RESUMO

BACKGROUND: Previously we have shown that acute exposure to thimerosal (Thi) can induce oxidative stress and DNA damage in a human conjunctival cell line. However, the long-term effect of Thi on Chang conjunctival cells is not clear. Therefore, the aim of this study was to further investigate the fate of the cells after acute exposure to Thi. METHOD: Cells were first exposed to various concentrations of Thi (0.00001 % ∼ 0.001 %) for 30 min, and then cells were assessed after a 24-h recovery period. Morphologic changes were observed under a light microscope and cell viability was evaluated. Cell apoptosis, cell cycle distribution and mitochondrial membrane potential (MMP) (rhodamine 123 assay) were detected by flow cytometry analysis. Poly (ADP-ribose) polymerase (PARP), activation of caspase-3 and microtubule-associated protein light chain 3 (LC-3) were examined by western blot analysis. RESULTS: DNA strand breaks were significantly increased in a dose-dependent manner with 30 min exposure to Thi, although no significant cell death was detected. However, after 24-h recovery, the ratio of apoptotic cells was significantly increased to 0.0005 % and 0.001 % in Thi treated groups (p < 0.001 compared to the control group). Apoptosis was confirmed by the cleavage of PARP and caspase-3 activation. In addition, G2/M cell cycle arrest and decrease of MMP were recorded. Finally, the LC-3 results indicated the occurrence of autophagy in Thi-treated cells. CONCLUSION: Acute exposure to Thi can induce DNA damage, and eventually can lead to cell death, probably through the caspase-dependent apoptosis pathway, while autophagy might also be involved.


Assuntos
Apoptose/efeitos dos fármacos , Autofagia/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Túnica Conjuntiva/patologia , Conservantes Farmacêuticos/toxicidade , Timerosal/toxicidade , Western Blotting , Caspase 3/metabolismo , Linhagem Celular , Túnica Conjuntiva/metabolismo , Dano ao DNA/efeitos dos fármacos , Células Epiteliais/metabolismo , Células Epiteliais/patologia , Citometria de Fluxo , Humanos , Potencial da Membrana Mitocondrial , Proteínas Associadas aos Microtúbulos/metabolismo , Poli(ADP-Ribose) Polimerases/metabolismo
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