RESUMEN
PURPOSE: Metabolite-specific balanced SSFP (MS-bSSFP) sequences are increasingly used in hyperpolarized [1-13C]Pyruvate (HP 13C) MRI studies as they improve SNR by refocusing the magnetization each TR. Currently, pharmacokinetic models used to fit conversion rate constants, kPL and kPB, and rate constant maps do not account for differences in the signal evolution of MS-bSSFP acquisitions. METHODS: In this work, a flexible MS-bSSFP model was built that can be used to fit conversion rate constants for these experiments. The model was validated in vivo using paired animal (healthy rat kidneys n = 8, transgenic adenocarcinoma of the mouse prostate n = 3) and human renal cell carcinoma (n = 3) datasets. Gradient echo (GRE) acquisitions were used with a previous GRE model to compare to the results of the proposed GRE-bSSFP model. RESULTS: Within simulations, the proposed GRE-bSSFP model fits the simulated data well, whereas a GRE model shows bias because of model mismatch. For the in vivo datasets, the estimated conversion rate constants using the proposed GRE-bSSFP model are consistent with a previous GRE model. Jointly fitting the lactate T2 with kPL resulted in less precise kPL estimates. CONCLUSION: The proposed GRE-bSSFP model provides a method to estimate conversion rate constants, kPL and kPB, for MS-bSSFP HP 13C experiments. This model may also be modified and used for other applications, for example, estimating rate constants with other hyperpolarized reagents or multi-echo bSSFP.
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Isótopos de Carbono , Imagen por Resonancia Magnética , Ácido Pirúvico , Animales , Ácido Pirúvico/farmacocinética , Ácido Pirúvico/metabolismo , Ratas , Imagen por Resonancia Magnética/métodos , Ratones , Isótopos de Carbono/farmacocinética , Humanos , Masculino , Riñón/diagnóstico por imagen , Riñón/metabolismo , Simulación por Computador , Neoplasias de la Próstata/diagnóstico por imagen , Neoplasias de la Próstata/metabolismo , Procesamiento de Imagen Asistido por Computador/métodos , Algoritmos , Relación Señal-Ruido , Neoplasias Renales/diagnóstico por imagen , Neoplasias Renales/metabolismo , Ratones TransgénicosRESUMEN
OBJECTIVE: Hyperpolarized [1-13C]-pyruvate magnetic resonance imaging is an emerging metabolic imaging method that offers unprecedented spatiotemporal resolution for monitoring tumor metabolism in vivo. To establish robust imaging biomarkers of metabolism, we must characterize phenomena that may modulate the apparent pyruvate-to-lactate conversion rate (kPL). Here, we investigate the potential effect of diffusion on pyruvate-to-lactate conversion, as failure to account for diffusion in pharmacokinetic analysis may obscure true intracellular chemical conversion rates. METHODS: Changes in hyperpolarized pyruvate and lactate signal were calculated using a finite-difference time domain simulation of a two-dimensional tissue model. Signal evolution curves with intracellular kPL values from 0.02 to 1.00 s-1 were analyzed using spatially invariant one-compartment and two-compartment pharmacokinetic models. A second spatially variant simulation incorporating compartmental instantaneous mixing was fit with the same one-compartment model. RESULTS: When fitting with the one-compartment model, apparent kPL underestimated intracellular kPL by approximately 50% at an intracellular kPL of 0.02 s-1. This underestimation increased for larger kPL values. However, fitting the instantaneous mixing curves showed that diffusion accounted for only a small part of this underestimation. Fitting with the two-compartment model yielded more accurate intracellular kPL values. SIGNIFICANCE: This work suggests diffusion is not a significant rate-limiting factor in pyruvate-to-lactate conversion given that our model assumptions hold true. In higher order models, diffusion effects may be accounted for by a term characterizing metabolite transport. Pharmacokinetic models used to analyze hyperpolarized pyruvate signal evolution should focus on carefully selecting the analytical model for fitting rather than accounting for diffusion effects.
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Imagen por Resonancia Magnética , Ácido Pirúvico , Ácido Pirúvico/análisis , Ácido Pirúvico/farmacocinética , Isótopos de Carbono/farmacocinética , Imagen por Resonancia Magnética/métodos , Simulación por Computador , Ácido LácticoRESUMEN
Isotope tracing has helped to determine the metabolic activities of organs. Methods to probe metabolic heterogeneity within organs are less developed. We couple stable-isotope-labeled nutrient infusion to matrix-assisted laser desorption ionization imaging mass spectrometry (iso-imaging) to quantitate metabolic activity in mammalian tissues in a spatially resolved manner. In the kidney, we visualize gluconeogenic flux and glycolytic flux in the cortex and medulla, respectively. Tricarboxylic acid cycle substrate usage differs across kidney regions; glutamine and citrate are used preferentially in the cortex and fatty acids are used in the medulla. In the brain, we observe spatial gradations in carbon inputs to the tricarboxylic acid cycle and glutamate under a ketogenic diet. In a carbohydrate-rich diet, glucose predominates throughout but in a ketogenic diet, 3-hydroxybutyrate contributes most strongly in the hippocampus and least in the midbrain. Brain nitrogen sources also vary spatially; branched-chain amino acids contribute most in the midbrain, whereas ammonia contributes in the thalamus. Thus, iso-imaging can reveal the spatial organization of metabolic activity.
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Encéfalo/metabolismo , Isótopos de Carbono/farmacocinética , Riñón/metabolismo , Isótopos de Nitrógeno/farmacocinética , Animales , Dieta , Enzimas , Gluconeogénesis , Ácido Glutámico/biosíntesis , Glucólisis , Masculino , Ratones Endogámicos C57BL , Imagen Molecular , Análisis de la Célula Individual , Espectrometría de Masa por Láser de Matriz Asistida de Ionización Desorción , Espectrometría de Masas en Tándem , Ácidos Tricarboxílicos/metabolismo , Flujo de TrabajoRESUMEN
Oleoylethanolamide is an endogenous molecule with neuroprotective effects. It has been reported that exogenous oleoylethanolamide can be administered therapeutically, but the confounding presence of the endogenous molecule has led to conflicting reports regarding the mechanisms of the effects and highlights a need for an adequate methodology to differentiate them. We have developed a liquid chromatography-tandem mass spectrometry method to study oleoylethanolamide in rat plasma and brain using a 13 C-labeled isotope, 13 C-oleoylethanolamide. 13 C-oleoylethanolamide was extracted using a liquid-liquid extraction employing acetonitrile and tert-butyl methyl ether (1:4). Analysis was performed using a gradient with a total run time of 12 min. 13 C-oleoylethanolamide, d4 -oleoylethanolamide (internal standard), and 12 C-oleoylethanolamide (endogenous background) eluted simultaneously at 1.64 min. The method was validated for specificity, sensitivity, accuracy, and precision and found to be capable of quantification within acceptable limits of ±15% over the calibration range of 0.39-25 ng/mL for the plasma and 1.17-75 ng/g for the brain. It was then applied to quantify 13 C-oleoylethanolamide over 90 min after intravenous administration of a solution (1 mg/kg) in rats. Results suggest that 13 C-oleoylethanolamide does not reach therapeutic concentrations in the brain, despite a relatively prolonged plasma circulation, suggesting that rapid degradation in the brain remains an obstacle to its clinical application to neurological disease.
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Encéfalo/metabolismo , Cromatografía Liquida/métodos , Etanolamina , Ácidos Oléicos , Plasma/metabolismo , Animales , Isótopos de Carbono/farmacocinética , Cromatografía Líquida de Alta Presión/métodos , Etanolamina/análisis , Etanolamina/farmacocinética , Extracción Líquido-Líquido/métodos , Ácidos Oléicos/análisis , Ácidos Oléicos/farmacocinética , Ratas , Reproducibilidad de los Resultados , Espectrometría de Masas en Tándem/métodosRESUMEN
Immune-based metabolic reprogramming of arginine utilization in the brain contributes to the neuronal pathology associated with Alzheimer's disease (AD). To enable our long-term goals of differentiation of AD mouse model genotypes, ages, and sexes based on activity of this pathway, we describe here the novel dosing (using uniformly labeled (13C615N4) arginine) and analysis methods using capillary electrophoresis high-resolution accurate-mass mass spectrometry for isotope tracing of metabolic products of arginine. We developed a pseudoprimed infusion-dosing regimen, using repeated injections, to achieve a steady state of uniformly labeled arginine in 135-195 min post bolus dose. Incorporation of stable isotope labeled carbon and nitrogen from uniformly labeled arginine into a host of downstream metabolites was measured in vivo in mice using serially sampled dried blood spots from the tail. In addition to the dried blood spot time course samples, total isotope incorporation into arginine-related metabolites was measured in the whole brain and plasma after 285 min. Preliminary demonstration of the technique identified differences isotope incorporation in arginine metabolites between male and female mice in a mouse-model of sporadic Alzheimer's disease (APOE4/huNOS2). The technique described herein will permit arginine pathway activity differentiation between mouse genotypes, ages, sexes, or drug treatments in order to elucidate the contribution of this pathway to Alzheimer's disease.
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Enfermedad de Alzheimer/metabolismo , Arginina/análisis , Electroforesis Capilar/métodos , Espectrometría de Masas/métodos , Enfermedad de Alzheimer/sangre , Animales , Apolipoproteína E4/genética , Arginina/sangre , Arginina/química , Encéfalo/metabolismo , Isótopos de Carbono/análisis , Isótopos de Carbono/farmacocinética , Modelos Animales de Enfermedad , Femenino , Humanos , Marcaje Isotópico , Masculino , Ratones Transgénicos , Óxido Nítrico Sintasa de Tipo II/genética , Isótopos de Nitrógeno/análisis , Isótopos de Nitrógeno/farmacocinética , Prueba de Estudio ConceptualRESUMEN
There is inconsistent information regarding the size effects of exogenously given hyaluronan on its in vivo fate. The data are often biased by the poor quality of hyaluronan and non-ideal labelling strategies used for resolving exogenous/endogenous hyaluronan, which only monitor the label and not hyaluronan itself. To overcome these drawbacks and establish the pharmacokinetics of intravenous hyaluronan in relation to its Mw, 13C-labelled HA of five Mws from 13.6-1562 kDa was prepared and administered to mice at doses 25-50 mg kg-1. The elimination efficiency increased with decreasing Mw. Low Mw hyaluronan was rapidly eliminated as small hyaluronan fragments in urine, while high Mw hyaluronan exhibited saturable kinetics and complete metabolization within 48 h. All tested Mws exhibited a similar uptake by liver cells and metabolization into activated sugars. 13C-labelling combined with LC-MS provides an excellent approach to elucidating in vivo fate and biological activities of hyaluronan.
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Ácido Hialurónico/farmacocinética , Marcaje Isotópico/métodos , Administración Intravenosa , Animales , Huesos/metabolismo , Isótopos de Carbono/química , Isótopos de Carbono/metabolismo , Isótopos de Carbono/farmacocinética , Cartílago/metabolismo , ADP-Ribosa Cíclica/metabolismo , Vías de Eliminación de Fármacos , Femenino , Ácido Hialurónico/química , Ácido Hialurónico/metabolismo , Ratones Endogámicos BALB C , Peso Molecular , Distribución Tisular , Uridina Difosfato Glucosa/metabolismo , Uridina Difosfato N-Acetilglucosamina/metabolismoRESUMEN
Imaging methods for hyperpolarized (HP) 13C agents must sample the evolution of signal from multiple agents with distinct chemical shifts within a very brief timeframe (typically < 1 min), which is challenging using conventional imaging methods. In this work, we compare two of the most commonly used HP spectroscopic imaging methods, spectral-spatial selective excitation and multi-echo chemical shift encoding (CSE, also referred to as IDEAL), for a typical preclinical HP [1-13C]pyruvate imaging scan at 7 T. Both spectroscopic encoding techniques were implemented and validated in HP experiments imaging enzyme phantoms and the murine kidney. SNR performance of these two spectroscopic imaging approaches was compared in numerical simulations and phantom experiments using a single-shot flyback EPI readout for spatial encoding. With identical effective excitation angles, the SNR of images acquired with spectral-spatial excitations and CSE were found to be effectively equivalent.
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Isótopos de Carbono/farmacocinética , Imagen por Resonancia Magnética/métodos , Ácido Pirúvico/farmacocinética , Animales , Imagen Eco-Planar/métodos , Procesamiento de Imagen Asistido por Computador , Riñón/diagnóstico por imagen , Ratones , Fantasmas de Imagen , Relación Señal-RuidoRESUMEN
The isotopic composition of baleen whales' epidermis structural layers can give information about dietary change over time. This study investigated if epidermis layers integrated isotopic values that record physiological changes from gestation to lactation. Epidermis tissues (n = 43) were collected from free ranging lactating female gray whale and calves during the beginning of three breeding seasons. Modelling of δ13C and δ15N values show intra- and inter-individual differences based on epidermal layers, age class and year of sampling. The isotopic composition of mother-calf pairs is correlated, and the estimates of the maximum mother-to-calf isotopic difference was ~1.4 for δ13C and between 1 and 1.5 for δ15N values. Change in δ15N values among epidermal layers in calves was associated with the transition from fetus to consumption of maternal milk. It is here proposed when lactation influences calf epidermis, δ15N values decrease consistently from the outermost to the innermost layer. However, if a calf was born only few days before collection, epidermis integrates more variable δ15N patterns because gestation still affects the isotopic composition of the layers. The possibility of calculating mother-to-calf nitrogen isotope fractionation, and the regularity of changes between calf layer δ15N values, allowed results of an isotopic clock model to predict the age of each calf when sampled with its mother. This model has the potential to be a straightforward method to estimate the beginning of lactation, therefore calf birth date when direct observations are not feasible. The non-lethal remote collection of epidermis appears to be an effective tool for the study of the physiology of reproduction of baleen whales. The parallel study of the three epidermal structural layers highlighted the importance of considering the unique mother-calf pair physiological status at the time of sampling time when stable isotope results are interpreted.
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Lactancia/fisiología , Embarazo/fisiología , Ballenas/fisiología , Animales , Isótopos de Carbono/farmacocinética , Epidermis/metabolismo , Femenino , Masculino , Isótopos de Nitrógeno/farmacocinéticaRESUMEN
BACKGROUND: The Logan graphical analysis (LGA) algorithm is widely used to quantify receptor density for parametric imaging in positron emission tomography (PET). Estimating receptor density, in terms of the non-displaceable binding potential (BPND), from the LGA using the ordinary least-squares (OLS) method has been found to be negatively biased owing to noise in PET data. This is because OLS does not consider errors in the X-variable (predictor variable). Existing bias reduction methods can either only reduce the bias slightly or reduce the bias accompanied by increased variation in the estimates. In this study, we addressed the bias reduction problem by applying a different regression method. METHODS: We employed least-squares cubic (LSC) linear regression, which accounts for errors in both variables as well as the correlation of these errors. Noise-free PET data were simulated, for 11C-carfentanil kinetics, with known BPND values. Statistical noise was added to these data and the BPNDs were re-estimated from the noisy data by three methods, conventional LGA, multilinear reference tissue model 2 (MRTM2), and LSC-based LGA; the results were compared. The three methods were also compared in terms of beta amyloid (A ß) quantification of 11C-Pittsburgh compound B brain PET data for two patients with Alzheimer's disease and differing A ß depositions. RESULTS: Amongst the three methods, for both synthetic and actual data, LSC was the least biased, followed by MRTM2, and then the conventional LGA, which was the most biased. Variations in the LSC estimates were smaller than those in the MRTM2 estimates. LSC also required a shorter computational time than MRTM2. CONCLUSIONS: The results suggest that LSC provides a better trade-off between the bias and variability than the other two methods. In particular, LSC performed better than MRTM2 in all aspects; bias, variability, and computational time. This makes LSC a promising method for BPND parametric imaging in PET studies.
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Enfermedad de Alzheimer/diagnóstico por imagen , Péptidos beta-Amiloides/química , Isótopos de Carbono/farmacocinética , Interpretación de Imagen Radiográfica Asistida por Computador/métodos , Algoritmos , Enfermedad de Alzheimer/metabolismo , Péptidos beta-Amiloides/metabolismo , Sesgo , Encéfalo/diagnóstico por imagen , Encéfalo/metabolismo , Isótopos de Carbono/química , Fentanilo/análogos & derivados , Fentanilo/química , Humanos , Análisis de los Mínimos Cuadrados , Tomografía de Emisión de Positrones , Relación Señal-RuidoRESUMEN
Kinetic modeling of the in vivo pyruvate-to-lactate conversion is crucial to investigating aberrant cancer metabolism that demonstrates Warburg effect modifications. Non-invasive detection of alterations to metabolic flux might offer prognostic value and improve the monitoring of response to treatment. In this clinical research project, hyperpolarized [1-13C] pyruvate was intravenously injected in a total of 10 brain tumor patients to measure its rate of conversion to lactate ( kPL ) and bicarbonate ( kPB ) via echo-planar imaging. Our aim was to investigate new methods to provide kPL and kPB maps with whole-brain coverage. The approach was data-driven and addressed two main issues: selecting the optimal model for fitting our data and determining an appropriate goodness-of-fit metric. The statistical analysis suggested that an input-less model had the best agreement with the data. It was also found that selecting voxels based on post-fitting error criteria provided improved precision and wider spatial coverage compared to using signal-to-noise cutoffs alone.
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Neoplasias Encefálicas , Encéfalo , Imagen Eco-Planar/métodos , Ácido Pirúvico , Encéfalo/diagnóstico por imagen , Encéfalo/metabolismo , Neoplasias Encefálicas/diagnóstico por imagen , Neoplasias Encefálicas/metabolismo , Isótopos de Carbono/análisis , Isótopos de Carbono/farmacocinética , Humanos , Interpretación de Imagen Asistida por Computador , Cinética , Ácido Láctico/análisis , Ácido Láctico/metabolismo , Ácido Pirúvico/análisis , Ácido Pirúvico/farmacocinéticaRESUMEN
In cancers, tumor cells are exposed to fluctuating nutrient microenvironments with limiting supplies of glucose and glutamine. While the metabolic program has been related to the expression of oncogenes, only fractional information is available on how variable precarious nutrient concentrations modulate the cellular levels of metabolites and their metabolic pathways. We thus sought to obtain an overview of the metabolic routes taken by 13C-glucose-derived metabolites in breast cancer MCF-7 cells growing in combinations of limiting glucose and glutamine concentrations. Isotopologue profiles of key metabolites were obtained by gas chromatography/mass spectrometry (GC/MS). They revealed that in limiting and standard saturating medium conditions, the same metabolic routes were engaged, including glycolysis, gluconeogenesis, as well as the TCA cycle with glutamine and pyruvate anaplerosis. However, the cellular levels of 13C-metabolites, for example, serine, alanine, glutamate, malate, and aspartate, were highly sensitive to the available concentrations and the ratios of glucose and glutamine. Notably, intracellular lactate concentrations did not reflect the Warburg effect. Also, isotopologue profiles of 13C-serine as well as 13C-alanine show that the same glucose-derived metabolites are involved in gluconeogenesis and pyruvate replenishment. Thus, anaplerosis and the bidirectional flow of central metabolic pathways ensure metabolic plasticity for adjusting to precarious nutrient conditions.
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Neoplasias de la Mama/metabolismo , Neoplasias de la Mama/patología , Isótopos de Carbono/farmacocinética , Glucosa/farmacocinética , Glutamina/farmacología , Ciclo del Ácido Cítrico/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Femenino , Glucosa/farmacología , Ácido Glutámico/metabolismo , Glucólisis/efectos de los fármacos , Humanos , Ácido Láctico/metabolismo , Células MCF-7 , Malatos/metabolismo , Redes y Vías Metabólicas/efectos de los fármacos , Redes y Vías Metabólicas/fisiología , Ácido Pirúvico/metabolismo , Microambiente Tumoral/efectos de los fármacosRESUMEN
To tackle urban water issues, the Chinese government has promoted the construction of sponge cities in recent years. Thirty cities have been designated as experimental sites to serve as models for future sponge city construction, as more than 80% of the built-up urban areas in China must reach the standards of sponge cities by 2030. Greening plants play an important role in sponge cities, and water-use efficiency (WUE) is a vital index to determine whether plants could adapt to and grow healthily in environments with water deficits. In this study, WUE of greening plants was quantified by measuring the stable carbon isotope fractionation. Suitable plants for the green spaces in Guyuan sponge city, in northern China, were selected based on their WUE, and the main factors affecting WUE were studied in four habitats within the city. Plant species identity had the greatest effect on WUE, while habitat and plant life form had lower effect, illustrating that WUE is a relatively stable and reliable index for the classification of plant species. We can improve the WUE and ecological function of green spaces in sponge cities by using isotope technology to select suitable plant species with high WUE. To our knowledge, this study is the first to select plant species for sponge city by using this method, providing a quick and scientific method for the selection of greening plants for future sponge cities.
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Isótopos de Carbono/farmacocinética , Viridiplantae/metabolismo , Agua/metabolismo , Isótopos de Carbono/química , China , Ciudades , Ecosistema , Agua Subterránea/análisis , Agua Subterránea/química , Plantas/química , Plantas/metabolismo , Viridiplantae/química , Agua/análisis , Recursos Hídricos/provisión & distribución , Abastecimiento de Agua/normasRESUMEN
During carcass decomposition, tissues undergo biochemical changes: Cells autolyze, enteric microbes ferment cellular products, and tissues degrade. Ultimately, decomposition fluids are released as an ephemeral nitrogen (N) and carbon source to the surrounding environment. However, decomposition fluids are δ15N-enriched relative to body tissues, leading to a disconnect between starting tissue composition and ending fluid composition. It remains largely unknown when or if tissues exhibit δ15N enrichment postmortem despite the importance of tissue stable isotopes to ecologists. To test our hypothesis that tissues would become progressively δ15N-enriched during decay, soft tissues and bone were collected from beaver carcasses at five time points. All soft tissues, including muscle, were significantly δ15N-enriched compared to fresh tissues, but were not as enriched as decomposition fluids. Tissue breakdown is initially dominated by anaerobic autolysis and later by microbe and insect infiltration, and partly explains decay fluid isotopic enrichment. We speculate that after rupture, preferential volatilization of δ15N-depleted compounds (especially ammonia) contributes to further enrichment. These results constrain the timing, rate, and potential mechanisms driving carcass isotopic enrichment during decay, and suggest that found carcasses (e.g., road kill) should be used with caution for inferring trophic ecology as decay can result in significant postmortem δ15N enrichment.
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Isótopos de Carbono/análisis , Isótopos de Carbono/farmacocinética , Isótopos de Nitrógeno/análisis , Isótopos de Nitrógeno/farmacocinética , Cambios Post Mortem , Animales , Roedores , Distribución TisularRESUMEN
Positron emission tomography (PET) radiopharmaceuticals can noninvasively measure free fatty acid (FFA) uptake into adipose tissue. We studied 29 volunteers to test whether abdominal and femoral subcutaneous adipose tissue FFA uptake measured using [1-11C]palmitate PET agrees with FFA storage rates measured using an intravenous bolus of [1-14C]palmitate and adipose biopsies. The dynamic left ventricular cavity PET images combined with blood sample radioactivity corrected for the 11CO2 content were used to create the blood time activity curve (TAC), and the constant (Ki) was determined using Patlak analysis of the TACs generated for regions of interest in abdominal subcutaneous fat. These data were used to calculate palmitate uptake rates in abdominal subcutaneous adipose tissue (µmol·kg-1·min-1). Immediately after the dynamic imaging, a static image of the thigh was taken to measure the standardized uptake value (SUV) in thigh adipose tissue, which was scaled to each participant's abdominal adipose tissue SUV to calculate thigh adipose palmitate uptake rates. Abdominal adipose palmitate uptake using PET [1-11C]palmitate was correlated with, but significantly (P < 0.001) greater than, FFA storage measured using [1-14C]palmitate and adipose biopsy. Thigh adipose palmitate measured using PET calculation was positively correlated (R2 = 0.44, P < 0.0001) with and not different from the biopsy approach. The relative differences between PET measured abdominal subcutaneous adipose tissue palmitate uptake and biopsy-measured palmitate storage were positively correlated (P = 0.03) with abdominal subcutaneous fat. We conclude that abdominal adipose tissue FFA uptake measured using PET does not equate to adipose FFA storage measured using biopsy techniques.
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Tejido Adiposo/patología , Ácidos Grasos no Esterificados/farmacocinética , Tomografía de Emisión de Positrones , Grasa Subcutánea/diagnóstico por imagen , Grasa Subcutánea/metabolismo , Tejido Adiposo/diagnóstico por imagen , Adiposidad/fisiología , Adulto , Biopsia , Distribución de la Grasa Corporal/métodos , Índice de Masa Corporal , Isótopos de Carbono/análisis , Isótopos de Carbono/farmacocinética , Radioisótopos de Carbono/análisis , Radioisótopos de Carbono/farmacocinética , Femenino , Humanos , Peso Corporal Ideal/fisiología , Lipólisis/fisiología , Masculino , Obesidad/metabolismo , Obesidad/patología , Sobrepeso/metabolismo , Sobrepeso/patología , Ácido Palmítico/química , Ácido Palmítico/farmacocinética , Tomografía de Emisión de Positrones/métodosRESUMEN
Indirect calorimetry is a tool used routinely by sport/exercise physiologist to assess the metabolic response to training and to nutritional interventions. There are different stoichiometric equations to estimate fat (FatOxR ) and carbohydrates (CHOOxR ) oxidation rates, however there are not enough information in literature about what are the most accurate equations. The purpose of this study was to determine the concordance between indirect calorimetry and a method of reference for stoichiometric equations used to estimate FatOxR and CHOOxR . Concordance between indirect calorimetry and the method of reference (13 C to 12 C ratio (13 C:12 C ratio) technique) for key stoichiometric equations was assessed in well-trained triathletes. Subjects carried out a carbohydrate depletion-repletion protocol, labeling the glycogen stores with 13 C, and a laboratory test to assess the 13 C metabolic response during a wide range of aerobic intensities during exercise. All the equations showed a narrow agreement interval (Δ) (CHOOxR nPC (protein component negligible): -0.308, 0.308, CHOOxR PC (protein component): -0.268, 0.268, FatOxR nPC and PC: -0.032, 0.032 (g·min-1 )). FatOxR showed a similar concordance (28-32%) with CHOOxR nPC ranging from 55% to 75%, and for CHOOxR PC between 51% to 71%. None of the stoichiometric equations met a perfect agreement with the method of reference. The Jeukendrup and Wallis equation showed the best concordance for CHOOxR nPC whilst the Frayn and Ferrannini (Glu) equations had the best agreement for CHOOxR PC. All FatOxR equations showed similar concordances and they are able to be used indistinctly.
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Algoritmos , Metabolismo de los Hidratos de Carbono , Metabolismo de los Lípidos , Acondicionamiento Físico Humano/fisiología , Adulto , Calorimetría Indirecta/métodos , Calorimetría Indirecta/normas , Dióxido de Carbono/metabolismo , Isótopos de Carbono/farmacocinética , Dieta , Glucógeno/metabolismo , Humanos , Masculino , Oxidación-Reducción , Acondicionamiento Físico Humano/métodos , Valores de Referencia , Reproducibilidad de los ResultadosRESUMEN
Cancers in the bone produce a number of severe symptoms including pain that compromises patient functional status, quality of life, and survival. The source of this pain is multifaceted and includes factors secreted from tumor cells. Malignant cells release the neurotransmitter and cell-signaling molecule glutamate via the oxidative stress-related cystine/glutamate antiporter, system xC-, which reciprocally imports cystine for synthesis of glutathione and the cystine/cysteine redox cycle. Pharmacological inhibition of system xC- has shown success in reducing and delaying the onset of cancer pain-related behavior in mouse models. This investigation describes the development of a stable siRNA-induced knockdown of the functional trans-membrane system xC- subunit xCT ( SLC7A11) in the human breast cancer cell line MDA-MB-231. Clones were verified for xCT knockdown at the transcript, protein, and functional levels. RNAseq was performed on a representative clone to comprehensively examine the transcriptional cellular signature in response to xCT knockdown, identifying multiple differentially regulated factors relevant to cancer pain including nerve growth factor, interleukin-1, and colony-stimulating factor-1. Mice were inoculated intrafemorally and recordings of pain-related behaviors including weight bearing, mechanical withdrawal, and limb use were performed. Animals implanted with xCT knockdown cancer cells displayed a delay until the onset of nociceptive behaviors relative to control cells. These results add to the body of evidence suggesting that a reduction in glutamate release from cancers in bone by inhibition of the system xC- transporter may decrease the severe and intractable pain associated with bone metastases.
Asunto(s)
Sistema de Transporte de Aminoácidos y+/metabolismo , Neoplasias de la Mama/complicaciones , Dolor en Cáncer/etiología , Dolor en Cáncer/genética , Regulación Neoplásica de la Expresión Génica/fisiología , Sistema de Transporte de Aminoácidos y+/genética , Animales , Neoplasias de la Mama/genética , Neoplasias de la Mama/metabolismo , Proteínas de Unión al Calcio/genética , Proteínas de Unión al Calcio/metabolismo , Isótopos de Carbono/farmacocinética , Carnitina O-Palmitoiltransferasa/genética , Carnitina O-Palmitoiltransferasa/metabolismo , Línea Celular Tumoral , Cistina/farmacocinética , Modelos Animales de Enfermedad , Femenino , Ácido Glutámico/metabolismo , Glutatión/metabolismo , Humanos , Interleucina-1/metabolismo , Factor Estimulante de Colonias de Macrófagos/genética , Factor Estimulante de Colonias de Macrófagos/metabolismo , Ratones Endogámicos BALB C , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/metabolismo , Factor de Crecimiento Nervioso/genética , Factor de Crecimiento Nervioso/metabolismo , ARN Interferente Pequeño/genética , ARN Interferente Pequeño/metabolismoRESUMEN
Spermatozoa from three feline species-the domestic cat (Felis catus), the cheetah (Acinonyx jubatus), and the clouded leopard (Neofelis nebulosa)-were analyzed using metabolomic profiling and 13C-based fluxomics to address questions raised regarding their energy metabolism. Metabolic profiles and utilization of 13C-labeled energy substrates were detected and quantified using gas chromatography-mass spectrometry (GC-MS). Spermatozoa were collected by electroejaculation and incubated in media supplemented with 1.0 mM [U13C]-glucose, [U13C]-fructose, or [U13C]-pyruvate. Evaluation of intracellular metabolites following GC-MS analysis revealed the uptake and utilization of labeled glucose and fructose in sperm, as indicated by the presence of heavy ions in glycolytic products lactate and pyruvate. Despite evidence of substrate utilization, neither glucose nor fructose had an effect on the sperm motility index of ejaculated spermatozoa from any of the three felid species, and limited entry of pyruvate derived from these hexose substrates into mitochondria and the tricarboxylic acid cycle was detected. However, pathway utilization was species-specific for the limited number of individuals (four to seven males per species) assessed in these studies. An inhibitor of fatty acid beta-oxidation (FAO), etomoxir, altered metabolic profiles of all three felid species but decreased motility only in the cheetah. While fluxomic analysis provided direct evidence that glucose and fructose undergo catabolic metabolism, other endogenous substrates such as endogenous lipids may provide energy to fuel motility.
Asunto(s)
Isótopos de Carbono/farmacocinética , Metabolismo Energético , Felidae/metabolismo , Metabolómica/métodos , Espermatozoides/metabolismo , Acinonyx/metabolismo , Animales , Animales Domésticos , Isótopos de Carbono/análisis , Gatos/metabolismo , Ciclo del Ácido Cítrico/fisiología , Felidae/clasificación , Glucólisis/fisiología , Ácido Láctico/metabolismo , Masculino , Ácido Pirúvico/metabolismo , Análisis de Semen/métodos , Análisis de Semen/veterinariaRESUMEN
AIMS: 13C-glucose breath tests are reported as an alternative non-invasive method to evaluate glucose metabolism. However, the metabolic results differ based on the site of the carbon atom in the glucose. The aim of this study was to evaluate changes in the metabolism of carbon atoms contained in glucose in patients with diabetes using [1, 2, 3-13C]glucose breath tests. METHODS: Sixteen healthy participants and 20 diabetic patients were enrolled in the study. Three types of breath tests, [1-13C], [2-13C], and [3-13C]glucose breath tests, were performed after an overnight fast. Breath samples were taken at baseline and at 10-min intervals over 150 min, and 13CO2 excretion curves were expressed using non-dispersive infrared isotope spectrometry. RESULTS: 13CO2 levels increased more rapidly, and the peak value of 13CO2 (Cmax) was highest after the administration of [3-13C]glucose followed by [2-13C] and [1-13C]glucose in controls. Delayed 13CO2 excretion and a low area under the curve through 150 min (AUC150) were obtained in diabetic patients. The group with severe diabetes had a significantly lower Cmax and AUC150 in the [1-13C]glucose breath test. CONCLUSIONS: The [1-13C]glucose breath test, which has been used to evaluate glucose metabolism, is suitable for patients with late-stage diabetes, whereas the [2-13C]glucose breath test is ideal in the early stages. Although the [3-13C]glucose breath test is theoretically useful for evaluating the uptake of glucose and the anaerobic glycolysis system, it can be used in practice to distinguish reduced uptake from impaired oxidation of glucose in combination with the other two tests.
Asunto(s)
Isótopos de Carbono/análisis , Isótopos de Carbono/farmacocinética , Diabetes Mellitus Tipo 2/metabolismo , Glucosa/análisis , Glucosa/farmacocinética , Adulto , Anciano , Anciano de 80 o más Años , Pruebas Respiratorias/métodos , Estudios de Casos y Controles , Diagnóstico Precoz , Femenino , Gluconeogénesis/efectos de los fármacos , Humanos , Masculino , Persona de Mediana Edad , Oxidación-Reducción , Adulto JovenRESUMEN
When activated, microglial cells have the potential not only to secrete typical proinflammatory mediators but also to release the neurotransmitter glutamate in amounts that may promote excitotoxicity. Here, we wished to determine the potential of the Parkinson's disease (PD) protein α-Synuclein (αS) to stimulate glutamate release using cultures of purified microglial cells. We established that glutamate release was robustly increased when microglial cultures were treated with fibrillary aggregates of αS but not with the native monomeric protein. Promotion of microglial glutamate release by αS aggregates (αSa) required concomitant engagement of TLR2 and P2X7 receptors. Downstream to cell surface receptors, the release process was mediated by activation of a signaling cascade sequentially involving phosphoinositide 3-kinase (PI3K) and NADPH oxidase, a superoxide-producing enzyme. Inhibition of the Xc- antiporter, a plasma membrane exchange system that imports extracellular l-cystine and exports intracellular glutamate, prevented the release of glutamate induced by αSa, indicating that system Xc- was the final effector element in the release process downstream to NADPH oxidase activation. Of interest, the stimulation of glutamate release by αSa was abrogated by dopamine through an antioxidant effect requiring D1 dopamine receptor activation and PI3K inhibition. Altogether, present data suggest that the activation of microglial cells by αSa may possibly result in a toxic build-up of extracellular glutamate contributing to excitotoxic stress in PD. The deficit in dopamine that characterizes this disorder may further aggravate this process in a vicious circle mechanism.
Asunto(s)
Dopamina/farmacología , Ácido Glutámico/metabolismo , Microglía/efectos de los fármacos , Agregado de Proteínas/efectos de los fármacos , alfa-Sinucleína/metabolismo , Adenosina Trifosfato/análogos & derivados , Adenosina Trifosfato/farmacología , Animales , Animales Recién Nacidos , Anticuerpos Monoclonales/farmacología , Isótopos de Carbono/farmacocinética , Células Cultivadas , Cistina/farmacocinética , Humanos , Lipopéptidos/farmacología , Lipopolisacáridos/farmacología , Ratones , Microglía/ultraestructura , Inhibidores de Agregación Plaquetaria/farmacología , Especies Reactivas de Oxígeno/metabolismo , Transducción de Señal/efectos de los fármacos , Receptor Toll-Like 2/inmunología , alfa-Sinucleína/farmacologíaRESUMEN
SCOPE: Quinic acid in its free form is broadly abundant in plants, and can accumulate in copious amounts in coffee, tea, and certain fruits. However, it has been mostly studied as chlorogenic acid, an ester of caffeic and quinic acids. When chlorogenic acid reaches the colon, it is hydrolyzed by microbial esterases releasing caffeic and quinic acids. While biotransformation of chlorogenic and caffeic acids have been elucidated by in vitro and in vivo studies, the gut metabolism of quinic acid has been so far overlooked. METHODS AND RESULTS: [U-13 C]-Quinic acid is submitted to a colonic model using human fecal microbiota for assessing its metabolic fate. The metabolite profiles formed along microbial biotransformation are monitored by a combined metabolomics approach, using both 2D GC- and ultra-HPLC-MS. Six metabolic intermediates are identified by incorporation of isotopic label. CONCLUSION: Two parallel degradation pathways could be proposed: (1) an oxidative route, leading to aromatization and accumulation of protocatechuic acid, and a (2) reductive route, including dehydroxylation to cyclohexane carboxylic acid. Elucidating the biotransformation of food bioactives by the gut microbiota is of relevance for understanding nutrition, interindividual variability and potential effects on human metabolism.