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1.
Nihon Yakurigaku Zasshi ; 154(3): 138-142, 2019.
Artigo em Japonês | MEDLINE | ID: mdl-31527364

RESUMO

Zinc, an essential trace element, plays an important role in a large number of biological functions. In mammalian brain, whereas the majority of brain zinc is bound to proteins including metallothionein, about 5-15% is stored in presynaptic vesicles of glutamatergic neurons throughout the forebrain, especially in the hippocampus, in a relatively free state. Thus, free zinc (Zn2+) concentration in the brain is considered to be regulated in order to maintain normal brain functions such as learning and memory. On the other hand, brain Zn2+ dyshomeostasis has been recognized as a mechanism for neuronal injury in brain disorders including Alzheimer's disease and brain ischemia. In particular, after transient brain ischemia, Zn2+ accumulates in hippocampal neurons via a zinc transport system, or via release from cytosolic zinc-binding proteins, which results in neuronal cell death. Recently, it has been demonstrated that Zn2+ dyshomeostasis also occurs in glial cells such as microglia, astrocytes and oligodendrocytes after brain ischemia. In oligodendrocytes, ischemic insult triggers intracellular Zn2+ accumulation, resulting in cell death via mitochondrial dysfunction. Increased extracellular Zn2+ inhibits astrocytic glutamate uptake. In addition, extracellular Zn2+ massively released from ischemic neurons primes microglia to enhance production of pro-inflammatory cytokines in response to stimuli that trigger M1 activation. This review aims to describe the impact of brain Zn2+ dyshomeostasis on alterations in glial cell survival and functions in post-ischemic brains.


Assuntos
Química Encefálica , Isquemia Encefálica/fisiopatologia , Acidente Vascular Cerebral/fisiopatologia , Zinco/fisiologia , Animais , Astrócitos , Encéfalo , Microglia , Neurônios , Oligodendroglia
2.
Chem Commun (Camb) ; 55(67): 9955-9958, 2019 Aug 28.
Artigo em Inglês | MEDLINE | ID: mdl-31364619

RESUMO

A silver nanocluster-based ratiometric fluorescent nanosensor was developed for the determination of ATP in the cerebrospinal fluid of a mouse brain. Using this useful tool with good stability and high selectivity as well as a wide linear detection range, it was found that the ATP concentration in a mouse brain with Alzheimer's disease was 2300-fold higher than that in a normal one.


Assuntos
Trifosfato de Adenosina/líquido cefalorraquidiano , Química Encefálica , DNA/química , Corantes Fluorescentes/química , Nanopartículas Metálicas/química , Doença de Alzheimer/metabolismo , Doença de Alzheimer/patologia , Animais , Encéfalo/citologia , Encéfalo/patologia , Córtex Cerebral/química , Hipocampo/química , Camundongos , Conformação de Ácido Nucleico , Prata/química , Espectrometria de Fluorescência/métodos
3.
Zhongguo Zhong Yao Za Zhi ; 44(13): 2841-2848, 2019 Jul.
Artigo em Chinês | MEDLINE | ID: mdl-31359699

RESUMO

Curcumin( Cur) is a natural active substance extracted from the roots or tubers of traditional Chinese medicinal materials. It has anti-inflammatory and anti-tumor activities on brain diseases. Due to the poor stability,low solubility,poor absorption and low bioavailability of curcumin,N-acetyl-L-cysteine( NAC) was used as an absorption enhancer and mixed with curcumin to improve the absorption of curcumin in the body. In this paper,curcumin was smashed by airflow pulverization,and Cur-NAC mixtures were prepared by being grinded with liquid. Then,the raw material and the product were analyzed by differential scanning calorimetry( DSC),X-ray diffraction( XRD) for structural characterization. The dissolution was determined by high performance liquid chromatography( HPLC) analysis. The characteristic peaks of the samples prepared by grinding method were similar to those of the raw materials,while the melting temperature and the accumulated dissolution degree were not significantly changed. The crystal forms of the products were not changed,and no new crystal form was formed after grinding. After the administration of intranasal powder,blood samples were collected from the orbit,while the whole brain tissues were removed from the skull and dissected into 10 anatomical regions. The concentrations of curcumin in these samples were determined by UPLC-MS/MS. The concentrations of curcumin in plasma and brain were compared at different time points. After intranasal administration of two drugs,it was found that the concentration of curcumin after sniffing up the mixtures in plasma was high,and the concentration of the drug in the olfactory bulb,hippocampus,and pons was increased significantly. Within 0. 083-0. 5 h,the olfactory bulb,piriform lobe and hippocampus remained high concentrations,the endodermis,striatum,hypothalamus and midbrain reached high concentrations within 1-3 h; and the cerebellum,pons and brain extension maintained relatively high concentrations within 3-7 h. The experiment showed that nasal administration of Cur-NAC mixtures can significantly improve the bioavailability of curcumin,and lead to significant differences in brain tissue distribution.


Assuntos
Acetilcisteína/farmacologia , Química Encefálica , Curcumina/farmacocinética , Administração Intranasal , Animais , Disponibilidade Biológica , Encéfalo , Cromatografia Líquida , Ratos , Espectrometria de Massas em Tandem , Distribuição Tecidual
4.
J Forensic Leg Med ; 65: 133-136, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31153008

RESUMO

BACKGROUND: Benefits and even dangers of plants are known since time began. The ancients used plants and herbs because of their effects on the human body. Poisoning is a logical consequence of their use: history is full of episodes of plants and herbs poisoning, whether intentional or accidental. AIM: Oleander poisoning is generally accidental; an intentional assumption of its leaves to commit suicide is uncommon because the population is not aware of the harmfulness of its cardiotoxic glycosides, therefore we report a fatal case of self-poisoning through the voluntary ingestion of oleander leaves. METHODS: A diagnosis of oleander self-poisoning was highly suspected on the basis of the circumstantial evidence and the autopsy findings. Toxicological investigations were performed on the samples collected during the autopsy and aimed at confirm the presence of oleandrin at a toxic level. RESULTS: The autopsy revealed a piece of oleander leaf on the posterior third of the tongue's body and several plant residues, similar to the one recovered on the tongue, into the gastric content; petechiae on the deep surface of the scalp, multi-organ congestion, and pulmonary edema were also observed. The histological study corroborated the pulmonary edema macroscopically observed but did not provide any other information. The detection of oleandrin in biological cadaveric samples revealed high, fatal, concentrations. CONCLUSIONS: Cases of voluntary ingestion of oleander with a suicidal intent prove to be uncommon: in the case reported the victim was aware about the possibility to commit suicide through the ingestion of oleander leaves.


Assuntos
Nerium/envenenamento , Folhas de Planta/envenenamento , Suicídio , Química Encefálica , Cardenolídeos/análise , Feminino , Vesícula Biliar/química , Mucosa Gástrica/química , Conteúdo Gastrointestinal/química , Humanos , Rim/química , Fígado/química , Pulmão/química , Pessoa de Meia-Idade , Edema Pulmonar/patologia , Baço/química
5.
Artigo em Russo | MEDLINE | ID: mdl-31166312

RESUMO

Evaluation of brain metabolism is an important part in examination of brain lesions. Phosphorus magnetic resonance spectroscopy opens up great opportunities for studying the energy metabolism and allows noninvasive examination of metabolic processes occurring both in healthy and in pathologic brain tissue by obtaining a spectrum of phosphorus-containing metabolites involved in the turnover of cell membrane phospholipids. The technique presented in this paper was used to conduct 31P MR spectroscopy and to estimate the ratio between the peaks of the main metabolites and intracellular pH of the healthy brain tissue of 23 volunteers in the age group under 30 years old in clinical settings. Based on the recorded stable phosphorus spectra of metabolites of the healthy brain tissue, the value of intracellular pH (6.963±0.044) and the ratio of the main PME/PDE peaks (1.17±0.20) were calculated. The database was created to subsequently analyze the metabolic changes in brain tissue spectra in norm and in pathology, as well as the intracellular pH variations that have diagnostic and prognostic value.


Assuntos
Encéfalo , Fósforo , Adulto , Encéfalo/diagnóstico por imagem , Química Encefálica , Humanos , Concentração de Íons de Hidrogênio , Espectroscopia de Ressonância Magnética , Voluntários
6.
Forensic Sci Int ; 301: 388-393, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31226641

RESUMO

Brain tissue is a useful supplement to blood in postmortem investigations, but reference concentrations are scarce for many opioids. Heroin cases may be difficult to distinguish from morphine cases as heroin and its metabolites are rapidly degraded. We present concentrations from brain and blood and brain-blood ratios of 98 cases where morphine was quantified. These cases were grouped according to the cause of death: A: The compound was solely assumed to have caused a fatal intoxication. B: The compound presumably contributed to a fatal outcome in combination with other drugs, alcohol or disease. C: The compound was not regarded to be related to the cause of death. The cases were further classified as heroin cases if 6-acetyl-morphine or noscapine were detected. The analyses were carried out using solid-phase extraction or protein precipitation followed by ultra high-performance liquid chromatography coupled to mass spectrometry. The average brain-blood ratios of morphine were 1.2 and 1.8 for 69 morphine and 29 heroin cases, respectively. Differences in the brain-blood ratios were found for cases where heroin and morphine were involved in the cause of death, either in combination or on its own (P<0.001 and P=0.004, respectively). However, the overlap between morphine and heroin cases precludes the use of the brain-blood ratio to distinguish heroin from morphine intake. Morphine-6-glucuronide and 6-acetyl-morphine were quantified in brain and blood in a subset of the samples, yielding median brain-blood ratios of 5.1 and 8.3, respectively. The brain concentrations may aid the toxicological investigation in cases where heroin or morphine intoxications are suspected, but blood is not available.


Assuntos
Química Encefálica , Heroína/análise , Morfina/análise , Entorpecentes/análise , Cromatografia Líquida , Overdose de Drogas/diagnóstico , Toxicologia Forense/métodos , Humanos , Espectrometria de Massas , Derivados da Morfina/análise , Noscapina/análise , Envenenamento/diagnóstico
7.
Artigo em Chinês | MEDLINE | ID: mdl-31189237

RESUMO

Objective: To investigate the dynamic changes of copper and iron contents in brain tissue, body fluids and barriers of rats exposed to lead at different periods in order to provide a theoretical basis for the study of the mechanism of lead nerve injury. Methods: Sixty-four healthy adult SPF male SD rats were randomly divided into control group and lead exposure group, after one week of adaptive feeding, rats in the lead exposure group were treated with 250 mg/L lead acetate, and rats in control group were treated with ordinary drinking water, the experimental period was 12 weeks. After exposure for 3, 6, 9 and 12 weeks, the samples including blood, choroid plexus, cerebrospinal fluid, cortex, hippocampus, striatum, hypothalamus, amygdala, substantia nigra and cerebellum were obtained. Lead, copper and iron content in all kinds of samples were detected by Inductively Coupled Plasma Mass Spectrometry(ICP-MS). The measurement data were presented as Mean±SD, Comparison of metal contents in different tissues of rats at different time analyzed using repeated measurement analysis of variance, Two-variable correlation analysis using Spearman correlation test.The relationship between lead exposure experiod and copper and iron in samples was studied by using trend test. Results: After 12 weeks of lead exposure compared with the control group, lead contents in cortex, hippocampus, striatum, hypothalamus, amygdala, substantia nigra and cerebellum of rats were 2.21, 2.44, 2.95, 3.53, 4.01, 1.85 and 2.86 folds of control group, and the differences were statistically significant(P<0.05). At the same time, lead content in blood, cerebrospinal fluid,choroid plexus, brain microvessels and bones increased. The increase rate in the amygdala and cerebrospinal fluid ranked first among brain tissue or barrier,which were 4.01 and 3.0 folds respectively. Compared with the control group, Compared with the control group, copper content in cortex,hippocampus, striatum, hypothalamus,amygdala, cerebellum,blood,cerebrospinal fluid,choroid plexus and cerebral microvasculature showed an increasing trend among rats following 3,6,9,12 weeks of lead exposure. Copper content change in the striatum was highest among all brain tissue. The increase rate of copper content in the striatum was at the top among brain tissues. After 12 weeks of lead exposure,copper content in brain microvessels was 4.98 folds higher than that of the control group (P<0.05). After lead exposure at different periods,the iron content in the cortex, hippocampus, striatum,cerebrospinal fluid,choroid plexus and brain microvessels of experimental rats all increased(P<0.05). And the iron increase rate in the hypothalamus or cerebrospinal fluid increase ranked first among brain tissues or body fluid the most obviously. Conclusion: With the increase of exposure time, lead exposure can changes in the contents of copper and iron in different brain tissues,body fluids and barriers in rats,among which, the contents of copper and iron in the amygdala,cerebrospinal fluid and brain microvessels increase significantly. This may be related to nerve damage from lead exposure.


Assuntos
Química Encefálica , Cobre , Ferro , Chumbo , Animais , Encéfalo , Cobre/farmacocinética , Ferro/farmacocinética , Chumbo/toxicidade , Masculino , Ratos , Ratos Sprague-Dawley
8.
Molecules ; 24(9)2019 May 03.
Artigo em Inglês | MEDLINE | ID: mdl-31058813

RESUMO

INTRODUCTION: Alcohol overuse may be related to increased aluminum (Al) exposure, the brain accumulation of which contributes to dementia. However, some reports indicate that silicon (Si) may have a protective role over Al-induced toxicity. Still, no study has ever explored the brain content of Al and Si in alcoholic use disorder (AUD). MATERIALS AND METHODS: To fill this gap, the present study employed inductively coupled plasma optical emission spectrometry to investigate levels of Al and Si in 10 brain regions and in the liver of AUD patients (n = 31) and control (n = 32) post-mortem. RESULTS: Al content was detected only in AUD patients at mean ± SD total brain content of 1.59 ± 1.19 mg/kg, with the highest levels in the thalamus (4.05 ± 12.7 mg/kg, FTH), inferior longitudinal fasciculus (3.48 ± 9.67 mg/kg, ILF), insula (2.41 ± 4.10 mg/kg) and superior longitudinal fasciculus (1.08 ± 2.30 mg/kg). Si content displayed no difference between AUD and control, except for FTH. Positive inter-region correlations between the content of both elements were identified in the cingulate cortex, hippocampus, and ILF. CONCLUSIONS: The findings of this study suggest that AUD patients may potentially be prone to Al-induced neurodegeneration in their brain-although this hypothesis requires further exploration.


Assuntos
Alcoolismo/complicações , Alumínio/análise , Química Encefálica , Doenças Neurodegenerativas/diagnóstico , Silício/análise , Adulto , Idoso , Alumínio/toxicidade , Autopsia , Estudos de Casos e Controles , Córtex Cerebral/química , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Doenças Neurodegenerativas/induzido quimicamente , Espectrofotometria Atômica , Tálamo/química
9.
Artigo em Inglês | MEDLINE | ID: mdl-31100605

RESUMO

The R&D of PET imaging agents is a complex system engineering, simplifying screening steps and increasing screening efficiency have become popular issues. The purpose of this study is to develop a new screening procedure using cassette-wave microdosing and LC-MS/MS to enhance the screening throughput of unradiolabeled candidate compounds as PET imaging agents. Nine compounds were divided into 3 sets and made into 3 cassettes. Fifteen rats were randomly divided into 3 groups, and every animal received three intravenous bolus injections at three different time points; the doses were at microdose levels. This dosing approach takes advantage of temporal and spatial differences and is likened to an input wave; therefore, this approach was named cassette-wave microdosing. The samples of different brain regions such as the hypothalamus, striatum, hippocampus, cortex, cerebellum and the remainder of the brain were detected by LC-MS/MS analysis. The research potential of the compounds as PET imaging agents is evaluated in terms of brain biodistribution data. The screening method is rapid, highly efficient, reliable and reduces animal usage. Additionally, it can shorten the evaluation process of radiopharmaceuticals and enhance the screening throughput of PET radiopharmaceuticals without the use of radioactive agents.


Assuntos
Compostos de Anilina , Química Encefálica/fisiologia , Cromatografia Líquida/métodos , Meios de Contraste , Tomografia por Emissão de Pósitrons , Espectrometria de Massas em Tandem/métodos , Administração Intravenosa , Compostos de Anilina/administração & dosagem , Compostos de Anilina/análise , Compostos de Anilina/farmacocinética , Animais , Encéfalo/metabolismo , Meios de Contraste/administração & dosagem , Meios de Contraste/análise , Meios de Contraste/farmacocinética , Limite de Detecção , Modelos Lineares , Masculino , Ratos , Ratos Sprague-Dawley , Reprodutibilidade dos Testes , Distribuição Tecidual
10.
Adv Exp Med Biol ; 1128: 1-11, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31062322

RESUMO

The brain or central nervous system (CNS) utilizes a vast amount of energy to sustain its basic functions, and most of the energy in the brain is derived from glucose. Whole-body energy and glucose homeostasis in the periphery of the human body are regulated by insulin, while the brain had been considered as an "insulin-insensitive" organ, because bulk brain glucose uptake is not affected by insulin in either rodents and humans. However, recently it has become clear that the actions of insulin are more widespread in the CNS and are a critical part of normal development, food intake, and energy balance, as well as plasticity throughout adulthood. Moreover, there are substantial evidence demonstrating that brain insulin is derived from pancreas, neurons, and astrocytes. In this chapter, I reviewed recent progress in roles of insulin in the brain, expression of insulin genes, and multiple origins of the brain insulin.


Assuntos
Química Encefálica , Encéfalo/fisiologia , Insulina/fisiologia , Ingestão de Alimentos , Metabolismo Energético , Humanos
11.
Anal Chim Acta ; 1071: 98-108, 2019 Sep 13.
Artigo em Inglês | MEDLINE | ID: mdl-31128761

RESUMO

In this work, we report a sensitive and selective electrochemical sensor for the detection of dopamine (DA) neurotransmitter based on VS2-SnS2/f-MWCNT hybrids. Herein, the binary metal sulfide (VS2-SnS2) was synthesized via single step hydrothermal route and hybrids with f-MWCNT via the ultrasonication process. The as-prepared VS2-SnS2/f-MWCNT hybrids were characterized through the FESEM, EDX and elemental mapping, TEM, XPS, Raman and XRD techniques. The electrochemical performance and catalytic activity of the modified electrodes were probed using electrochemical impedance spectra (EIS), cyclic voltammetry (CV) and differential pulse voltammetry (DPV). Interestingly, DPV results exhibits an appreciable linear range from 0.025 to 1017 µM and LOD of 0.008 µM. The selectivity study was performed to prove the high selectivity of the VS2-SnS2/f-MWCNT hybrids modified electrode. Furthermore, the practical applicability of the DA sensor was scrutinized in human serum sample and rat brain sample.


Assuntos
Dopamina/sangue , Nanotubos de Carbono/química , Neurotransmissores/sangue , Sulfetos/química , Compostos de Estanho/química , Compostos de Vanádio/química , Animais , Química Encefálica , Técnicas Eletroquímicas/instrumentação , Técnicas Eletroquímicas/métodos , Eletrodos , Humanos , Concentração de Íons de Hidrogênio , Limite de Detecção , Ratos , Reprodutibilidade dos Testes , Sulfetos/síntese química , Estanho/química , Compostos de Estanho/síntese química , Vanádio/química , Compostos de Vanádio/síntese química
12.
Forensic Sci Int ; 299: 34-40, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-30954005

RESUMO

We report a fatal γ-hydroxybutyric acid (GHB) intoxication of a forty-year old man. According to an acquaintances' statement, the deceased had drunk a beverage containing GHB approximately five hours before he was found. Postmortem GHB concentrations were determined using gas chromatography coupled to single quadrupole mass spectrometry after simple protein precipitation with methanol and derivatization with BSTFA (1% TMCS). Concentrations in body fluids and tissues of the deceased were as follows: cardiac blood 384 mg/L, femoral blood 358 mg/L, urine 864 mg/L, brain tissue 211 mg/kg, liver tissue 201 mg/kg, kidney tissue 492 mg/kg, bile 334 mg/L and gastric content 2025 mg/L. In an exhibit (liquid in a plastic bottle found next to the decedent) analyzed 29 days after the intake 27.6 g/L GHB were found with an increasing content during storage depending on the pH of the liquid (17 months after the intake: 70.0 g/L GHB and 121.2 g/L after adjusting the exhibit to a pH of 10 before extraction). GHB concentrations in head hair of the deceased (overall length approx. 4 cm, measured in segments of 0.5 cm) were measured using liquid chromatography coupled to triple quadrupole mass spectrometry. Concentrations in unwashed and washed hair samples were 91.9-174 ng/mg and 49.2-134 ng/mg, respectively. All cut-off values for postmortem matrices generally used for the identification of an exogenous GHB intake, which are further discussed within this publication, were exceeded. A lethal GHB intoxication can be assumed by a combination of toxicological findings, police investigations and exclusion of other causes of death.


Assuntos
Hidroxibutiratos/envenenamento , Adulto , Bebidas , Bile/química , Química Encefálica , Cromatografia Líquida , Toxicologia Forense , Cromatografia Gasosa-Espectrometria de Massas , Conteúdo Gastrointestinal/química , Cabelo/química , Humanos , Hidroxibutiratos/análise , Rim/química , Fígado/química , Masculino , Transtornos Relacionados ao Uso de Substâncias/complicações , Transtornos Relacionados ao Uso de Substâncias/diagnóstico
13.
Nat Protoc ; 14(5): 1509-1529, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-30962606

RESUMO

Here, we describe an optimized and detailed protocol for block-face serial microscopy tomography (FAST). FAST enables high-speed serial section fluorescence imaging of fixed brains at an axonal spatial resolution and subsequent image data processing. It renders brain-wide anatomical and functional analyses, including structural profiling of nuclear-stained brain at the single-cell level, cell-type-specific mapping with reporter animal brains and neuronal tracing with anterograde/retrograde labeling. Light-sheet fluorescence microscopy of cleared brains is advantageous in regard to imaging speed, but its spatial resolution is generally limited, whereas the opposite is true for conventional confocal microscopy. FAST offers a solution to overcome these technical limitations. This protocol describes detailed procedures for assembling the FAST hardware, sample preparation, imaging and image processing. A single imaging session takes as little as 2.4 h per mouse brain, and sample preparation requires 1 to several days, depending on pretreatments; however, multiple samples can be prepared simultaneously. We anticipate that FAST will contribute to unbiased and hypothesis-free approaches for a better understanding of brain systems.


Assuntos
Encéfalo/diagnóstico por imagem , Imagem Tridimensional/métodos , Microscopia de Fluorescência/métodos , Imagem Molecular/métodos , Tomografia/métodos , Animais , Química Encefálica/fisiologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL
14.
Nat Protoc ; 14(5): 1634-1660, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-30996262

RESUMO

In vitro reconstitutions of microtubule assemblies have provided essential mechanistic insights into the molecular bases of microtubule dynamics and their interactions with associated proteins. The tubulin code has emerged as a regulatory mechanism for microtubule functions, which suggests that tubulin isotypes and post-translational modifications (PTMs) play important roles in controlling microtubule functions. To investigate the tubulin code mechanism, it is essential to analyze different tubulin variants in vitro. Until now, this has been difficult, as most reconstitution experiments have used heavily post-translationally modified tubulin purified from brain tissue. Therefore, we developed a protocol that allows purification of tubulin with controlled PTMs from limited sources through cycles of polymerization and depolymerization. Although alternative protocols using affinity purification of tubulin also yield very pure tubulin, our protocol has the unique advantage of selecting for fully functional tubulin, as non-polymerizable tubulin is excluded in the successive polymerization cycles. It thus provides a novel procedure for obtaining tubulin with controlled PTMs for in vitro reconstitution experiments. We describe specific procedures for tubulin purification from adherent cells, cells grown in suspension cultures and single mouse brains. The protocol can be combined with drug treatment, transfection of cells before tubulin purification or enzymatic treatment during the purification process. The amplification of cells and their growth in spinner bottles takes ~13 d; the tubulin purification takes 6-7 h. The tubulin can be used in total internal reflection fluorescence (TIRF)-microscopy-based experiments or pelleting assays for the investigation of intrinsic properties of microtubules and their interactions with associated proteins.


Assuntos
Processamento de Proteína Pós-Traducional/genética , Tubulina (Proteína)/química , Tubulina (Proteína)/isolamento & purificação , Animais , Reatores Biológicos , Química Encefálica , Linhagem Celular , Células HeLa , Humanos , Camundongos , Polimerização , Tubulina (Proteína)/genética , Tubulina (Proteína)/metabolismo , Ultracentrifugação
15.
Anal Bioanal Chem ; 411(15): 3373-3382, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31016328

RESUMO

Titania-grafted magnetic sporopollenin is synthesized by the liquid-phase deposition (LPD) technique, characterized by SEM, EDX, and nitrogen adsorption porosimetry, and used for the selective enrichment of phosphorylated peptides. The material is low cost because of easier availability of pollens and has rich surface chemistry which enables strong attachment of titania onto magnetic sporopollenin. The material shows higher selectivity of 1:1000 with ß-casein spiked in BSA. Higher sensitivity of 10 fmol is recorded for phosphopeptides from standard ß-casein digest. Twenty phosphorylated peptides are enriched from milk digest and four endogenous phosphopeptides from diluted human serum. The magnetic property of titania-coated magnetic sporopollenin facilitates the fast and effective isolation of phosphopeptides from complex mixtures through external magnet. The material is finally applied to tryptic digest of rat brain cell lysate for phosphopeptide enrichment where 2718 phosphopeptides are identified by using LC-MS/MS with C18 column. Titania-coated magnetic sporopollenin captures both mono-phosphorylated (2489) and multi-phosphorylated peptides (229) due to strong affinity of TiO2 with phosphates. TiO2-coated magnetic material also shows better enrichment efficiency in comparison to commercial TiO2. Graphical abstract.


Assuntos
Biopolímeros/química , Química Encefálica , Carotenoides/química , Imãs/química , Fosfopeptídeos/análise , Titânio/química , Adsorção , Animais , Carotenoides/síntese química , Caseínas/química , Bovinos , Magnetismo/métodos , Fosfopeptídeos/isolamento & purificação , Ratos , Soroalbumina Bovina/química , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz/métodos
16.
Nat Protoc ; 14(6): 1708-1733, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-31028373

RESUMO

Understanding the structure-function relationships between diverse cell types in a complex organ environment requires detailed in situ reconstruction of cell-associated molecular properties in the context of 3D, macro-scale tissue architecture. We recently developed clearing-enhanced 3D (Ce3D), a simple and effective method for tissue clearing that achieves excellent transparency; preserves cell morphology, tissue architecture, and reporter molecule fluorescence; and is robustly compatible with direct immunolabeling. These characteristics permit high-quality multiplex fluorescence microscopy of large tissue volumes, as well as image analysis using advanced platforms such as volumetric histocytometry, collectively allowing quantitative characterization of cells with respect to their spatial positioning within tissues on the basis of phenotypic and functional markers. Ce3D clearing is fast, achieving robust transparency of most tissues within 24 h, albeit still necessitating additional time for staining, imaging, and analysis (1-2 weeks). Here, we provide detailed procedures for tissue clearing using Ce3D, including optimized workflows for tissue processing and staining, as well as treatment of difficult-to-clear organs such as the brain. We also describe a new procedure for RNA detection in Ce3D-treated tissues, as well as provide additional details for the volumetric histocytometry data processing steps. Finally, we discuss limitations and work-around strategies for improving antibody-based tissue immunolabeling, fluorophore multiplexing, large-volume microscopy, and computational analysis of large image datasets. Together, these detailed procedures and solutions for high-resolution volumetric microscopy with Ce3D enable quantitative visualization of cells and tissues at a high level of detail, allowing exploration of cellular spatial relationships in a variety of tissue settings.


Assuntos
Imagem Tridimensional/métodos , Microscopia Confocal/métodos , Microscopia de Fluorescência/métodos , Animais , Química Encefálica , Feminino , Imuno-Histoquímica/métodos , Masculino , Camundongos , RNA/análise , Coloração e Rotulagem/métodos , Fixação de Tecidos/métodos
17.
N Engl J Med ; 380(18): 1716-1725, 2019 05 02.
Artigo em Inglês | MEDLINE | ID: mdl-30969506

RESUMO

BACKGROUND: Chronic traumatic encephalopathy (CTE) is a neurodegenerative disease that has been associated with a history of repetitive head impacts. The neuropathological diagnosis is based on a specific pattern of tau deposition with minimal amyloid-beta deposition that differs from other disorders, including Alzheimer's disease. The feasibility of detecting tau and amyloid deposition in the brains of living persons at risk for CTE has not been well studied. METHODS: We used flortaucipir positron-emission tomography (PET) and florbetapir PET to measure deposition of tau and amyloid-beta, respectively, in the brains of former National Football League (NFL) players with cognitive and neuropsychiatric symptoms and in asymptomatic men with no history of traumatic brain injury. Automated image-analysis algorithms were used to compare the regional tau standardized uptake value ratio (SUVR, the ratio of radioactivity in a cerebral region to that in the cerebellum as a reference) between the two groups and to explore the associations of SUVR with symptom severity and with years of football play in the former-player group. RESULTS: A total of 26 former players and 31 controls were included in the analysis. The mean flortaucipir SUVR was higher among former players than among controls in three regions of the brain: bilateral superior frontal (1.09 vs. 0.98; adjusted mean difference, 0.13; 95% confidence interval [CI], 0.06 to 0.20; P<0.001), bilateral medial temporal (1.23 vs. 1.12; adjusted mean difference, 0.13; 95% CI, 0.05 to 0.21; P<0.001), and left parietal (1.12 vs. 1.01; adjusted mean difference, 0.12; 95% CI, 0.05 to 0.20; P = 0.002). In exploratory analyses, the correlation coefficients in these three regions between the SUVRs and years of play were 0.58 (95% CI, 0.25 to 0.79), 0.45 (95% CI, 0.07 to 0.71), and 0.50 (95% CI, 0.14 to 0.74), respectively. There was no association between tau deposition and scores on cognitive and neuropsychiatric tests. Only one former player had levels of amyloid-beta deposition similar to those in persons with Alzheimer's disease. CONCLUSIONS: A group of living former NFL players with cognitive and neuropsychiatric symptoms had higher tau levels measured by PET than controls in brain regions that are affected by CTE and did not have elevated amyloid-beta levels. Further studies are needed to determine whether elevated CTE-associated tau can be detected in individual persons. (Funded by Avid Radiopharmaceuticals and others.).


Assuntos
Encéfalo/patologia , Encefalopatia Traumática Crônica/patologia , Futebol Americano/lesões , Tomografia por Emissão de Pósitrons , Tauopatias/patologia , Proteínas tau/análise , Adulto , Idoso , Compostos de Anilina , Encéfalo/diagnóstico por imagem , Química Encefálica , Concussão Encefálica/complicações , Estudos de Casos e Controles , Encefalopatia Traumática Crônica/diagnóstico por imagem , Encefalopatia Traumática Crônica/etiologia , Transtornos Cognitivos/etiologia , Etilenoglicóis , Humanos , Masculino , Transtornos Mentais/etiologia , Pessoa de Meia-Idade , Placa Amiloide/diagnóstico por imagem , Compostos Radiofarmacêuticos , Tauopatias/diagnóstico por imagem
18.
N Engl J Med ; 380(15): 1408-1420, 2019 Apr 11.
Artigo em Inglês | MEDLINE | ID: mdl-30970186

RESUMO

BACKGROUND: Prodromal Alzheimer's disease offers an opportunity to test the effect of drugs that modify the deposition of amyloid in the brain before the onset of dementia. Verubecestat is an orally administered ß-site amyloid precursor protein-cleaving enzyme 1 (BACE-1) inhibitor that blocks production of amyloid-beta (Aß). The drug did not prevent clinical progression in a trial involving patients with mild-to-moderate dementia due to Alzheimer's disease. METHODS: We conducted a randomized, double-blind, placebo-controlled, 104-week trial to evaluate verubecestat at doses of 12 mg and 40 mg per day, as compared with placebo, in patients who had memory impairment and elevated brain amyloid levels but whose condition did not meet the case definition of dementia. The primary outcome was the change from baseline to week 104 in the score on the Clinical Dementia Rating Scale-Sum of Boxes (CDR-SB; scores range from 0 to 18, with higher scores indicating worse cognition and daily function). Secondary outcomes included other assessments of cognition and daily function. RESULTS: The trial was terminated for futility after 1454 patients had been enrolled; 485 had been assigned to receive verubecestat at a dose of 12 mg per day (the 12-mg group), 484 to receive verubecestat at a dose of 40 mg per day (the 40-mg group), and 485 to receive placebo. A total of 234 patients, 231 patients, and 239 patients per group, respectively, completed 104 weeks of the trial regimen. The estimated mean change from baseline to week 104 in the CDR-SB score was 1.65 in the 12-mg group, 2.02 in the 40-mg group, and 1.58 in the placebo group (P = 0.67 for the comparison between the 12-mg group and the placebo group and P = 0.01 for the comparison between the 40-mg group and the placebo group), suggesting a worse outcome in the higher-dose group than in the placebo group. The estimated rate of progression to dementia due to Alzheimer's disease was 24.5, 25.5, and 19.3 events per 100 patient-years in the 12-mg group, the 40-mg group, and the placebo group, respectively (hazard ratio for 40 mg vs. placebo, 1.38; 97.51% confidence interval, 1.07 to 1.79, not adjusted for multiple comparisons), favoring placebo. Adverse events were more common in the verubecestat groups than in the placebo group. CONCLUSIONS: Verubecestat did not improve clinical ratings of dementia among patients with prodromal Alzheimer's disease, and some measures suggested that cognition and daily function were worse among patients who received verubecestat than among those who received placebo. (Funded by Merck Sharp & Dohme; ClinicalTrials.gov number, NCT01953601.).


Assuntos
Doença de Alzheimer/prevenção & controle , Precursor de Proteína beta-Amiloide/antagonistas & inibidores , Disfunção Cognitiva/tratamento farmacológico , Óxidos S-Cíclicos/uso terapêutico , Tiadiazinas/uso terapêutico , Idoso , Peptídeos beta-Amiloides/análise , Química Encefálica , Disfunção Cognitiva/patologia , Óxidos S-Cíclicos/efeitos adversos , Progressão da Doença , Método Duplo-Cego , Inibidores Enzimáticos/efeitos adversos , Inibidores Enzimáticos/uso terapêutico , Feminino , Hipocampo/patologia , Humanos , Análise de Intenção de Tratamento , Imagem por Ressonância Magnética , Masculino , Tamanho do Órgão , Placa Amiloide/diagnóstico por imagem , Tomografia por Emissão de Pósitrons , Sintomas Prodrômicos , Tiadiazinas/efeitos adversos , Falha de Tratamento
19.
Nutrients ; 11(4)2019 Apr 11.
Artigo em Inglês | MEDLINE | ID: mdl-30978920

RESUMO

Harman and norharman, two neuroactive ß-carbolines, are present in several plants and in thermally processed foods. They exhibited a wide spectrum of biological and pharmacological effects, including antioxidant, neuroprotective, and anti-inflammatory effects. In this article, we review the progress of recent research on the presence of these compounds in food, as well as their various biological and neuroactive properties. Our findings strongly suggest that some foods, especially coffee, can act as a rich source of ß-carbolines, which may possibly be associated with a reduced risk for serious neurodegenerative diseases, such as Parkinson's and Alzheimer's.


Assuntos
Carbolinas/análise , Alimentos , Animais , Encéfalo/efeitos dos fármacos , Encéfalo/fisiologia , Química Encefálica , Carbolinas/administração & dosagem , Carbolinas/química , Carbolinas/farmacologia , Tremor Essencial/induzido quimicamente , Tremor Essencial/metabolismo , Manipulação de Alimentos , Harmina/administração & dosagem , Harmina/análogos & derivados , Harmina/análise , Humanos , Doenças Neurodegenerativas/induzido quimicamente , Doenças Neurodegenerativas/metabolismo , Fármacos Neuroprotetores , Estresse Oxidativo/efeitos dos fármacos , Doença de Parkinson/metabolismo , Extratos Vegetais/química
20.
Nutrients ; 11(4)2019 Apr 11.
Artigo em Inglês | MEDLINE | ID: mdl-30979047

RESUMO

Unhealthy diet promotes progression of metabolic disorders and brain dysfunction with aging. Green tea extracts (GTEs) have various beneficial effects and alleviate metabolic disorders. GTEs have neuroprotective effects in rodent models, but their effects against brain dysfunction in models of aging fed unhealthy diets are still unclear. Here, we showed that GTEs attenuate high-fat (HF) diet-induced brain dysfunction in senescence-accelerated mouse prone-8 (SAMP8), a murine model of senescence. SAMP8 mice were fed a control diet, HF diet, or HF diet with 0.5% GTEs (HFGT) for four months. The HF diet reduced memory retention and induced amyloid ß1-42 accumulation, whereas GTEs attenuated these changes. In HF diet-fed mice, lipid oxidative stress, assessed by malondialdehyde levels, was increased. The levels of proteins that promote synaptic plasticity, such as brain-derived neurotrophic factor (BDNF) and postsynaptic density protein 95 (PSD95), were reduced. These alterations related to brain dysfunction were not observed in HFGT diet-fed mice. Overall, our data suggest that GTEs intake might attenuate brain dysfunction in HF diet-fed SAMP8 mice by protecting synaptic plasticity as well as via anti-oxidative effects. In conclusion, GTEs might ameliorate unhealthy diet-induced brain dysfunction that develops with aging.


Assuntos
Encefalopatias/tratamento farmacológico , Dieta Hiperlipídica/efeitos adversos , Fármacos Neuroprotetores , Extratos Vegetais/administração & dosagem , Chá , Envelhecimento , Peptídeos beta-Amiloides/análise , Animais , Encéfalo/patologia , Química Encefálica , Encefalopatias/etiologia , Encefalopatias/fisiopatologia , Fator Neurotrófico Derivado do Encéfalo/análise , Disfunção Cognitiva/tratamento farmacológico , Disfunção Cognitiva/etiologia , Proteína 4 Homóloga a Disks-Large/análise , Masculino , Memória , Camundongos , Plasticidade Neuronal , Tamanho do Órgão , Estresse Oxidativo/efeitos dos fármacos , Fitoterapia , Sinaptofisina/análise
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