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1.
Am J Physiol Endocrinol Metab ; 318(6): E943-E955, 2020 06 01.
Artigo em Inglês | MEDLINE | ID: mdl-32369414

RESUMO

Myokines, such as irisin, have been purported to exert physiological effects on skeletal muscle in an autocrine/paracrine fashion. In this study, we aimed to investigate the mechanistic role of in vivo fibronectin type III domain-containing 5 (Fndc5)/irisin upregulation in muscle. Overexpression (OE) of Fndc5 in rat hindlimb muscle was achieved by in vivo electrotransfer, i.e., bilateral injections of Fndc5 harboring vectors for OE rats (n = 8) and empty vector for control rats (n = 8). Seven days later, a bolus of D2O (7.2 mL/kg) was administered via oral gavage to quantify muscle protein synthesis. After an overnight fast, on day 9, 2-deoxy-d-glucose-6-phosphate (2-DG6P; 6 mg/kg) was provided during an intraperitoneal glucose tolerance test (2 g/kg) to assess glucose handling. Animals were euthanized, musculus tibialis cranialis muscles and subcutaneous fat (inguinal) were harvested, and metabolic and molecular effects were evaluated. Muscle Fndc5 mRNA increased with OE (~2-fold; P = 0.014), leading to increased circulating irisin (1.5 ± 0.9 to 3.5 ± 1.2 ng/mL; P = 0.049). OE had no effect on protein anabolism or mitochondrial biogenesis; however, muscle glycogen was increased, along with glycogen synthase 1 gene expression (P = 0.04 and 0.02, respectively). In addition to an increase in glycogen synthase activation in OE (P = 0.03), there was a tendency toward increased glucose transporter 4 protein (P = 0.09). However, glucose uptake (accumulation of 2-DG6P) was identical. Irisin elicited no endocrine effect on mitochondrial biogenesis or uncoupling proteins in white adipose tissue. Hindlimb overexpression led to physiological increases in Fndc5/irisin. However, our data indicate limited short-term impacts of irisin in relation to muscle anabolism, mitochondrial biogenesis, glucose uptake, or adipose remodeling.


Assuntos
Fibronectinas/genética , Músculo Esquelético/metabolismo , Gordura Subcutânea/metabolismo , Animais , Desoxiglucose/metabolismo , Óxido de Deutério , Eletroporação , Fibronectinas/metabolismo , Expressão Gênica , Glucose/metabolismo , Teste de Tolerância a Glucose , Transportador de Glucose Tipo 4/genética , Glucose-6-Fosfato/análogos & derivados , Glucose-6-Fosfato/metabolismo , Glicogênio/metabolismo , Glicogênio Sintase/genética , Glicogênio Sintase/metabolismo , Membro Posterior , Masculino , Proteínas de Desacoplamento Mitocondrial/genética , Biogênese de Organelas , Biossíntese de Proteínas , RNA Mensageiro/metabolismo , Ratos
2.
J Phys Chem Lett ; 11(10): 4156-4162, 2020 May 21.
Artigo em Inglês | MEDLINE | ID: mdl-32370505

RESUMO

Excited state proton transfer (ESPT) is thought to be responsible for the photostability of biological molecules, including DNA and proteins, and natural dyes such as indigo. However, the mechanistic role of the solvent interaction in driving ESPT is not well understood. Here, the electronic excited state deactivation dynamics of indigo carmine (InC) is mapped by visible pump-infrared probe and two-dimensional electronic-vibrational (2DEV) spectroscopy and complemented by electronic structure calculations. The observed dynamics reveal notable differences between InC in a protic solvent, D2O, and an aprotic solvent, deuterated dimethyl sulfoxide (dDMSO). Notably, an acceleration in the excited state decay is observed in D2O (<10 ps) compared to dDMSO (130 ps). Our data reveals clear evidence for ESPT in D2O accompanied by a significant change in dipole moment, which is found not to occur in dDMSO. We conclude that the ability of protic solvents to form intermolecular H-bonds with InC enables ESPT, which facilitates a rapid nonradiative S1 → S0 transition via the monoenol intermediate.


Assuntos
Corantes/química , Óxido de Deutério/química , Dimetil Sulfóxido/química , Índigo Carmim/química , Prótons , Estrutura Molecular , Solventes/química
3.
Am J Physiol Endocrinol Metab ; 319(1): E175-E186, 2020 07 01.
Artigo em Inglês | MEDLINE | ID: mdl-32459526

RESUMO

Little is known about the effects of the development of metabolic syndrome (MS) on protein and amino acid (AA) metabolism. During this study, we took advantage of the variability in interindividual susceptibility to high fat diet-induced MS to study the relationships between MS, protein synthesis, and AA catabolism in multiple tissues in rats. After 4 mo of high-fat feeding, an MS score (ZMS) was calculated as the average of the z-scores for individual MS components [weight, adiposities, homeostasis model for the assessment of insulin resistance (HOMA-IR), and triglycerides]. In the small intestine, liver, plasma, kidneys, heart, and muscles, tissue protein synthesis was measured by 2H2O labeling, and we evaluated the proportion of tissue AA catabolism (relative to protein synthesis) and nutrient routing to nonindispensable AAs in tissue proteins using natural nitrogen and carbon isotopic distances between tissue proteins and nutrients (Δ15N and Δ13C), respectively. In the liver, protein mass and synthesis increased, whereas the proportion of AA catabolism decreased with ZMS. By contrast, in muscles, we found no association between ZMS and protein mass, protein synthesis (except for a weak positive association in the gastrocnemius muscle only), and proportion of AA catabolism. The development of MS was also associated with altered metabolic flexibility and fatty acid oxidation, as shown by less routing of dietary lipids to nonindispensable AA synthesis in liver and muscle. In conclusion, MS development is associated with a greater gain of both fat and protein masses, with higher protein anabolism that mainly occurs in the liver, whereas muscles probably develop anabolic resistance due to insulin resistance.


Assuntos
Aminoácidos/metabolismo , Dieta Hiperlipídica , Intestino Delgado/metabolismo , Rim/metabolismo , Fígado/metabolismo , Síndrome Metabólica/metabolismo , Músculo Esquelético/metabolismo , Miocárdio/metabolismo , Animais , Isótopos de Carbono , Óxido de Deutério , Gorduras na Dieta/metabolismo , Ácidos Graxos/metabolismo , Resistência à Insulina , Masculino , Isótopos de Nitrogênio , Obesidade/metabolismo , Plasma , Biossíntese de Proteínas , Proteínas/metabolismo , Ratos
4.
Nat Commun ; 11(1): 1454, 2020 03 19.
Artigo em Inglês | MEDLINE | ID: mdl-32193396

RESUMO

Enzymes dependent on nicotinamide cofactors are important components of the expanding range of asymmetric synthetic techniques. New challenges in asymmetric catalysis are arising in the field of deuterium labelling, where compounds bearing deuterium (2H) atoms at chiral centres are becoming increasingly desirable targets for pharmaceutical and analytical chemists. However, utilisation of NADH-dependent enzymes for 2H-labelling is not straightforward, owing to difficulties in supplying a suitably isotopically-labelled cofactor ([4-2H]-NADH). Here we report on a strategy that combines a clean reductant (H2) with a cheap source of 2H-atoms (2H2O) to generate and recycle [4-2H]-NADH. By coupling [4-2H]-NADH-recycling to an array of C=O, C=N, and C=C bond reductases, we demonstrate asymmetric deuteration across a range of organic molecules under ambient conditions with near-perfect chemo-, stereo- and isotopic selectivity. We demonstrate the synthetic utility of the system by applying it in the isolation of the heavy drug (1S,3'R)-[2',2',3'-2H3]-solifenacin fumarate on a preparative scale.


Assuntos
Biocatálise , Técnicas de Química Sintética/métodos , Deutério/química , Marcação por Isótopo/métodos , Oxirredutases/química , Óxido de Deutério/química , Estrutura Molecular , Niacinamida/química , Succinato de Solifenacina/química , Estereoisomerismo
5.
J Vis Exp ; (156)2020 02 20.
Artigo em Inglês | MEDLINE | ID: mdl-32150166

RESUMO

Body condition scoring systems and body condition indices are common techniques used for assessing the health status or fitness of a species. Body condition scoring systems are evaluator dependent and have the potential to be highly subjective. Body condition indices can be confounded by foraging, the effects of body weight, as well as statistical and inferential problems. An alternative to body condition scoring systems and body condition indices is using a stable isotope such as deuterium oxide to determine body composition. The deuterium oxide dilution method is a repeatable, quantitative technique used to estimate body composition in humans, wildlife, and domestic species. Additionally, the deuterium oxide dilution technique can be used to determine the water consumption of an individual animal. Here, we describe the adaption of the deuterium oxide dilution technique for assessing body composition in big brown bats (Eptesicus fuscus) and for assessing water consumption in cats (Felis catis).


Assuntos
Tecido Adiposo/química , Composição Corporal , Peso Corporal , Óxido de Deutério/química , Ingestão de Líquidos , Espectroscopia de Infravermelho com Transformada de Fourier/métodos , Animais , Gatos , Quirópteros
6.
Phys Chem Chem Phys ; 22(10): 5463-5475, 2020 Mar 11.
Artigo em Inglês | MEDLINE | ID: mdl-32096510

RESUMO

The calcium sensor protein calmodulin is ubiquitous among eukaryotes. It translates intracellular Ca2+ influx (by a decrease of conformational flexibility) into increased target recognition affinity. Here we demonstrate that by using the IR reporter -SCN in combination with 2D-IR spectroscopy, global structure changes and local dynamics, degree of solvent exposure and protein-ligand interaction can be characterised in great detail. The long vibrational lifetime of the -SCN label allows for centerline slope analysis of the 2D-IR line shape up to 120 ps to deduce the frequency-frequency correlation function (FFCF) of the -SCN label in various states and label positions in the protein. Based on that we show clear differences between a solvent exposed site, the environment close to the Ca2+ binding motif and three highly conserved positions for ligand binding. Furthermore, we demonstrate how these dynamics are affected by conformational change induced by the addition of Ca2+ ions and by interaction with a short helical peptide mimicking protein binding. We show that the binding mode is strongly heterogeneous among the probed key binding methionine residues. SCN's vibrational relaxation is dominated by intermolecular contributions. Changes in the vibrational lifetime upon changing between H2O and D2O buffer therefore provide a robust measure for water accessibility of the label. Characterising -SCN's extinction coefficient, vibrational lifetime in light and heavy water and its FFCF we demonstrate the vast potential it has as a label especially for nonlinear spectroscopies, such as 2D-IR spectroscopy.


Assuntos
Calmodulina/química , Espectrofotometria Infravermelho , Calmodulina/metabolismo , Óxido de Deutério/química , Ligação Proteica , Conformação Proteica , Domínios e Motivos de Interação entre Proteínas , Solventes/química , Vibração , Água/química
7.
J Photochem Photobiol B ; 204: 111787, 2020 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-31958676

RESUMO

The sensitivity for singlet oxygen (1O2) of two convenient 1O2 probes, 1,3-diphenylisobenzofuran (DPBF) and 9,10-Anthracenediyl-bis(methylene)dimalonic acid (ABDA), has been investigated in different aqueous environments. Both probes are commercially available at reasonable cost and can be used with standard UV-vis spectrometers. Although DPBF is not soluble in neat water and is not specific to the detection of 1O2, it has very high, essentially diffusion-limited, reactivity towards 1O2; it can trap up to 50% of all 1O2 created in alcohol/water or micellar solution, and even more when replacing H2O by D2O, which makes it highly useful when the process under investigation does not yield much 1O2. On the other hand, ABDA has a much lower reactivity, reacting with only 2% of the singlet oxygen generated in H2O, as well as a smaller extinction coefficient, resulting in a much smaller spectroscopic response, but is soluble in neat water and is specific for 1O2, allowing for discrimination from other reactive oxygen species. The results presented here not only allow a comparative assessment of the usefulness of the two 1O2 probes, but also provide a reference for an accurate absolute quantification of the amount of 1O2 generated in an experiment from the observed absorbance bleach.


Assuntos
Antracenos/química , Benzofuranos/química , Oxigênio Singlete/análise , Água/química , Óxido de Deutério/química , Luz , Solventes/química , Espectrofotometria
8.
Diabetes ; 69(3): 300-312, 2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-31806624

RESUMO

Branched esters of palmitic acid and hydroxystearic acid (PAHSA) are anti-inflammatory and antidiabetic lipokines that connect glucose and lipid metabolism. We aimed to characterize involvement of the 5-PAHSA regioisomer in the adaptive metabolic response of white adipose tissue (WAT) to cold exposure (CE) in mice, exploring the cross talk between glucose utilization and lipid metabolism. CE promoted local production of 5- and 9-PAHSAs in WAT. Metabolic labeling of de novo lipogenesis (DNL) using 2H2O revealed that 5-PAHSA potentiated the effects of CE and stimulated triacylglycerol (TAG)/fatty acid (FA) cycling in WAT through impacting lipogenesis and lipolysis. Adipocyte lipolytic products were altered by 5-PAHSA through selective FA re-esterification. The impaired lipolysis in global adipose triglyceride lipase (ATGL) knockout mice reduced free PAHSA levels and uncovered a metabolite reservoir of TAG-bound PAHSAs (TAG estolides) in WAT. Utilization of 13C isotope tracers and dynamic metabolomics documented that 5-PAHSA primes adipocytes for glucose metabolism in a different way from insulin, promoting DNL and impeding TAG synthesis. In summary, our data reveal new cellular and physiological mechanisms underlying the beneficial effects of 5-PAHSA and its relation to insulin action in adipocytes and independently confirm a PAHSA metabolite reservoir linked to ATGL-mediated lipolysis.


Assuntos
Adipócitos/metabolismo , Tecido Adiposo Branco/metabolismo , Glucose/metabolismo , Lipase/genética , Ácido Palmítico/metabolismo , Ácidos Esteáricos/metabolismo , Triglicerídeos/metabolismo , Animais , Isótopos de Carbono , Temperatura Baixa , Óxido de Deutério , Ácidos Graxos/metabolismo , Lipase/metabolismo , Lipogênese/genética , Lipólise , Metabolômica , Camundongos , Camundongos Knockout
9.
Br J Nutr ; 123(2): 232-240, 2020 01 28.
Artigo em Inglês | MEDLINE | ID: mdl-31623693

RESUMO

Saliva and urine are the two main body fluids sampled when breast milk intake is measured with the 2H oxide dose-to-mother technique. However, these two body fluids may generate different estimates of breast milk intake due to differences in isotope enrichment. Therefore, we aimed to assess how the estimated amount of breast milk intake differs when based on saliva and urine samples and to explore whether the total energy expenditure of the mothers is related to breast milk output. We used a convenience sample of thirteen pairs of mothers and babies aged 2-4 months, who were exclusively breastfed and apparently healthy. To assess breast milk intake, we administered doubly labelled water to the mothers and collected saliva samples from them, while simultaneously collecting both saliva and urine from their babies over a 14-d period. Isotope ratio MS was used to analyse the samples for 2H and 18O enrichments. Mean breast milk intake based on saliva samples was significantly higher than that based on urine samples (854·5 v. 812·8 g/d, P = 0·029). This can be attributed to slightly higher isotope enrichments in saliva and to a poorer model fit for urine samples as indicated by a higher square root of the mean square error (14·6 v. 10·4 mg/kg, P = 0·001). Maternal energy expenditure was not correlated with breast milk output. Our study suggests that saliva sampling generates slightly higher estimates of breast milk intake and is more precise as compared with urine and that maternal energy expenditure does not influence breast milk output.


Assuntos
Óxido de Deutério/administração & dosagem , Óxido de Deutério/urina , Fenômenos Fisiológicos da Nutrição do Lactente , Leite Humano , Saliva/química , Adulto , Água Corporal/química , Aleitamento Materno , Óxido de Deutério/análise , Metabolismo Energético , Feminino , Humanos , Técnicas de Diluição do Indicador , Lactente , Masculino , Espectrometria de Massas , Mães , Estado Nutricional , Isótopos de Oxigênio/análise , Isótopos de Oxigênio/urina
10.
Artigo em Inglês | MEDLINE | ID: mdl-31861542

RESUMO

BACKGROUND: Screening methods for childhood obesity are based largely on the published body mass index (BMI) criteria. Nonetheless, their accuracy in African children is largely unknown. The diagnostic accuracies of the World Health Organization (WHO), the Centers for Disease Control and Prevention (CDC), and the International Obesity Taskforce (IOTF) BMI-based criteria in defining obesity using deuterium dilution as a criterion method in a sample of Ghanaian children are presented. METHODS: Data on anthropometric indices and percent body fat were collected from 183 children aged 8-11 years. The sensitivity, specificity, and predictive values were calculated. The overall performance of the BMI criteria was evaluated using the receiver operating characteristics area under the curve (AUC). RESULTS: Overall sensitivity of WHO, CDC, and IOTF were 59.4% (40.6-76.3), 53.1% (34.7-70.9), and 46.9% (29.1-65.3) respectively. The overall specificity was high, ranging from 98.7% by WHO to 100.0% by IOTF. The AUC were 0.936 (0.865-1.000), 0.924 (0.852-0.995), and 0.945 (0.879-1.000) by the WHO, CDC, and IOTF criteria respectively for the overall sample. Prevalence of obesity by the WHO, CDC, IOTF, and deuterium oxide-derived percent body fat were 11.5%, 10.4%, 8.2%, and 17.5% respectively, with significant positive correlations between the BMI z-scores and percent body fat. CONCLUSIONS: The BMI-based criteria were largely specific but with moderate sensitivity in detecting excess body fat in Ghanaian children. To improve diagnostic accuracy, direct measurement of body fat and other health risk factors should be considered in addition to BMI.


Assuntos
Índice de Massa Corporal , Óxido de Deutério , Obesidade Pediátrica/diagnóstico , Tecido Adiposo , Antropometria , Criança , Estudos Transversais , Feminino , Gana/epidemiologia , Humanos , Masculino , Obesidade Pediátrica/epidemiologia , Prevalência , Curva ROC , Fatores de Risco , Sensibilidade e Especificidade
11.
Anal Chem ; 91(23): 15171-15178, 2019 12 03.
Artigo em Inglês | MEDLINE | ID: mdl-31687804

RESUMO

The metabolic activity of bacterial cells largely differentiates even within a clonal population. Such metabolic divergence among cells is thought to play an important role for phenotypic adaptation to ever-changing environmental conditions, such as antibiotic persistence. It has long been thought that persisters are in a state called dormancy, in which cells are metabolically inactive and do not grow. However, recent studies suggest that some types of persisters are not necessarily dormant, triggering a debate about the mechanisms of persisters. Here, we combined single-cell Raman imaging spectroscopy and D2O labeling to analyze metabolic activities of bacterial persister cells. Metabolically active cells uptake deuterium through metabolic processes and give distinct C-D Raman bands, which are direct indicators of metabolic activity. Using this imaging method, we characterized the metabolic activity of Mycobacterium smegmatis, a fast-growing model for Mycobacterium tuberculosis. We found that persister cells of M. smegmatis show certain metabolic activity and active cell growth in the presence of the antibiotic rifampicin. Interestingly, persistence is not correlated with growth rate prior to antibiotic exposure. These results show that dormancy is not responsible for the persistence of M. smegmatis cells against rifampicin, suggesting that the mechanism of persistence largely varies depending on the type of antibiotics and bacteria. Our results successfully demonstrate the potential of our perfusion-based single-cell D2O Raman imaging system for the analysis of the metabolic activity and growth of bacterial persister cells.


Assuntos
Óxido de Deutério/metabolismo , Mycobacterium smegmatis/citologia , Mycobacterium smegmatis/metabolismo , Mycobacterium tuberculosis/citologia , Mycobacterium tuberculosis/metabolismo , Análise de Célula Única , Antibacterianos/farmacologia , Óxido de Deutério/química , Testes de Sensibilidade Microbiana , Mycobacterium smegmatis/efeitos dos fármacos , Mycobacterium tuberculosis/efeitos dos fármacos , Rifampina/farmacologia , Análise Espectral Raman
12.
Ann Nutr Metab ; 75(2): 109-113, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31743917

RESUMO

An objective method of assessing breastfeeding practices is required to evaluate progress toward the World Health Organization Global Target 2025: to increase exclusive breastfeeding (EBF) rates in the first 6 months to at least 50% by 2025. Currently, assessment of EBF at the population level is based on mother or caregiver reporting, which risks recall and social desirability bias. A more objective method is the deuterium oxide dose to mother (DTM) technique, in which lactating mothers are given a small amount of deuterium-labeled water. The infant receives deuterium during breastfeeding, and a compartmental model is used to determine the amount of human milk consumed by the infant, and the exclusivity of breastfeeding practices. If the amount of human milk consumed by an infant is determined using the DTM technique and the concentration of nutritional components or potentially toxic contaminants is measured, then the infant's intake of essential nutrients or environmental contaminants can be ascertained.


Assuntos
Aleitamento Materno , Óxido de Deutério/farmacocinética , Ingestão de Alimentos , Leite Humano/química , Adulto , África ao Sul do Saara , Ásia , Peso Corporal , Óxido de Deutério/administração & dosagem , Óxido de Deutério/análise , Feminino , Infecções por HIV , Humanos , Lactente , Recém-Nascido , Masculino , México , Micronutrientes/análise , Resíduos de Praguicidas/análise , Saliva/química , Deficiência de Vitamina A/etiologia , Abastecimento de Água
13.
Sheng Li Xue Bao ; 71(5): 725-731, 2019 Oct 25.
Artigo em Chinês | MEDLINE | ID: mdl-31646326

RESUMO

The purpose of this study was to investigate the anti-injury effect and protective mechanism of hydrogen-enriched water in a rat model of acute liver injury induced by aflatoxin B1 (AFB1). Healthy male Sprague-Dawley (SD) rats were randomly divided into control group, model group (AFB1 group) and hydrogen-enriched water treatment group (AFB1+H2 group). The rat model of acute liver injury induced by AFB1 was established by single intragastric administration of AFB1 (2.0 mg/kg), and then the rats were treated with hydrogen-enriched water intragastrically. HE staining was used to observe the pathological changes of liver tissue. Blood samples were taken from vena cava to measure serum liver function indexes. Live tissue was sampled to detect malondialdehyde (MDA) and reduced glutathione (GSH) contents. Western blot was used to detect phosphorylation levels of MAPK signaling pathway proteins (ERK, JNK and p38 MAPK). The results showed that, compared with the AFB1 group, the AFB1+H2 group exhibited increased body weights, alleviated acute liver injury, decreased activities of serum glutamic-pyruvic transaminase and glutamic oxaloacetic transaminase, as well as total bilirubin level in the serum. Meanwhile, hydrogen-enriched water decreased MDA content and increased GSH content in liver tissue. AFB1-increased phosphorylation levels of ERK, JNK and p38 MAPK in liver tissue were down-regulated significantly by hydrogen-enriched water treatment. These results suggest that hydrogen-enriched water can alleviate liver injury induced by AFB1, and its mechanism may be related to the reduction of oxidative stress and the inhibition of MAPK signal transduction pathway activation.


Assuntos
Aflatoxina B1 , Doença Hepática Induzida por Substâncias e Drogas/prevenção & controle , Óxido de Deutério/uso terapêutico , Animais , Doença Hepática Induzida por Substâncias e Drogas/patologia , Fígado/patologia , Sistema de Sinalização das MAP Quinases , Masculino , Estresse Oxidativo , Ratos , Ratos Sprague-Dawley
14.
Nat Commun ; 10(1): 4094, 2019 09 25.
Artigo em Inglês | MEDLINE | ID: mdl-31554785

RESUMO

The hydrogen isotope deuterium is widely used in the synthesis of isotopically-labeled compounds and in the fabrication of semiconductors and optical fibers. However, the facile production of deuterium gas (D2) and hydrogen deuteride (HD) in a controlled manner is a challenging task, and rational heterogeneously-catalyzed protocols are still lacking. Herein, we demonstrate the selective production of hydrogen isotope compounds from a combination of formic acid and D2O, through cooperative action by a PdAg nanocatalyst on a silica substrate whose surface is modified with amine groups. In this process, D2 is predominantly evolved by the assist of weakly basic amine moieties, while nanocatalyst particles in the vicinity of strongly basic amine groups promote the preferential formation of HD. Kinetic data and calculations based on semi-classically corrected transition state theory coupled with density functional theory suggest that quantum tunneling dominates the hydrogen/deuterium exchange reaction over the metallic PdAg surfaces.


Assuntos
Formiatos/química , Hidrogênio/farmacologia , Catálise , Preparações de Ação Retardada/farmacologia , Deutério/química , Óxido de Deutério/química , Hidrogenação , Nanopartículas/química , Água/química
15.
Chem Phys Lipids ; 225: 104816, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31525381

RESUMO

To optimize dermal and transdermal administration of drugs, the barrier function of the skin, particularly the stratum corneum (SC), needs to be reduced reversibly. For this purpose, penetration/diffusion enhancers such as DMSO can be applied. However, there is the question whether DMSO is an aggressive penetration/diffusion enhancer in pharmaceutical and cosmetical relevant concentrations? Until now, it is unclear if this penetration/diffusion enhancement is caused by an interaction with the SC lipid matrix or related to effects within the corneocytes. Therefore, the effects of the hydrophilic enhancer DMSO on SC models with different dimensionality ranging from bilayers (liposomes) via oligo-layers to multilayers have been investigated in this study. The effects of DMSO should be compared to that of other relevant hydrophilic enhancers such as urea and taurine. An innovative spectrum of methods was applied to ascertain the mode of action of DMSO in relevant concentrations on a molecular scale. The experiments reveal that there is no specific interaction of 10% and 30% DMSO solutions with the SC model systems. Hence, if DMSO is applied in pharmaceutically and cosmetically relevant concentrations, it has no influence on the SC model systems used. Neither an additional water uptake in the head group region nor a decrease of the lipid chain packing density have been observed. The leakage studies on liposomes show that 10% DMSO is causing just a very slight leakage of 8%, lower than the leakage of 19.4% caused by 10% urea (Müller et al., 2016). Consequently, the interactions of DMSO with the SC model lipids used are very low in concentrations of 10% and 30%, respectively. Since the lipid composition in native SC lipid matrix is far more complex than this model mixture, the results can not be directly transferred to the native SC lipid matrix. However, the outcome of this study, together with various findings in the literature give rise to the assumption that the enhancing effect of DMSO concerning the diffusion of relevant hydrophilic drugs and actives appears to be realized via the corneocytes.


Assuntos
Dimetil Sulfóxido/química , Lipídeos/química , Óxido de Deutério/química , Difusão , Concentração de Íons de Hidrogênio , Interações Hidrofóbicas e Hidrofílicas , Lipossomos/química , Modelos Moleculares , Tamanho da Partícula , Propriedades de Superfície
16.
Anal Chem ; 91(17): 11063-11069, 2019 09 03.
Artigo em Inglês | MEDLINE | ID: mdl-31397558

RESUMO

In metabolomics, nuclear magnetic resonance (NMR) spectroscopy allows to identify and quantify compounds in biological samples. The sample preparation generally requires only few steps; however, an indispensable factor is the addition of a locking substance into the biofluid sample, such as deuterium oxide (D2O). While creatinine loss in pure D2O is well-described, the effects of different D2O concentrations on the signal profile of biological samples are unknown. In this work, we investigated the effect of D2O levels in the NMR buffer system in urine samples, in dependence on dwell time and temperature exposition. We reveal a decrease of the urinary creatinine peak area up to 35% after 24 h of dwell time at room temperature (RT) using 25% (v/v) D2O, but only 4% loss using 2.5% D2O. 1H, inverse-gated (IG) 13C, DEPT-HSQC NMR, and mass spectrometry (MS) experiments confirmed a proton-deuterium (H/D) exchange at the CH2. This leads to underestimation of creatinine levels and has an extensive effect when creatinine is used for normalization. This work offers a sample stability examination, depending on the D2O concentration, dwell time, and temperature and enables a method to correct for the successive loss. We propose an equation to correct the creatinine loss for samples prepared with various D2O concentrations and storage temperatures for dwell times up to 24 h. The correction function was validated against an external data set with n = 26 samples. To ensure sufficient creatinine stability in future studies, we suggest that a maximum of 10% D2O should be used at 4 °C or 2.5% D2O at RT, respectively.


Assuntos
Creatinina/urina , Óxido de Deutério/urina , Espectroscopia de Ressonância Magnética/normas , Metabolômica/normas , Artefatos , Medição da Troca de Deutério , Humanos , Guias de Prática Clínica como Assunto , Temperatura
17.
Scand Cardiovasc J ; 53(6): 329-336, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31455109

RESUMO

Objectives. Although deuterium oxide (D2O) has preservative property on the extracted organ, whether D2O also protects the in situ myocardial injury remains unknown. Using cardiac microdialysis, local administration of D2O through dialysis probe was applied in situ rat heart. We examined the effect of the D2O on the myocardial injury induced ischemia, reperfusion, and chemical hypoxia. Methodology. We measured dialysate myoglobin levels during 30 min of coronary occlusion and reperfusion in the absence and presence of D2O. Furthermore, to confirm the effect of D2O on NaCN induced myocardial injury, we measured the dialysate myoglobin levels with local perfusion of NaCN in the absence and presence of D2O. Results. The dialysate myoglobin levels increased from 177 ± 45 ng/mL at baseline to 3030 ± 1523 ng/mL during 15-30 min of coronary occlusion and further increased to 8588 ± 1684ng/mL at 0-15 min of reperfusion. The dialysate myoglobin levels with 60 min local perfusion of NaCN increased to 1214 ± 279 ng/mL. D2O attenuated myocardial myoglobin release during 15-30 min of coronary occlusion and 0-30 min of reperfusion and 15-60 min of local perfusion of NaCN. Conclusions. D2O might have a beneficial effect of myocardium against ischemia, reperfusion and chemical hypoxia.


Assuntos
Óxido de Deutério/farmacologia , Cardiopatias/prevenção & controle , Traumatismo por Reperfusão Miocárdica/prevenção & controle , Miocárdio/patologia , Animais , Modelos Animais de Doenças , Cardiopatias/induzido quimicamente , Cardiopatias/metabolismo , Cardiopatias/patologia , Masculino , Traumatismo por Reperfusão Miocárdica/metabolismo , Traumatismo por Reperfusão Miocárdica/patologia , Miocárdio/metabolismo , Mioglobina/metabolismo , Ratos Sprague-Dawley , Cianeto de Sódio , Fatores de Tempo
18.
Lab Chip ; 19(15): 2598-2609, 2019 08 07.
Artigo em Inglês | MEDLINE | ID: mdl-31259340

RESUMO

Early reaction intermediates in protein folding, such as those resulting in ß-amyloid formation due to transient misfolding, emerge within a few hundred microseconds. Here, we report a method to obtain sub-millisecond temporal resolution and molecular structural information of protein (mis-)folding events by using a microfluidic continuous-flow mixer (MCFM) in combination with Fourier transform infrared (FT-IR) imaging. The MCFMs are made out of cyclic olefin copolymer (COC) films, because this approach allows for rapid prototyping of different mixer designs. Furthermore, COC offers high IR transparency between 1500 and 2500 cm-1, thus maximizing the signal to noise ratio of the IR data obtained from a sample of interest. By combining narrow and wide channel widths in MCFM design, the platform provides fast mixing (460 µs) to induce protein (mis-)folding, and it maximizes the residence time in the observing area, so a wide range of reaction timescales can be captured in a single image. We validated the platform for its ability to induce and observe sub-millisecond processes by studying two systems: (i) the mixing of H2O and D2O and (ii) the mixing induced deprotonation of carboxylic acid. First, we observed excellent agreement between simulated and experimental data of the on-chip mixing of H2O and D2O, which verifies the distance-reaction time relationships based on simulation. Second, deprotonation of carboxylic acid by on-chip mixing with sodium hydroxide solution validates the ability of the platform to induce rapid pH jump that is needed for some biomolecular reactions. Finally, we studied the methanol-induced partial-unfolding of ubiquitin to show that our platform can be used to study biomolecular events 'on-pathway' using FT-IR imaging. We successfully extracted kinetic and structural details of the conformational changes along the channel. Our results are in agreement with prior studies that required more elaborate stopped flow approaches to acquire data for different time points. In summary, the reported method uses an easy-to-fabricate microfluidic mixer platform integrated with hyperspectral FT-IR imaging for rapid acquisition of structural details and kinetic parameters of biomolecular reactions. This approach does not need stopped flow or molecular imaging probes, as required respectively for alternative FT-IR spectroscopy and fluorescence approaches.


Assuntos
Dispositivos Lab-On-A-Chip , Imagem Molecular/instrumentação , Espectroscopia de Infravermelho com Transformada de Fourier , Ácido Acético/química , Óxido de Deutério/química , Concentração de Íons de Hidrogênio , Conformação Proteica , Desdobramento de Proteína , Razão Sinal-Ruído , Ubiquitina/química
19.
PLoS One ; 14(7): e0219563, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31291373

RESUMO

INTRODUCTION: Wrist-mounted motion sensors can quantify the volume and intensity of physical activities, but little is known about their long-term validity. Our aim was to validate a wrist motion sensor in estimating daily energy expenditure, including any change induced by long-term participation in endurance and strength training. Supplemental heart rate monitoring during weekly exercise was also investigated. METHODS: A 13-day doubly labeled water (DLW) measurement of total energy expenditure (TEE) was performed twice in healthy male subjects: during two last weeks of a 12-week Control period (n = 15) and during two last weeks of a 12-week combined strength and aerobic Training period (n = 13). Resting energy expenditure was estimated using two equations: one with body weight and age, and another one with fat-free mass. TEE and activity induced energy expenditure (AEE) were determined from motion sensor alone, and from motions sensor combined with heart rate monitor, the latter being worn during exercise only. RESULTS: When body weight and age were used in the calculation of resting energy expenditure, the motion sensor data alone explained 78% and 62% of the variation in TEE assessed by DLW at the end of Control and Training periods, respectively, with a bias of +1.75 (p <.001) and +1.19 MJ/day (p = .002). When exercise heart rate data was added to the model, the combined wearable device approach explained 85% and 70% of the variation in TEE assessed by DLW with a bias of +1.89 and +1.75 MJ/day (p <.001 for both). While significant increases in TEE and AEE were detected by all methods as a result of participation in regular training, motion sensor approach underestimated the change measured by DLW: +1.13±0.66 by DLW, +0.59±0.69 (p = .004) by motion sensor, and +0.98±0.70 MJ/day by combination of motion sensor and heart rate. Use of fat-free mass in the estimation of resting energy expenditure removed the biases between the wearable device estimations and the golden standard reference method of TEE and demonstrated a training-induced increase in resting energy expenditure by +0.18±0.13 MJ/day (p <.001). CONCLUSIONS: Wrist motion sensor combined with a heart rate monitor during exercise sessions, showed high agreement with the golden standard measurement of daily TEE and its change induced by participation in a long-term training protocol. The positive findings concerning the validity, especially the ability to follow-up the change associated with a lifestyle modification, can be considered significant because they partially determine the feasibility of wearable devices as quantifiers of health-related behavior.


Assuntos
Metabolismo Energético/fisiologia , Monitorização Fisiológica/métodos , Condicionamento Físico Humano/fisiologia , Resistência Física/fisiologia , Acelerometria/instrumentação , Adulto , Peso Corporal , Óxido de Deutério/análise , Ingestão de Líquidos/fisiologia , Frequência Cardíaca/fisiologia , Humanos , Estilo de Vida , Masculino , Monitorização Fisiológica/instrumentação , Água/química , Dispositivos Eletrônicos Vestíveis , Punho , Adulto Jovem
20.
Obes Surg ; 29(10): 3299-3308, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31230202

RESUMO

OBJECTIVE: To investigate the influence of changes in energy balance and body composition on the rate of weight loss throughout 1 year of Roux-en-Y gastric bypass. METHODS: Variables were collected pre-, 6, and 12 months (M) post-surgery from 18 women (BMI ≥ 40 and ≤ 50 kg m-2, 20 to 45 years). Total energy expenditure (TEEm), fat-free mass (FFM), and fat mass (FM) were measured by doubly labeled water. Self-reported energy intake (EIsr) was obtained from three non-consecutive food diaries. Metabolic adaptation was assessed via deviations from TEE predictive equation, and the calculated energy intake (EIc) via the sum of TEE and change in body stores. RESULTS: BMI significantly decreased (mean ± SD) from 45 ± 2 kg m-2 to 32 ± 3 kg m-2 at 6 M, and to 30 ± 3 kg m-2 at 12 M after surgery. The TEEm reduced significantly at both time points when compared with pre-surgery (6 M: - 612 ± 317 kcal day-1; 12 M: - 447 ± 516 kcal day-1). At 6 M, a metabolic adaptation was observed and the energy balance was - 1151 ± 195 kcal day-1, while at 12 M it was - 332 ± 158 kcal day-1. Changes in the values of TEEm were associated with changes in body weight at 12 M post-surgery. A significant underreporting was observed for EIsr (1057 ± 385 kcal day-1) vs. EIc (2083 ± 309 kcal day-1) at 12 M post-operative. CONCLUSION: The higher rate of weight loss at 6 M post-surgery was a response to energy imbalance, which was caused by high restriction in energy intake even with the presence of metabolic adaptation at this time. The EIsr was not sufficiently accurate to assess the energy consumption of this population. REGISTRATION OF CLINICAL TRIALS (OBSERVATIONAL STUDY): Brazilian Clinical Trials Registry: RBR-8k5jsj. Universal Trial Number: U1111-1206-0858.


Assuntos
Metabolismo Energético/fisiologia , Derivação Gástrica , Perda de Peso/fisiologia , Adulto , Óxido de Deutério , Humanos , Estudos Longitudinais , Pessoa de Meia-Idade , Adulto Jovem
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