Deuterated water imaging of the rat brain following metabolism of [2 H7 ]glucose.

PURPOSE: To determine whether deuterated water (HDO) generated from the metabolism of [2 H7 ]glucose is a sensitive biomarker of cerebral glycolysis and oxidative flux. METHODS: A bolus of [2 H7 ]glucose was injected through the tail vein at 1.95 g/kg into Sprague-Dawley rats. A 2 H surface coil was placed on top of the head to record 2 H spectra of the brain every 1.3 minutes to measure glucose uptake and metabolism to HDO, lactate, and glutamate/glutamine. A two-point Dixon method based on a gradient-echo sequence was used to reconstruct deuterated glucose and water (HDO) images selectively. RESULTS: The background HDO signal could be detected and imaged before glucose injection. The 2 H NMR spectra showed arrival of [2 H7 ]glucose and its metabolism in a time-dependent manner. A ratio of the HDO to glutamate/glutamine resonances demonstrates a pseudo-steady state following injection, in which cerebral metabolism dominates wash-in of HDO generated by peripheral metabolism. Brain spectroscopy reveals that HDO generation is linear with lactate and glutamate/glutamine appearance in the appropriate pseudo-steady state window. Selective imaging of HDO and glucose is easily accomplished using a gradient-echo method. CONCLUSION: Metabolic imaging of HDO, as a marker of glucose, lactate, and glutamate/glutamine metabolism, has been shown here for the first time. Cerebral glucose metabolism can be assessed efficiently using a standard gradient-echo sequence that provides superior in-plane resolution compared with CSI-based techniques.

Detecting altered hepatic lipid oxidation by MRI in an animal model of MASLD.

Metabolic dysfunction-associated steatotic liver disease (MASLD) prevalence is increasing annually and affects over a third of US adults. MASLD can progress to metabolic dysfunction-associated steatohepatitis (MASH), characterized by severe hepatocyte injury, inflammation, and eventual advanced fibrosis or cirrhosis. MASH is predicted to become the primary cause of liver transplant by 2030. Although the etiology of MASLD/MASH is incompletely understood, dysregulated fatty acid oxidation is implicated in disease pathogenesis. Here, we develop a method for estimating hepatic ß-oxidation from the metabolism of [D15]octanoate to deuterated water and detection with deuterium magnetic resonance methods. Perfused livers from a mouse model of MASLD reveal dysregulated hepatic ß-oxidation, findings that corroborate in vivo imaging. The high-fat-diet-induced MASLD mouse studies indicate that decreased ß-oxidative efficiency in the fatty liver could serve as an indicator of MASLD progression. Furthermore, our method provides a clinically translatable imaging approach for determining hepatic ß-oxidation efficiency.

Nanomaterials for Molecular Detection and Analysis of Extracellular Vesicles.

Extracellular vesicles (EVs) have emerged as a novel resource of biomarkers for cancer and certain other diseases. Probing EVs in body fluids has become of major interest in the past decade in the development of a new-generation liquid biopsy for cancer diagnosis and monitoring. However, sensitive and specific molecular detection and analysis are challenging, due to the small size of EVs, low amount of antigens on individual EVs, and the complex biofluid matrix. Nanomaterials have been widely used in the technological development of protein and nucleic acid-based EV detection and analysis, owing to the unique structure and functional properties of materials at the nanometer scale. In this review, we summarize various nanomaterial-based analytical technologies for molecular EV detection and analysis. We discuss these technologies based on the major types of nanomaterials, including plasmonic, fluorescent, magnetic, organic, carbon-based, and certain other nanostructures. For each type of nanomaterial, functional properties are briefly described, followed by the applications of the nanomaterials for EV biomarker detection, profiling, and analysis in terms of detection mechanisms.

Ex Vivo Hepatic Perfusion Through the Portal Vein in Mouse.

Metabolic diseases such as diabetes, pre-diabetes, non-alcoholic fatty liver disease (NAFLD), and nonalcoholic steatohepatitis (NASH) are becoming increasingly common. Ex vivo liver perfusions allow for a comprehensive analysis of liver metabolism using nuclear magnetic resonance (NMR), in nutritional conditions that can be rigorously controlled. As in silico simulations remain a primarily theoretical means of assessing hormone actions and the effects of pharmaceutical intervention, the perfused liver remains one of the most valuable test beds for understanding hepatic metabolism. As these studies guide basic insights into hepatic physiology, results must be accurate and reproducible. The greatest factor in the reproducibility of ex vivo hepatic perfusion is the quality of surgery. Therefore, we have introduced an organized and streamlined method to perform ex vivo mouse liver perfusions in the context of in situ NMR experiments. We also describe a unique application and discuss common issues encountered in these studies. The overall purpose is to provide an uncomplicated guide to a technique we have refined over several years that we deem the golden standard for obtaining reproducible results in hepatic resections and perfusions in the context of in situ NMR experiments. The distance to the center of the field for the magnet as well as the inaccessibility of the tissue to intervention during the NMR experiment makes our methods novel.

Synergistic Effect of ß-Lapachone and Aminooxyacetic Acid on Central Metabolism in Breast Cancer.

The compound ß-lapachone, a naturally derived naphthoquinone, has been utilized as a potent medicinal nutrient to improve health. Over the last twelve years, numerous reports have demonstrated distinct associations of ß-lapachone and NAD(P)H: quinone oxidoreductase 1 (NQO1) protein in the amelioration of various diseases. Comprehensive research of NQO1 bioactivity has clearly confirmed the tumoricidal effects of ß-lapachone action through NAD+-keresis, in which severe DNA damage from reactive oxygen species (ROS) production triggers a poly-ADP-ribose polymerase-I (PARP1) hyperactivation cascade, culminating in NAD+/ATP depletion. Here, we report a novel combination strategy with aminooxyacetic acid (AOA), an aspartate aminotransferase inhibitor that blocks the malate-aspartate shuttle (MAS) and synergistically enhances the efficacy of ß-lapachone metabolic perturbation in NQO1+ breast cancer. We evaluated metabolic turnover in MDA-MB-231 NQO1+, MDA-MB-231 NQO1-, MDA-MB-468, and T47D cancer cells by measuring the isotopic labeling of metabolites from a [U-13C]glucose tracer. We show that ß-lapachone treatment significantly hampers lactate secretion by ~85% in NQO1+ cells. Our data demonstrate that combinatorial treatment decreases citrate, glutamate, and succinate enrichment by ~14%, ~50%, and ~65%, respectively. Differences in citrate, glutamate, and succinate fractional enrichments indicate synergistic effects on central metabolism based on the coefficient of drug interaction. Metabolic modeling suggests that increased glutamine anaplerosis is protective in the case of MAS inhibition.

Hyperpolarized Dihydroxyacetone Is a Sensitive Probe of Hepatic Gluconeogenic State.

Type II diabetes and pre-diabetes are widely prevalent among adults. Elevated serum glucose levels are commonly treated by targeting hepatic gluconeogenesis for downregulation. However, direct measurement of hepatic gluconeogenic capacity is accomplished only via tracer metabolism approaches that rely on multiple assumptions, and are clinically intractable due to expense and time needed for the studies. We previously introduced hyperpolarized (HP) [2-13C]dihydroxyacetone (DHA) as a sensitive detector of gluconeogenic potential, and showed that feeding and fasting produced robust changes in the ratio of detected hexoses (6C) to trioses (3C) in the perfused liver. To confirm that this ratio is robust in the setting of treatment and hormonal control, we used ex vivo perfused mouse livers from BLKS mice (glucagon treated and metformin treated), and db/db mice. We confirm that the ratio of signal intensities of 6C to 3C in 13C nuclear magnetic resonance spectra post HP DHA administration is sensitive to hepatic gluconeogenic state. This method is directly applicable in vivo and can be implemented with existing technologies without the need for substantial modifications.

Application of Carbon-13 Isotopomer Analysis to Assess Perinatal Myocardial Glucose Metabolism in Sheep.

Ovine models of pregnancy have been used extensively to study maternal-fetal interactions and have provided considerable insight into nutrient transfer to the fetus. Ovine models have also been utilized to study congenital heart diseases. In this work, we demonstrate a comprehensive assessment of heart function and metabolism using a perinatal model of heart function with the addition of a [U-13C]glucose as tracer to study central energy metabolism. Using nuclear magnetic resonance spectroscopy, and metabolic modelling, we estimate myocardial citric acid cycle turnover (normalized for oxygen consumption), substrate selection, and anaplerotic fluxes. This methodology can be applied to studying acute and chronic effects of hormonal signaling in future studies.