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1.
Nat Commun ; 12(1): 5293, 2021 09 06.
Artigo em Inglês | MEDLINE | ID: mdl-34489448

RESUMO

The ATP hydrolysis transition state of motor proteins is a weakly populated protein state that can be stabilized and investigated by replacing ATP with chemical mimics. We present atomic-level structural and dynamic insights on a state created by ADP aluminum fluoride binding to the bacterial DnaB helicase from Helicobacter pylori. We determined the positioning of the metal ion cofactor within the active site using electron paramagnetic resonance, and identified the protein protons coordinating to the phosphate groups of ADP and DNA using proton-detected 31P,1H solid-state nuclear magnetic resonance spectroscopy at fast magic-angle spinning > 100 kHz, as well as temperature-dependent proton chemical-shift values to prove their engagements in hydrogen bonds. 19F and 27Al MAS NMR spectra reveal a highly mobile, fast-rotating aluminum fluoride unit pointing to the capture of a late ATP hydrolysis transition state in which the phosphoryl unit is already detached from the arginine and lysine fingers.


Assuntos
Difosfato de Adenosina/química , Trifosfato de Adenosina/química , Proteínas de Bactérias/química , DNA Bacteriano/química , DnaB Helicases/química , Helicobacter pylori/enzimologia , Difosfato de Adenosina/metabolismo , Trifosfato de Adenosina/metabolismo , Compostos de Alumínio/química , Compostos de Alumínio/metabolismo , Arginina/química , Arginina/metabolismo , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Domínio Catalítico , Clonagem Molecular , DNA Bacteriano/genética , DNA Bacteriano/metabolismo , DnaB Helicases/genética , DnaB Helicases/metabolismo , Escherichia coli/enzimologia , Escherichia coli/genética , Fluoretos/química , Fluoretos/metabolismo , Expressão Gênica , Helicobacter pylori/genética , Hidrólise , Lisina/química , Lisina/metabolismo , Modelos Moleculares , Ressonância Magnética Nuclear Biomolecular , Ligação Proteica , Conformação Proteica em alfa-Hélice , Conformação Proteica em Folha beta , Domínios e Motivos de Interação entre Proteínas , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Especificidade por Substrato , Termodinâmica
2.
Eur J Pharm Sci ; 166: 105985, 2021 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-34455087

RESUMO

Non-ideal behaviour of mixed ions is disclosed in skin absorption experiments of mixed halide anions in excised pig skin. Comparison of skin absorption of pure and mixed ions shows enhanced penetration of chaotropic ions from mixed solutions. An experimental design and statistical analysis using a Scheffé {3,2} simplex-lattice allows investigating the full ternary diagram of anion mixtures of fluoride, bromide and iodide. Synergism in mixed absorption is observed for chaotropic bromide and iodide anions. A refined analysis highlighting specific interactions is made by considering the ratio of the absorbed amount to the ion activity instead of the directly measured absorbed amount. Statistical analysis discards non-significant effects and discloses specific interactions. Such interactions between bromide and iodide cause an absorption enhancement of their partner by a factor of 2-3 with respect to the case of ideal mixing. It is proposed that enhanced absorption from mixed solution involves the formation of neutral complex species of mixed bromide and iodide with endogenous magnesium or calcium inside stratum corneum.


Assuntos
Absorção Cutânea , Água , Animais , Ânions/metabolismo , Fluoretos/metabolismo , Pele/metabolismo , Soluções , Suínos , Água/metabolismo
3.
Elife ; 102021 07 12.
Artigo em Inglês | MEDLINE | ID: mdl-34250906

RESUMO

Fluc family fluoride channels protect microbes against ambient environmental fluoride by undermining the cytoplasmic accumulation of this toxic halide. These proteins are structurally idiosyncratic, and thus the permeation pathway and mechanism have no analogy in other known ion channels. Although fluoride-binding sites were identified in previous structural studies, it was not evident how these ions access aqueous solution, and the molecular determinants of anion recognition and selectivity have not been elucidated. Using x-ray crystallography, planar bilayer electrophysiology, and liposome-based assays, we identified additional binding sites along the permeation pathway. We used this information to develop an oriented system for planar lipid bilayer electrophysiology and observed anion block at one of these sites, revealing insights into the mechanism of anion recognition. We propose a permeation mechanism involving alternating occupancy of anion-binding sites that are fully assembled only as the substrate approaches.


Assuntos
Ânions/metabolismo , Fluoretos/metabolismo , Canais Iônicos/metabolismo , Ânions/química , Proteínas de Bactérias/metabolismo , Sítios de Ligação , Fenômenos Biofísicos , Cristalografia por Raios X/métodos , Eletrofisiologia/métodos , Fluoretos/química , Ácido Glutâmico/metabolismo , Canais Iônicos/química , Modelos Moleculares , Conformação Proteica
4.
Plant Cell Rep ; 40(9): 1751-1772, 2021 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-34173048

RESUMO

KEY MESSAGE: Early induction of OsFEX was insufficient for fluoride adaptation in IR-64. Overexpression of OsFEX in yeast and Nicotiana benthamiana enhanced fluoride tolerance. The present study delineates the regulation of fluoride exporter (FEX) in the fluoride-sensitive rice cultivar, IR-64 and its efficacy in generating high fluoride tolerance in transgenic Nicotiana benthamiana. Gene and protein expression profiling revealed that OsFEX exhibited early induction during fluoride stress in the vegetative and reproductive tissues of IR-64, although the expression was suppressed upon prolonged stress treatment. Analysis of OsFEX promoter in transgenic N. benthamiana, using ß-glucuronidase reporter assay confirmed its early inducible nature, since the reporter expression and activity peaked at 12 h of NaF stress, after which it was lowered. OsFEX expression was up regulated in the presence of gibberellic acid (GA) and melatonin, while it was suppressed by abscisic acid (ABA). Complementation of ΔFEX1ΔFEX2 yeast mutants with OsFEX enabled high fluoride tolerance, thus validating the functional efficiency of the transgene. Bioassay of transgenic N. benthamiana lines, expressing OsFEX either under its own promoter or under CaMV35S promoter, established that constitutive overexpression, rather than early induction of OsFEX was essential and crucial for generating fluoride tolerance in the transgenics. Overall, the suppression of OsFEX in the later growth phases of stressed IR-64 due to enhanced ABA conservation and lowered synthesis of GA, as supported by the application of the respective phytohormone biosynthetic inhibitors, such as sodium tungstate and paclobutrazol, accounted for the fluoride-hyperaccumulative nature of the rice cultivar.


Assuntos
Fluoretos/toxicidade , Oryza/efeitos dos fármacos , Oryza/fisiologia , Proteínas de Plantas/genética , Tabaco/genética , Ácido Abscísico/farmacologia , Adaptação Fisiológica/efeitos dos fármacos , Adaptação Fisiológica/genética , Proteínas de Transporte/genética , Proteínas de Transporte/metabolismo , Fluoretos/metabolismo , Regulação da Expressão Gênica de Plantas , Teste de Complementação Genética , Giberelinas/farmacologia , Proteínas de Membrana/genética , Microrganismos Geneticamente Modificados , Mutação , Proteínas de Plantas/metabolismo , Plantas Geneticamente Modificadas , Regiões Promotoras Genéticas , Saccharomyces cerevisiae/efeitos dos fármacos , Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/genética , Estresse Fisiológico/efeitos dos fármacos , Estresse Fisiológico/fisiologia
5.
Pol J Vet Sci ; 24(1): 151-157, 2021 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-33847101

RESUMO

The last biomonitoring study in Poland on intoxication with fluoride compounds of deer was conducted almost two decades ago. Given the fact that fluoride level in air and water is not widely monitored in Poland, it is justified to undertake monitoring of F- levels in people and other long-lived mammals. This paper provides the assessment of the present level of fluoride accumulation in mineralized tissue of large herbivorous mammals. The aim of the present study was to determine fluoride concentration in teeth of deer inhabiting the areas of Poland which are industrially uncontaminated with fluoride compounds, to establish possible correlations between the analysed parameters, and to provide a comparison of the present results with those obtained in other studies. Mean concentration of fluoride in all analysed samples amounted to 231.0 F mg/kg, with the minimum value of 22.0 F mg/kg and the maximum of 935.0 F mg/kg. This results from the development of industry and a widespread use of fluoride-supplemented caries prevention products which contributes to an intense accumulation of these substances in vertebrates, predominantly in mineralized tissue.


Assuntos
Cervos/metabolismo , Poluentes Ambientais/química , Fluoretos/química , Resíduos Industriais , Dente/química , Animais , Cervos/classificação , Monitoramento Ambiental , Poluentes Ambientais/metabolismo , Fluoretos/metabolismo , Polônia , Dente/metabolismo
6.
Ecotoxicol Environ Saf ; 215: 112108, 2021 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-33799132

RESUMO

Fluoride which is widespread in our environment and food due to its geological origin and industrial pollution has been identified as a developmental neurotoxicant. Gut-brain axis provides new insight into brain-derived injury. We previously found the psychoactive effects of a probiotic strain, Lactobacillus johnsonii BS15 against fluoride-induced memory dysfunction in mice by modulating the gut-brain axis. In this study, we aimed to detect the link between the reconstruction of gut microbiota and gut-brain axis through which probiotic alleviate fluoride-induced memory impairment. We also added an hour of water avoidance stress (WAS) before behavioral tests and sampling, aiming to demonstrate the preventive effects of the probiotic on fluoride-induced memory impairment after psychological stress. Mice were given fluoridated drinking water (sodium fluoride 100 ppm, corresponding to 37.8 ± 2.4 ppm F¯) for 70 days and administered with PBS or a probiotic strain, Lactobacillus johnsonii BS15 for 28 days prior to and throughout a 70 day exposure to sodium fluoride. Results showed that fluoride increases the hyperactivity of hypothalamic-pituitary-adrenal (HPA) axis and reduces the exploration ratio in novel object recognition (NOR) test and the spontaneous exploration during the T-maze test in mice following WAS, which were significantly improved by the probiotic. 16S rRNA sequencing showed a significant separation in ileal microbiota between the fluoride-treated mice and control mice. Lactobacillus was the main targeting bacteria and significantly reduced in fluoride-treated mice. BS15 reconstructed the fluoride-post microbiota and increased the relative abundance of Lactobacillus. D-lactate content and diamine oxidase (DAO) activity, two biomarkers of gut permeability were reduced in the serum of probiotic-inoculated mice. ZO-1, an intestinal tight junction protein was reduced by fluoride in mRNA, and its protein levels were increased by the probiotic treatment. Moreover, the hippocampus which is essential to learning and memory, down-regulated mRNA level of both the myelin-associated glycoprotein (MAG), and protein levels of brain-derived neurotrophic factor (BDNF), including the improvement of cAMP response element-binding protein (CREB) by BS15 in fluoride-exposed mice after WAS. Via spearman correlation analysis, Lactobacillus displayed significantly positive associations with the behavioral tests, levels of nerve development related factors, and intestinal tight junction proteins ZO-1, and negative association with TNF-α of the hippocampus, highlighting regulatory effects of gut bacteria on memory potential and gut barrier. These results suggested the psychoactive effects of BS15 on fluoride-induced memory dysfunction after psychological stress. In addition, there may be some correlations between fluoride-induced memory dysfunction and reconstruction of gut microbiota. AVAILABILITY OF DATA AND MATERIALS: 16S rRNA sequencing reads have uploaded to NCBI. The accession code of 16S rRNA sequencing reads in the National Center for Biotechnology Information (NCBI) BioProject database: PRJNA660154.


Assuntos
Fluoretos/metabolismo , Microbioma Gastrointestinal/fisiologia , Probióticos/farmacologia , Animais , Encéfalo/metabolismo , Fator Neurotrófico Derivado do Encéfalo/metabolismo , Hipocampo/metabolismo , Sistema Hipotálamo-Hipofisário/metabolismo , Lactobacillus/metabolismo , Masculino , Memória , Transtornos da Memória/induzido quimicamente , Camundongos , Microbiota , Sistema Hipófise-Suprarrenal/metabolismo , RNA Ribossômico 16S/metabolismo , Fator de Necrose Tumoral alfa/metabolismo , Proteína da Zônula de Oclusão-1/metabolismo
7.
Ecotoxicol Environ Saf ; 217: 112225, 2021 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-33864983

RESUMO

Long-term exposure to excessive fluoride causes chronic damage in the body tissues and could lead to skeletal and dental fluorosis. Cartilage damage caused by excessive fluoride intake has gained wide attention, but how fluoride accumulation blocks the development of chondrocytes is still unclear. Here, we report a negative correlation between the length and growth plate width after NaF treatments via apoptosis and autophagy, with shrinkage of cells, nuclear retraction, dissolution of chondrocytes. Whereas, fluoride exposure had no significant effect on the number and distribution of the osteoclasts which were well aligned. More importantly, fluoride exposure induced apoptosis of tibial bone through CytC/Bcl-2/P53 pathways via targeting Caspase3, Caspase9, Bak1, and Bax expressions. Meanwhile, the Beclin1, mTOR, Pakin, Pink, and p62 were elevated in NaF treatment group, which indicated that long-term excessive fluoride triggered the autophagy in the tibial bone and produced the chondrocyte injury. Altogether, fluoride exposure induced the chondrocyte injury by regulating the autophagy and apoptosis in the tibial bone of ducks, which demonstrates that fluoride exposure is a risk factor for cartilage development. These findings revealed the essential role of CytC/Bcl-2/P53 pathways in long-term exposure to fluoride pollution and block the development of chondrocytes in ducks, and CytC/Bcl-2/P53 can be targeted to prevent fluoride induced chondrocyte injury.


Assuntos
Condrócitos/fisiologia , Patos/fisiologia , Fluoretos/toxicidade , Animais , Apoptose/efeitos dos fármacos , Autofagia/efeitos dos fármacos , Proteína Beclina-1/metabolismo , Condrócitos/efeitos dos fármacos , Condrogênese , Fluoretos/metabolismo , Lâmina de Crescimento
8.
J Am Chem Soc ; 143(10): 3753-3763, 2021 03 17.
Artigo em Inglês | MEDLINE | ID: mdl-33630577

RESUMO

The lack of efficient [18F]fluorination processes and target-specific organofluorine chemotypes remains the major challenge of fluorine-18 positron emission tomography (PET). We report here an ultrafast isotopic exchange method for the radiosynthesis of novel PET agent aryl [18F]fluorosulfate enabled by the emerging sulfur fluoride exchange (SuFEx) click chemistry. The method has been applied to the fully automated 18F-radiolabeling of 25 structurally and functionally diverse aryl fluorosulfates with excellent radiochemical yield (83-100%, median 98%) and high molar activity (280 GBq µmol-1) at room temperature in 30 s. The purification of radiotracers requires no time-consuming HPLC but rather a simple cartridge filtration. We further demonstrate the imaging application of a rationally designed poly(ADP-ribose) polymerase 1 (PARP1)-targeting aryl [18F]fluorosulfate by probing subcutaneous tumors in vivo.


Assuntos
Química Click , Fluoretos/química , Compostos Radiofarmacêuticos/síntese química , Compostos de Enxofre/química , Animais , Linhagem Celular Tumoral , Meios de Contraste/síntese química , Meios de Contraste/química , Meios de Contraste/metabolismo , Teoria da Densidade Funcional , Estabilidade de Medicamentos , Fluoretos/síntese química , Fluoretos/metabolismo , Radioisótopos de Flúor/química , Humanos , Camundongos , Neoplasias/diagnóstico por imagem , Poli(ADP-Ribose) Polimerases/química , Poli(ADP-Ribose) Polimerases/metabolismo , Tomografia por Emissão de Pósitrons , Compostos Radiofarmacêuticos/metabolismo , Compostos de Enxofre/síntese química , Compostos de Enxofre/metabolismo , Transplante Heterólogo
9.
ACS Appl Mater Interfaces ; 13(2): 2327-2335, 2021 Jan 20.
Artigo em Inglês | MEDLINE | ID: mdl-33401893

RESUMO

This study presents a controlled synthesis of NaYbF4@NaYF4 core-shell upconversion nanoparticles using the hot-injection technique. NaYF4 shells with tunable morphologies including long-rod, short-rod, and quasi-sphere are grown on identical NaYbF4 core nanoparticles by controlled injection of acetate or trifluoroacetate precursors. Mechanistic investigations reveal that anisotropic interfacial strain accounts for the preferential growth of shell layers along the c-axis. However, the strain effect can be offset by the fast injection of shell precursors, leading to nearly isotropic growth of NaYF4 shells over the NaYbF4 core nanoparticles. The core-shell nanoparticles are further modified with DNA molecules and incubated with adenocarcinomic human alveolar basal epithelial cells. Based on a combination of characterizations by flow cytometry and confocal microscopy, favorable cellular uptake and DNA delivery are observed for the quasi-sphere nanoparticles, owing to the high dispersibility and easy membrane wrapping. The method described here could be extended to synthesize other types of functional nanostructures for the study of morphology-dependent properties.


Assuntos
Portadores de Fármacos/química , Fluoretos/química , Nanopartículas/química , Itérbio/química , Ítrio/química , Células A549 , DNA/administração & dosagem , DNA/farmacocinética , Portadores de Fármacos/metabolismo , Fluoretos/metabolismo , Técnicas de Transferência de Genes , Humanos , Nanopartículas/metabolismo , Nanopartículas/ultraestrutura , Nanotecnologia , Itérbio/metabolismo , Ítrio/metabolismo
10.
Life Sci ; 270: 119122, 2021 Apr 01.
Artigo em Inglês | MEDLINE | ID: mdl-33508294

RESUMO

The adrenal glands have striking morpho-biochemical features that render them vulnerable to the effects of toxins. AIMS: This study was conducted to explore the therapeutic utility of extracellular vesicles derived from bone marrow mesenchymal stem cells (BMSC-EVs) against fluoride-induced adrenal toxicity. MATERIALS AND METHODS: The work included isolation and further identification of BMSC-EVs by transmission electron microscopy and flow cytometric analysis. Adrenal toxicity in rats was induced by oral administration of 300 ppm of sodium fluoride (NaF) in drinking water for 60 days followed by a single dose injection of BMSC-EVs. The effects of BMSC-EVs against NaF was evaluated by adrenal oxidant/antioxidant biomarkers, hormonal assay of plasma adrenocorticotrophic hormone (ACTH) and corticosterone (CORT) and mRNA gene expression quantitation for adrenal cortical steroidogenic pathway-encoding genes. Histopathological examination of the adrenal tissue was performed. KEY FINDINGS: BMSC-EVs were effectively isolated and characterized. NaF exposure decreased adrenal superoxide dismutase and catalase activities, increased adrenal malondialdehyde levels, elevated plasma ACTH, diminished CORT concentrations and downregulated the adrenal cortical steroidogenic pathway-encoding genes. In addition, NaF-induced marked adrenal histopathological lesions. SIGNIFICANCE: BMSC-EVs treatment repaired damaged adrenal tissue and recovered its function greatly following NaF consumption. BMSC-EVs reversed the toxic effects of NaF and reprogramed injured adrenal cells by activating regenerative processes.


Assuntos
Glândulas Suprarrenais/metabolismo , Vesículas Extracelulares/transplante , Células-Tronco Mesenquimais/metabolismo , Animais , Células da Medula Óssea/citologia , Diferenciação Celular/efeitos dos fármacos , Vesículas Extracelulares/metabolismo , Feminino , Fluoretos/efeitos adversos , Fluoretos/metabolismo , Células-Tronco Mesenquimais/citologia , Células-Tronco Mesenquimais/fisiologia , Microscopia Eletrônica de Transmissão/métodos , Ratos
11.
Ecotoxicol Environ Saf ; 208: 111437, 2021 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-33096359

RESUMO

Long-term exposure to high concentrations of fluoride (F) can damage mineralized and soft tissues such as bones, liver, kidney, intestine, and nervous system of adult rats. The high permeability of the blood-brain barrier and placenta to F during pregnancy and lactation may be critical to neurological development. Therefore, this study aimed to investigate the effects of F exposure during pregnancy and lactation on molecular processes and oxidative biochemistry of offspring rats' hippocampus. Pregnant Wistar rats were randomly assigned into 3 groups in accordance with the drinking water received: G1 - deionized water (control); G2 - 10 mg/L of F and G3 - 50 mg/L of F. The exposure to fluoridated water began on the first day of pregnancy and lasted until the 21st day of breastfeeding (when the offspring rats were weaned). Blood plasma samples of the offspring rats were collected to determine F levels. Hippocampi samples were collected for oxidative biochemistry analyses through antioxidant capacity against peroxyl (ACAP), lipid peroxidation (LPO), and nitrite (NO2-) levels. Also, brain-derived neurotrophic factor (BDNF) gene expression (RT-qPCR) and proteomic profile analyses were performed. The results showed that exposure to both F concentrations during pregnancy and lactation increased the F bioavailability, triggered redox imbalance featured by a decrease of ACAP, increase of LPO and NO2- levels, BDNF overexpression and changes in the hippocampus proteome. These findings raise novel questions regarding potential repercussions on the hippocampus structure and functioning in the different cognitive domains.


Assuntos
Poluentes Ambientais/toxicidade , Fluoretos/toxicidade , Hipocampo/efeitos dos fármacos , Estresse Oxidativo/fisiologia , Animais , Antioxidantes/metabolismo , Fator Neurotrófico Derivado do Encéfalo , Feminino , Fluoretos/metabolismo , Hipocampo/crescimento & desenvolvimento , Lactação , Peroxidação de Lipídeos/efeitos dos fármacos , Masculino , Estresse Oxidativo/efeitos dos fármacos , Gravidez , Efeitos Tardios da Exposição Pré-Natal/metabolismo , Proteoma/metabolismo , Proteômica , Ratos , Ratos Wistar
12.
J Sci Food Agric ; 101(2): 379-387, 2021 Jan 30.
Artigo em Inglês | MEDLINE | ID: mdl-32623727

RESUMO

Tea is the one of the most popular non-alcoholic caffeinated beverages in the world. Tea is produced from the tea plant (Camellia sinensis (L.) O. Kuntze), which is known to accumulate fluoride. This article systematically analyzes the literature concerning fluoride absorption, transportation and fluoride tolerance mechanisms in tea plants. Fluoride bioavailability and exposure levels in tea infusions are also reviewed. The circulation of fluoride within the tea plantation ecosystems is in a positive equilibrium, with greater amounts of fluoride introduced to tea orchards than removed. Water extractable fluoride and magnesium chloride (MgCl2 ) extractable fluoride in plantation soil are the main sources of absorption by tea plant root via active trans-membrane transport and anion channels. Most fluoride is readily transported through the xylem as F- /F-Al complexes to leaf cell walls and vacuole. The findings indicate that tea plants employ cell wall accumulation, vacuole compartmentalization, and F-Al complexes to co-detoxify fluoride and aluminum, a possible tolerance mechanism through which tea tolerates higher levels of fluoride than most plants. Furthermore, dietary and endogenous factors influence fluoride bioavailability and should be considered when exposure levels of fluoride in commercially available dried tea leaves are interpreted. The relevant current challenges and future perspectives are also discussed. © 2020 Society of Chemical Industry.


Assuntos
Camellia sinensis/química , Fluoretos/análise , Fluoretos/metabolismo , Alumínio/análise , Alumínio/metabolismo , Disponibilidade Biológica , Transporte Biológico , Camellia sinensis/metabolismo , Parede Celular/química , Parede Celular/metabolismo , Exposição Dietética/efeitos adversos , Exposição Dietética/análise , Humanos , Folhas de Planta/química , Folhas de Planta/metabolismo , Medição de Risco , Solo/química , Chá/química
13.
Chemosphere ; 262: 127796, 2021 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-32755695

RESUMO

The literature is sparse in terms of the effect of exercise on the pharmacokinetics of fluoride (F) in humans. In a 4-treatment repeated measures cross-over study, we investigated F pharmacokinetics following no exercise (control) and three exercise intensity conditions (light, moderate and vigorous) in healthy adults. At a pre-experimental session, 8 participants (18-30y) residing in a non-fluoridated-area, underwent a VO2 max test to guide the three exercise intensities for the experimental sessions. Participants were on a F-free regime one week before and throughout the four experimental weeks. We measured urinary F excretion (UFE), maximum plasma concentration (Cmax), lag time of Cmax (Tmax), and Area Under the Curve (AUC) for plasma F concentration against time, following F ingestion then no, light, moderate and vigorous exercise. Results showed no statistically significant difference in Tmax among all sessions; whereas Cmax for moderate exercise (226.2 ng/ml) was significantly higher than for no (27.0 ng/ml; p < 0.001), light (105.6 ng/ml; p = 0.016) and vigorous (94.2 ng/ml; p = 0.008) exercise. Mean AUC over 0-90 min following F ingestion was also significantly higher in moderate exercise than for no (p < 0.001), light (p = 0.004) and vigorous (p = 0.001) exercise. Mean UFE over 0-14h was 638.8, 718.7, 574.6 and 450.5  µg for no, light, moderate and vigorous exercise, with no statistically significant differences among different sessions. In conclusion, this human experimental study suggests that moderate exercise may increase the fraction of F absorbed systemically which is therefore available to produce a biological effect. Future studies should be conducted with larger samples, different age groups and using different F doses.


Assuntos
Poluentes Ambientais/metabolismo , Fluoretos/metabolismo , Adulto , Área Sob a Curva , Estudos Cross-Over , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Minerais
14.
Nat Commun ; 11(1): 5045, 2020 10 07.
Artigo em Inglês | MEDLINE | ID: mdl-33028813

RESUMO

Fluorine is a key element in the synthesis of molecules broadly used in medicine, agriculture and materials. Addition of fluorine to organic structures represents a unique strategy for tuning molecular properties, yet this atom is rarely found in Nature and approaches to integrate fluorometabolites into the biochemistry of living cells are scarce. In this work, synthetic gene circuits for organofluorine biosynthesis are implemented in the platform bacterium Pseudomonas putida. By harnessing fluoride-responsive riboswitches and the orthogonal T7 RNA polymerase, biochemical reactions needed for in vivo biofluorination are wired to the presence of fluoride (i.e. circumventing the need of feeding expensive additives). Biosynthesis of fluoronucleotides and fluorosugars in engineered P. putida is demonstrated with mineral fluoride both as only fluorine source (i.e. substrate of the pathway) and as inducer of the synthetic circuit. This approach expands the chemical landscape of cell factories by providing alternative biosynthetic strategies towards fluorinated building-blocks.


Assuntos
Redes Reguladoras de Genes , Halogenação/genética , Engenharia Metabólica/métodos , Pseudomonas putida/metabolismo , Biologia Sintética/métodos , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Vias Biossintéticas/genética , RNA Polimerases Dirigidas por DNA/genética , Fluoretos/metabolismo , Flúor/metabolismo , Proteínas de Membrana Transportadoras/genética , Proteínas de Membrana Transportadoras/metabolismo , Mutação , Pseudomonas putida/genética , RNA Bacteriano/genética , Riboswitch/genética , Proteínas Virais/genética
15.
Nat Commun ; 11(1): 5459, 2020 10 29.
Artigo em Inglês | MEDLINE | ID: mdl-33122649

RESUMO

Biocontainment systems are needed to neutralize genetically modified organisms (GMOs) that pose ecological threats outside of controlled environments. In contrast, benign selection markers complement GMOs with reduced fitness. Benign selection agents serve as alternatives to antibiotics, which are costly and risk spread of antibiotic resistance. Here, we present a yeast biocontainment strategy leveraging engineered fluoride sensitivity and DNA vectors enabling use of fluoride as a selection agent. The biocontainment system addresses the scarcity of platforms available for yeast despite their prevalent use in industry and academia. In the absence of fluoride, the biocontainment strain exhibits phenotypes nearly identical to those of the wildtype strain. Low fluoride concentrations severely inhibit biocontainment strain growth, which is restored upon introduction of fluoride-based vectors. The biocontainment strategy is stringent, easily implemented, and applicable to several eukaryotes. Further, the DNA vectors enable genetic engineering at reduced costs and eliminate risks of propagating antibiotic resistance.


Assuntos
Fluoretos , Proteínas de Membrana/genética , Organismos Geneticamente Modificados/genética , Proteínas de Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/genética , Seleção Genética , Fluoretos/metabolismo , Fluoretos/farmacologia , Engenharia Genética , Marcadores Genéticos , Saccharomyces cerevisiae/efeitos dos fármacos , Biologia Sintética/métodos
16.
J Immunol Methods ; 487: 112875, 2020 12.
Artigo em Inglês | MEDLINE | ID: mdl-33031794

RESUMO

The expression of Kv1.3 and KCa channels in human T cells is essential for maintaining cell activation, proliferation and migration during an inflammatory response. Recently, an additional residual current, sensitive to anandamide and A293, compounds specifically inhibiting currents mediated by TASK channels, was observed after complete pharmacological blockade of Kv1.3 and KCa channels. This finding was not consistently observed throughout different studies and, an in-depth review of the different recording conditions used for the electrophysiological analysis of K+ currents in T cells revealed fluoride as major anionic component of the pipette intracellular solutions in the initial studies. While fluoride is frequently used to stabilize electrophysiological recordings, it is known as G-protein activator and to influence the intracellular Ca2+ concentration, which are mechanisms known to modulate TASK channel functioning. Therefore, we systemically addressed different fluoride- and chloride-based pipette solutions in whole-cell patch-clamp experiments in human T cells and used specific blockers to identify membrane currents carried by TASK and Kv1.3 channels. We found that fluoride increased the decay time constant of K+ outward currents, reduced the degree of the sustained current component and diminished the effect of the specific TASK channels blocker A293. These findings indicate that the use of fluoride-based pipette solutions may hinder the identification of a functional TASK channel component in electrophysiological experiments.


Assuntos
Fluoretos/farmacologia , Potenciais da Membrana/efeitos dos fármacos , Canais de Potássio de Domínios Poros em Tandem/metabolismo , Compostos de Potássio/farmacologia , Linfócitos T/efeitos dos fármacos , Células Cultivadas , Fluoretos/metabolismo , Humanos , Canal de Potássio Kv1.3/efeitos dos fármacos , Canal de Potássio Kv1.3/metabolismo , Cloreto de Magnésio/metabolismo , Cloreto de Magnésio/farmacologia , Técnicas de Patch-Clamp , Bloqueadores dos Canais de Potássio/farmacologia , Canais de Potássio de Domínios Poros em Tandem/efeitos dos fármacos , Compostos de Potássio/metabolismo , Linfócitos T/metabolismo , Fatores de Tempo
17.
Ecotoxicol Environ Saf ; 203: 110951, 2020 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-32678752

RESUMO

The growing use of rare-earth doped upconversion nanoparticles (UCNPs) has caused increasing concern about their biosafety. Here, to understand the toxicity of UCNPs and their mechanism in HepG2 cells, we systematically study the cytotoxicity, uptake and elimination behaviors of three types of UCNPs combined multiple cytotoxicity evaluation means with inductively coupled plasma mass spectrometry (ICP-MS) detection. Sodium yttrium fluoride, doped with 18% (molar ratio) ytterbium and 2% erbium (NaYF4: Yb3+, Er3+) was selected as the model UCNPs with two sizes (35 and 55 nm), and the poly(acrylic acid) and polyethylenimine were selected as the representatives of negative and positive surface coating of UCNPs, respectively. UCNPs were found to induce cytotoxicity in time- and dose-dependent manners, which might be mediated by reactive oxygen species generation and oxidative stress. Apoptosis, inflammation, and metabolic process were enhanced after cells exposed to 200 mg/L UCNPs for 48 h. Increase in the protein levels of cleaved caspased-9, cleaved caspase-3 and Bax and decrease in the anti-apoptotic protein, Bcl-2 suggested that the mitochondria mediated pathway was involved in UCNP-induced apoptosis. With the aid of ICP-MS, it demonstrated that the cytotoxicity was associated with internalized amount of UCNPs, which largely relied on their surface properties rather than size in the tested range. By comparing UCNPs with Y3+ ions, it demonstrated that NPs properties played a nonnegligible role in the cytotoxicity of UCNPs. These findings provide new insights for fundamental understanding of cytotoxicity of UCNPs and may contribute to more rational use of these materials in the future.


Assuntos
Endocitose/efeitos dos fármacos , Érbio/toxicidade , Fluoretos/toxicidade , Nanopartículas/toxicidade , Itérbio/toxicidade , Ítrio/toxicidade , Apoptose/efeitos dos fármacos , Técnicas de Cultura de Células , Sobrevivência Celular , Érbio/química , Érbio/metabolismo , Fluoretos/química , Fluoretos/metabolismo , Expressão Gênica/efeitos dos fármacos , Células Hep G2 , Humanos , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Nanopartículas/química , Nanopartículas/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Tamanho da Partícula , Propriedades de Superfície , Itérbio/química , Itérbio/metabolismo , Ítrio/química , Ítrio/metabolismo
18.
Arch Oral Biol ; 117: 104777, 2020 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-32592930

RESUMO

OBJECTIVE: To clarify the fluoride/mineral kinetics in an oral biofilm following concurrent application of fluoride and other mineral ions released from experimental toothpaste containing S-PRG filler using depth-specific analysis. STUDY DESIGN: Twenty subjects wore in situ plaque-generating devices, comprised of a pair of enamel slabs, and a biofilm was allowed to form. The devices were removed after three days, immersed in the toothpaste filtrate containing Al, B, Sr and F ions for 1 min, and then reinserted at the same location. After 30 min, the devices were removed and samples were obtained by sectioning into outer, middle and inner biofilm layers (300-µm thick). Samples treated with filtrate containing F without S-PRG filler extract served as the control. Fluoride and the three other mineral ions extracted from 4-µm sections were quantified using a fluoride electrode and ICP-AES, respectively. The results were corrected for biomass volume, estimated by the area measurement of stained 2-µm sections. RESULTS: The mean uptake ratios (S-PRG/control, ng/mm3) of Al, B, Sr and F were 186.6/53.7, 58.4/25.0, 456.9/125.7 and 43.6/12.0, respectively, in the outer layer, indicating that the mineral ions could easily diffuse into the biofilm. F concentrations in the outer biofilm treated using filtrate with S-PRG filler extract were significantly higher than those in controls, although both biofilms were exposed to filtrates containing the same level of F. CONCLUSIONS: The results suggest that toothpaste containing S-PRG filler promotes fluoride retention in oral biofilms via the uptake of other mineral ions.


Assuntos
Biofilmes , Placa Dentária , Fluoretos/metabolismo , Cimentos de Ionômeros de Vidro/química , Minerais/metabolismo , Cremes Dentais/química , Esmalte Dentário , Placa Dentária/tratamento farmacológico , Humanos
19.
J Agric Food Chem ; 68(32): 8568-8579, 2020 Aug 12.
Artigo em Inglês | MEDLINE | ID: mdl-32559071

RESUMO

The accumulation of fluoride in tea leaves from various cultivars exhibits significant differences. However, the molecular basis and mechanism remain largely unknown. Here, we reported that two genes of CsFEX (fluoride export genes in Camellia sinensis), CsFEX1 and CsFEX2, transport fluoride out of cells, alleviate the cellular fluoride toxin, and rescue the yeast mutant (FEX1ΔFEX2Δ) and Arabidopsis mutant (fex), as their efflux activities are coupled with proton gradients. Further analysis found that CsFEX1 and CsFEX2 localize to the plasma membrane both in yeast and Arabidopsis cells. CsFEX2 is more effective to reduce fluoride toxicity in yeast and Arabidopsis compared with CsFEX1 even at low pH. CsFEX2 induced by fluoride treatment is around tenfold higher in a low-fluoride cultivar (Yunkang 10) than that in a high-fluoride cultivar (Pingyang Tezaocha), suggesting that CsFEX2 possibly plays a critical role in reducing fluoride accumulation in tea leaves.


Assuntos
Camellia sinensis/metabolismo , Proteínas de Transporte/metabolismo , Fluoretos/metabolismo , Proteínas de Plantas/metabolismo , Transporte Biológico , Camellia sinensis/química , Camellia sinensis/genética , Proteínas de Transporte/genética , Fluoretos/análise , Folhas de Planta/química , Folhas de Planta/genética , Folhas de Planta/metabolismo , Proteínas de Plantas/genética
20.
J Sci Food Agric ; 100(15): 5498-5509, 2020 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-32567049

RESUMO

BACKGROUND: The East African Rift Valley (EARV) area is characterized by an intense volcanic activity, which largely influences the nature of soils, ground and surface waters causing a transfer of fluoride from volcanic emissions to the environment. Field experiments were conducted in fluorine-contaminated areas of Ngarenanyuki (Arumeru district) in north Tanzania. In order to evaluate the potential fluoride exposure from diet and the related health risk for the local population, the content of fluoride in soil and plant tissues was assessed, focusing on the edible portions (leaves, fruits or seeds) of the main cultivated and consumed food crops in the area. RESULTS: Average fluoride contents of 8.0, 11.4, 11.3 and 14.2 mg kg-1 of dry matter were observed respectively for maize (Zea mays L.), tomato (Lycopersicon esculentum Mill.), bean (Phaseolus vulgaris L.) and kale (Brassica sp. pl.) edible parts. The cumulative estimated average daily dose (EADD) ranged from 0.026 to 0.165 mg F d-1 kg-1 among different rural population groups and considering two different hypotheses of absorption fraction (75% or 100%), i.e. the amount of fluoride that is absorbed during the digestion process. The associated hazard index (HI) values varied from 0.43 to 2.75. CONCLUSIONS: Considering the dietary habits of the local population, the outcomes of the present study suggest that the investigated crops can substantially contribute to fluoride related diseases, especially in earlier ages. © 2020 Society of Chemical Industry.


Assuntos
Produtos Agrícolas/crescimento & desenvolvimento , Produtos Agrícolas/metabolismo , Fluoretos/metabolismo , Transporte Biológico , Brassica/química , Brassica/crescimento & desenvolvimento , Brassica/metabolismo , Exposição Dietética/análise , Fabaceae/química , Fabaceae/crescimento & desenvolvimento , Fabaceae/metabolismo , Fluoretos/análise , Humanos , Lycopersicon esculentum/química , Lycopersicon esculentum/crescimento & desenvolvimento , Lycopersicon esculentum/metabolismo , Solo/química , Poluentes do Solo/análise , Poluentes do Solo/metabolismo , Tanzânia , Zea mays/química , Zea mays/crescimento & desenvolvimento , Zea mays/metabolismo
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