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In an acute contact test with bees the compound of interest is dissolved in a carrier solvent (frequently acetone) and then a droplet of the solution is placed on the dorsal thorax of the bee. The volume of the droplet is standardised to 1 µL for honeybees and to 2 µL for bumblebees. In practice the same droplet volume is used for bees with very different sizes. In this research the effect of the droplet volume was evaluated with acute contact tests with dimethoate for the alfalfa leafcutter bee, the red mason bee, the honeybee and the bumblebee. The results were analysed with a ToxicoKinetic ToxicoDynamic (TKTD) model to separate kinetic from dynamic effects. This allows to compare the sensitivity of the bee based on the effect threshold and not on the time, species and test dependent LD50s. The analysis of the test results indicates that the magnitude of the response of the bees increased with increasing droplet size. The results also showed that the manifestation of effects over time is slower for the red mason bee and the bumblebee compared to the honeybee and the alfalfa leafcutter bee. This implies that the result of a 2 day test with a fixed dosing volume results in different response for a bumblebee compared to the alfalfa leafcutter bee, not because of different sensitivities of the bees involved but due to the difference of relative dosed surface ratio. So comparing the sensitivity of bee species, based on standardised tests is biased and amplifies the sensitivity for the smaller bee species.

Abstract Background The Asian tiger mosquito Aedes albopictus is an extremely invasive, globally distributed and medically important vector of various human and veterinary pathogens. In Germany, where this species was recently introduced, its establishment may become modulated by interspecific competition from autochthonous mosquito species, especially Culex pipiens (s.l.). While competitive superiority of Ae. albopictus to Cx. pipiens (s.l.) has been described elsewhere, it has not been assessed in the epidemiological conditions of Germany. The present study aimed to determine if such superiority exists under the physicochemical and microclimatic conditions typical for container habitats in Germany. Methods In a replacement series experiment, the larval and pupal responses of Ae. albopictus and Cx. pipiens (s.l.) (mortality, development time, growth) to interspecific interaction (five larval ratios) at (sub-)optimal temperatures (15, 20 and 25 °C) and differing food supply (3 and 6 mg animal-based food larva-1) were investigated using a randomized split-plot design. In addition to physicochemical measurements of the test media, natural physicochemical conditions were determined for comparative analyses in mosquito breeding sites across the Rhine-Main metropolitan region of Germany. Results Under the physicochemical and microclimatic conditions similar to the breeding sites of the Rhine-Main region, competitive superiority of Cx. pipiens (s.l.) to Ae. albopictus in terms of larval survival was more frequently observed than balanced coexistence. Food regime and multifactorial interactions, but not temperature alone, were controlling factors for interspecific competition. Larval food regime and the larval ratio of Ae. albopictus influenced the physicochemistry and algal growth at 15 °C, with increased Ae. albopictus mortality linked to a decreasing number of Scenedesmus, Oocystis and Anabaena algae. Conclusions Under the present environmental conditions, the spread of Ae. albopictus from isolated foci in Germany may generally be slowed by biotic interactions with the ubiquitous Cx. pipiens (s.l.) (and potentially other container-breeding mosquito species) and by limnic microalgae in microhabitats with high resource levels. Detailed knowledge of the context dependency in temperate mosquito ecology, and interrelations of physicochemistry and phycology may help to achieve a better understanding of the upcoming Ae. albopictus colonization processes in central and northern Europe.

Abstract Melamine is a key compound used as a clarifying agent for waste lubricating oil primarily due to its excellent adsorbent properties. Moreover, considerable interest exists for the further modification of melamine in order to provide a remediation agent with improved clarification capacity. In this study, hexamethylolmelamine was prepared using a solution of formaldehyde, which provided an agent capable of incorporation into a silicate polymer framework. Subsequently, the resultant monomer was added to a solution of silicate to produce the melamine-silica polymer. The melamine and melamine-silica polymer were characterized using the techniques of XRD, FTIR, SSA and thermal analysis to confirm structural and morphological characteristics. These characterizations indicated that the increase in the surface area of the 0.315 m2/g to melamine to 26.71 m2/g of melamine-silica suggests the effective introduction of silanols groups to hexamethylolmelamine and, therefore, corresponds to thehigh performance in relation to melamine as clarifying of waste lubricant oil.

Identifying initial triggering events in neurodegenerative disorders is critical to developing preventive therapies. In Huntington's disease (HD), hyperdopaminergia-probably triggered by the dysfunction of the most affected neurons, indirect pathway spiny projection neurons (iSPNs)-is believed to induce hyperkinesia, an early stage HD symptom. However, how this change arises and contributes to HD pathogenesis is unclear. Here, we demonstrate that genetic disruption of iSPNs function by Ntrk2/Trkb deletion in mice results in increased striatal dopamine and midbrain dopaminergic neurons, preceding hyperkinetic dysfunction. Transcriptomic analysis of iSPNs at the pre-symptomatic stage showed de-regulation of metabolic pathways, including upregulation of Gsto2, encoding glutathione S-transferase omega-2 (GSTO2). Selectively reducing Gsto2 in iSPNs in vivo effectively prevented dopaminergic dysfunction and halted the onset and progression of hyperkinetic symptoms. This study uncovers a functional link between altered iSPN BDNF-TrkB signalling, glutathione-ascorbate metabolism and hyperdopaminergic state, underscoring the vital role of GSTO2 in maintaining dopamine balance., Competing Interests: Competing interests: The authors declare no competing interests., (© 2024. The Author(s).)

Doxorubicin (DOX) is a widely used chemotherapeutic agent that can cause serious cardiotoxic side effects, leading to heart failure (HF). Impaired mitochondrial function is thought to be key factor driving progression into HF. We have previously shown in a rat model of DOX-HF that heart failure with reduced ejection fraction correlates with mitochondrial loss and dysfunction. Adenosine monophosphate-dependent kinase (AMPK) is a cellular energy sensor, regulating mitochondrial biogenesis and energy metabolism, including fatty acid oxidation. We hypothesised that AMPK activation could restore mitochondrial function and therefore be a novel cardioprotective strategy for the prevention of DOX-HF. Consequently, we set out to assess whether 5-aminoimidazole-4-carboxamide 1-β-D-ribofuranoside (AICAR), an activator of AMPK, could prevent cardiac functional decline in this chronic intravenous rat model of DOX-HF. In line with our hypothesis, AICAR improved cardiac systolic function. AICAR furthermore improved cardiac mitochondrial fatty acid oxidation, independent of mitochondrial number, and in the absence of observable AMPK-activation. In addition, we found that AICAR prevented loss of myocardial mass. RNAseq analysis showed that this may be driven by normalisation of pathways associated with ribosome function and protein synthesis, which are impaired in DOX-treated rat hearts. AICAR furthermore prevented dyslipidemia and excessive body-weight loss in DOX-treated rats, which may contribute to preservation of myocardial mass. Though it is unclear whether AICAR exerted its cardioprotective effect through cardiac or extra-cardiac AMPK-activation or via an AMPK-independent effect, these results show promise for the use of AICAR as a cardioprotective agent in DOX-HF to both preserve cardiac function and mass., Competing Interests: Declaration of competing interest Conflict of Interest: none declared., (Copyright © 2024 The Authors. Published by Elsevier Ltd.. All rights reserved.)

The use of the adjuvant therapeutic antibody trastuzumab in breast cancer is associated with a range of cardiotoxic side effects despite successfully reducing the severity of outcomes cancer patients,. The most common cardiac effect, a reduction in left ventricular ejection fraction (LVEF), is a known precursor to heart failure and often requires interruption of chemotherapy to avoid endangering patients further. An understanding of trastuzumab's cardiac-specific interactions is therefore critical in devising new methods to not only avoid permanent cardiac damage, but also prolong treatment time, and therefore effectiveness, for breast cancer patients. Increasingly, the use of exercise as a treatment has been indicated across the field of cardio-oncology due to encouraging evidence that it can protect against LVEF reductions and heart failure. This review explores the mechanisms of trastuzumab-mediated cardiotoxicity, as well as the physiological effects of exercise on the heart, in order to assess the suitability of exercise intervention for breast cancer patients on trastuzumab antibody-therapy. We furthermore draw comparison to existing evidence for exercise intervention as a cardioprotective treatment in doxorubicin-induced cardiotoxicity. Although preclinical evidence seems to support exercise-based approaches also in trastuzumab-cardiotoxicity, current clinical evidence is too limited to confidently recommend it as a treatment, largely owing to issues of adherence. Future studies should therefore examine how the variety and duration of exercise can be adjusted to improve treatment effectiveness at a more personalised level., (© 2023. The Author(s).)

Doxorubicin (DOX) is a successful chemotherapeutic widely used for the treatment of a range of cancers. However, DOX can have serious side-effects, with cardiotoxicity and hepatotoxicity being the most common events. Oxidative stress and changes in metabolism and bioenergetics are thought to be at the core of these toxicities. We have previously shown in a clinically-relevant rat model that a low DOX dose of 2 mg kg -1 week -1 for 6 weeks does not lead to cardiac functional decline or changes in cardiac carbohydrate metabolism, assessed with hyperpolarized [1- 13 C]pyruvate magnetic resonance spectroscopy (MRS). We now set out to assess whether there are any signs of liver damage or altered liver metabolism using this subclinical model. We found no increase in plasma alanine aminotransferase (ALT) activity, a measure of liver damage, following DOX treatment in rats at any time point. We also saw no changes in liver carbohydrate metabolism, using hyperpolarized [1- 13 C]pyruvate MRS. However, using metabolomic analysis of liver metabolite extracts at the final time point, we found an increase in most acyl-carnitine species as well as increases in high energy phosphates, citrate and markers of oxidative stress. This may indicate early signs of steatohepatitis, with increased and decompensated fatty acid uptake and oxidation, leading to oxidative stress., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Timm, Ball, Miller, Savic, West, Griffin and Tyler.)

Long-term cardiovascular complications of cancer therapy are becoming ever more prevalent due to increased numbers of cancer survivors. Cancer therapy-induced cardiotoxicity (CTIC) is an incompletely understood consequence of various chemotherapies, targeted anti-cancer agents and radiation therapy. It is typically detected clinically by a reduction in cardiac left ventricular ejection fraction, assessed by echocardiography. However, once cardiac functional decline is apparent, this indicates irreversible cardiac damage, highlighting a need for the development of diagnostics which can detect CTIC prior to the onset of functional decline. There is increasing evidence to suggest that pathological alterations to cardiac metabolism play a crucial role in the development of CTIC. This review discusses the metabolic alterations and mechanisms which occur in the development of CTIC, with a focus on doxorubicin, trastuzumab, imatinib, ponatinib, sunitinib and radiotherapy. Potential methods to diagnose and predict CTIC prior to functional cardiac decline in the clinic are evaluated, with a view to both biomarker and imaging-based approaches. Finally, the therapeutic potential of therapies which manipulate cardiac metabolism in the context of adjuvant cardioprotection against CTIC is examined. Together, an integrated view of the role of metabolism in pathogenesis, diagnosis and treatment is presented.

The diabetic heart has a decreased ability to metabolize glucose. The anti-ischemic drug meldonium may provide a route to counteract this by reducing l-carnitine levels, resulting in improved cardiac glucose utilization. Therefore, the aim of this study was to use the novel technique of hyperpolarized magnetic resonance to investigate the in vivo effects of treatment with meldonium on cardiac metabolism and function in control and diabetic rats. Thirty-six male Wistar rats were injected either with vehicle, or with streptozotocin (55 mg/kg) to induce a model of type 1 diabetes. Daily treatment with either saline or meldonium (100 mg/kg/day) was undertaken for three weeks. in vivo cardiac function and metabolism were assessed with CINE MRI and hyperpolarized magnetic resonance respectively. Isolated perfused hearts were challenged with low-flow ischemia/reperfusion to assess the impact of meldonium on post-ischemic recovery. Meldonium had no significant effect on blood glucose concentrations or on baseline cardiac function. However, hyperpolarized magnetic resonance revealed that meldonium treatment elevated pyruvate dehydrogenase flux by 3.1-fold and 1.2-fold in diabetic and control animals, respectively, suggesting an increase in cardiac glucose oxidation. Hyperpolarized magnetic resonance further demonstrated that meldonium reduced the normalized acetylcarnitine signal by 2.1-fold in both diabetic and control animals. The increase in pyruvate dehydrogenase flux in vivo was accompanied by an improvement in post-ischemic function ex vivo, as meldonium elevated the rate pressure product by 1.3-fold and 1.5-fold in the control and diabetic animals, respectively. In conclusion, meldonium improves in vivo pyruvate dehydrogenase flux in the diabetic heart, contributing to improved cardiac recovery after ischemia., (© 2021 The Authors. NMR in Biomedicine published by John Wiley & Sons Ltd.)

The diabetic heart is energetically and metabolically abnormal, with increased fatty acid oxidation and decreased glucose oxidation. One factor contributing to the metabolic dysfunction in diabetes may be abnormal handling of acetyl and acyl groups by the mitochondria. L-carnitine is responsible for their transfer across the mitochondrial membrane, therefore, supplementation with L-carnitine may provide a route to improve the metabolic state of the diabetic heart. The primary aim of this study was to use hyperpolarized magnetic resonance imaging (MRI) to investigate the effects of L-carnitine supplementation on the in vivo metabolism of [1- 13 C]pyruvate in diabetes. Male Wistar rats were injected with either vehicle or streptozotocin (55 mg/kg) to induce type-1 diabetes. Three weeks of daily i.p. treatment with either saline or L-carnitine (3 g/kg/day) was subsequently undertaken. In vivo cardiac function and metabolism were assessed with CINE and hyperpolarized MRI, respectively. L-carnitine supplementation prevented the progression of hyperglycemia, which was observed in untreated streptozotocin injected animals and led to reductions in plasma triglyceride and ß-hydroxybutyrate concentrations. Hyperpolarized MRI revealed that L-carnitine treatment elevated pyruvate dehydrogenase flux by 3-fold in the diabetic animals, potentially through increased buffering of excess acetyl-CoA units in the mitochondria. Improved functional recovery following ischemia was also observed in the L-carnitine treated diabetic animals.

Purpose: Hyperpolarized imaging experiments have conflicting requirements of high spatial, temporal, and spectral resolution. Spectral-spatial RF excitation has been shown to form an attractive magnetization-efficient method for hyperpolarized imaging, but the optimum readout strategy is not yet known., Methods: In this work, we propose a novel 3D hybrid-shot spiral sequence which features two constant density regions that permit the retrospective reconstruction of either high spatial or high temporal resolution images post hoc, (adaptive spatiotemporal imaging) allowing greater flexibility in acquisition and reconstruction., Results: We have implemented this sequence, both via simulation and on a preclinical scanner, to demonstrate its feasibility, in both a 1 H phantom and with hyperpolarized 13 C pyruvate in vivo., Conclusions: This sequence forms an attractive method for acquiring hyperpolarized imaging datasets, providing adaptive spatiotemporal imaging to ameliorate the conflict of spatial and temporal resolution, with significant potential for clinical translation., (© 2020 The Authors. Magnetic Resonance in Medicine published by Wiley Periodicals LLC on behalf of International Society for Magnetic Resonance in Medicine.)

Objectives: To enhance detection of the products of hyperpolarized [2- 13 C]dihydroxyacetone metabolism for assessment of three metabolic pathways in the liver in vivo. Hyperpolarized [2- 13 C]DHAc emerged as a promising substrate to follow gluconeogenesis, glycolysis and the glycerol pathways. However, the use of [2- 13 C]DHAc in vivo has not taken off because (i) the chemical shift range of [2- 13 C]DHAc and its metabolic products span over 144 ppm, and (ii) 1 H decoupling is required to increase spectral resolution and sensitivity. While these issues are trivial for high-field vertical-bore NMR spectrometers, horizontal-bore small-animal MR scanners are seldom equipped for such experiments., Methods: Real-time hepatic metabolism of three fed mice was probed by 1 H-decoupled 13 C-MR following injection of hyperpolarized [2- 13 C]DHAc. The spectra of [2- 13 C]DHAc and its metabolic products were acquired in a 7 T small-animal MR scanner using three purpose-designed spectral-spatial radiofrequency pulses that excited a spatial bandwidth of 8 mm with varying spectral bandwidths and central frequencies (chemical shifts)., Results: The metabolic products detected in vivo include glycerol 3-phosphate, glycerol, phosphoenolpyruvate, lactate, alanine, glyceraldehyde 3-phosphate and glucose 6-phosphate. The metabolite-to-substrate ratios were comparable to those reported previously in perfused liver., Discussion: Three metabolic pathways can be probed simultaneously in the mouse liver in vivo, in real time,  using hyperpolarized DHAc.

Doxorubicin (DOX) is a widely used chemotherapeutic agent that can cause serious cardiotoxic side effects culminating in congestive heart failure (HF). There are currently no clinical imaging techniques or biomarkers available to detect DOX-cardiotoxicity before functional decline. Mitochondrial dysfunction is thought to be a key factor driving functional decline, though real-time metabolic fluxes have never been assessed in DOX-cardiotoxicity. Hyperpolarized magnetic resonance imaging (MRI) can assess real-time metabolic fluxes in vivo. Here we show that cardiac functional decline in a clinically relevant rat-model of DOX-HF is preceded by a change in oxidative mitochondrial carbohydrate metabolism, measured by hyperpolarized MRI. The decreased metabolic fluxes were predominantly due to mitochondrial loss and additional mitochondrial dysfunction, and not, as widely assumed hitherto, to oxidative stress. Since hyperpolarized MRI has been successfully translated into clinical trials this opens up the potential to test cancer patients receiving DOX for early signs of cardiotoxicity.

Nicotinic acid receptor agonists have previously been shown to cause acute reductions in cardiac contractility. We sought to uncover the changes in cardiac metabolism underlying these alterations in function. In nine humans, we recorded cardiac energetics and function before and after a single oral dose of nicotinic acid using cardiac MRI to demonstrate contractile function and Phosphorus-31 ( 31 P) magnetic resonance spectroscopy to demonstrate myocardial energetics. Nicotinic Acid 400 mg lowered ejection fraction by 4% (64 ± 8% to 60 ± 7%, P = .03), and was accompanied by a fall in phosphocreatine/ATP ratio by 0.4 (2.2 ± 0.4 to 1.8 ± 0.1, P = .04). In four groups of eight Wistar rats, we used pyruvate dehydrogenase (PDH) flux studies to demonstrate changes in carbohydrate metabolism induced by the nicotinic acid receptor agonist, Acipimox, using hyperpolarized Carbon-13 ( 13 C) magnetic resonance spectroscopy. In rats which had been starved overnight, Acipimox caused a fall in ejection fraction by 7.8% (67.5 ± 8.9 to 60 ± 3.1, P = .03) and a nearly threefold rise in flux through PDH (from 0.182 ± 0.114 to 0.486 ± 0.139, P = .002), though this rise did not match pyruvate dehydrogenase flux observed in rats fed carbohydrate rich chow (0.726 ± 0.201). In fed rats, Acipimox decreased pyruvate dehydrogenase flux (to 0.512 ± 0.13, P = .04). Concentration of plasma insulin fell by two-thirds in fed rats administered Acipimox (from 1695 ± 891 ng/L to 550 ± 222 ng/L, P = .005) in spite of glucose concentrations remaining the same. In conclusion, we demonstrate that nicotinic acid receptor agonists impair cardiac contractility associated with a decline in cardiac energetics and show that the mechanism is likely a combination of reduced fatty acid availability and a failure to upregulate carbohydrate metabolism, essentially starving the heart of fuel., (© 2020 The Authors. The FASEB Journal published by Wiley Periodicals LLC on behalf of Federation of American Societies for Experimental Biology.)

Cardiac energetic dysfunction has been reported in patients with type 2 diabetes (T2D) and is an independent predictor of mortality. Identification of the mechanisms driving mitochondrial dysfunction, and therapeutic strategies to rescue these modifications, will improve myocardial energetics in T2D. We demonstrate using 31P-magnetic resonance spectroscopy (31P-MRS) that decreased cardiac ATP and phosphocreatine (PCr) concentrations occurred before contractile dysfunction or a reduction in PCr/ATP ratio in T2D. Real-time mitochondrial ATP synthesis rates and state 3 respiration rates were similarly depressed in T2D, implicating dysfunctional mitochondrial energy production. Driving this energetic dysfunction in T2D was an increase in mitochondrial protein acetylation, and increased ex vivo acetylation was shown to proportionally decrease mitochondrial respiration rates. Treating T2D rats in vivo with the mitochondrial deacetylase SIRT3 activator honokiol reversed the hyperacetylation of mitochondrial proteins and restored mitochondrial respiration rates to control levels. Using 13C-hyperpolarized MRS, respiration with different substrates, and enzyme assays, we localized this improvement to increased glutamate dehydrogenase activity. Finally, honokiol treatment increased ATP and PCr concentrations and increased total ATP synthesis flux in the T2D heart. In conclusion, hyperacetylation drives energetic dysfunction in T2D, and reversing acetylation with the SIRT3 activator honokiol rescued myocardial and mitochondrial energetics in T2D.

Doxorubicin is a commonly used chemotherapeutic agent for the treatment of a range of cancers, but despite its success in improving cancer survival rates, doxorubicin is cardiotoxic and can lead to congestive heart failure. Therapeutic options for this patient group are limited to standard heart failure medications with the only drug specific for doxorubicin cardiotoxicity to reach FDA approval being dexrazoxane, an iron-chelating agent targeting oxidative stress. However, dexrazoxane has failed to live up to its expectations from preclinical studies while also bringing up concerns about its safety. Despite decades of research, the molecular mechanisms of doxorubicin cardiotoxicity are still poorly understood and oxidative stress is no longer considered to be the sole evil. Mitochondrial impairment, increased apoptosis, dysregulated autophagy and increased fibrosis have also been shown to be crucial players in doxorubicin cardiotoxicity. These cellular processes are all linked by one highly conserved intracellular kinase: adenosine monophosphate-activated protein kinase (AMPK). AMPK regulates mitochondrial biogenesis via PGC1α signalling, increases oxidative mitochondrial metabolism, decreases apoptosis through inhibition of mTOR signalling, increases autophagy through ULK1 and decreases fibrosis through inhibition of TGFβ signalling. AMPK therefore sits at the control point of many mechanisms shown to be involved in doxorubicin cardiotoxicity and cardiac AMPK signalling itself has been shown to be impaired by doxorubicin. In this review, we introduce different agents known to activate AMPK (metformin, statins, resveratrol, thiazolidinediones, AICAR, specific AMPK activators) as well as exercise and dietary restriction, and we discuss the existing evidence for their potential role in cardioprotection from doxorubicin cardiotoxicity.

Hyperpolarized [1- 13 C] pyruvate MRS can measure cardiac pyruvate dehydrogenase (PDH) flux in vivo through 13 C-label incorporation into bicarbonate. Using this technology, substrate availability as well as pathology have been shown to modulate PDH flux. Clinical protocols attempt to standardize PDH flux with oral glucose loading prior to scanning, while rodents in preclinical studies are usually scanned in the fed state. We aimed to establish which strategy was optimal to maximize PDH flux and minimize its variability in both control and Type II diabetic rats, without affecting the pathological variation being assessed. We found similar variances in the bicarbonate to pyruvate ratio, reflecting PDH flux, in fed and fasted/glucose-loaded animals, which showed no statistically significant differences. Furthermore, fasting/glucose loading did not alter the low PDH flux seen in Type II diabetic rats. Overall this suggests that preclinical cardiac hyperpolarized magnetic resonance studies could be performed either in the fed or in the fasted/glucose-loaded state. Centres planning to start new clinical studies with cardiac hyperpolarized magnetic resonance in man may find it beneficial to run small proof-of-concept trials to determine whether metabolic standardizations by oral or intravenous glucose load are beneficial compared with scanning patients in the fed state., (© 2018 The Authors. NMR in Biomedicine published by John Wiley & Sons Ltd.)

Cardiovascular disease is the leading cause of death world-wide. It is increasingly recognised that cardiac pathologies show, or may even be caused by, changes in metabolism, leading to impaired cardiac energetics. The heart turns over 15 times its own weight in ATP every day and thus relies heavily on the availability of substrates and on efficient oxidation to generate this ATP. A number of old and emerging drugs that target different aspects of metabolism are showing promising results with regard to improved cardiac outcomes in patients. A non-invasive imaging technique that could assess the role of different aspects of metabolism in heart disease, as well as measure changes in cardiac energetics due to treatment, would be valuable in the routine clinical care of cardiac patients. Hyperpolarised magnetic resonance spectroscopy and imaging have revolutionised metabolic imaging, allowing real-time metabolic flux assessment in vivo for the first time. In this review we summarise metabolism in the healthy and diseased heart, give an introduction to the hyperpolarisation technique, 'dynamic nuclear polarisation' (DNP), and review the preclinical studies that have thus far explored healthy cardiac metabolism and different models of human heart disease. We furthermore show what advances have been made to translate this technique into the clinic, what technical challenges still remain and what unmet clinical needs and unexplored metabolic substrates still need to be assessed by researchers in this exciting and fast-moving field., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Rapid cancer cell proliferation promotes the production of reducing equivalents, which counteract the effects of relatively high levels of reactive oxygen species. Reactive oxygen species levels increase in response to chemotherapy and cell death, whereas an increase in antioxidant capacity can confer resistance to chemotherapy and is associated with an aggressive tumor phenotype. The pentose phosphate pathway is a major site of NADPH production in the cell, which is used to maintain the main intracellular antioxidant, glutathione, in its reduced state. Previous studies have shown that the rate of hyperpolarized [1- 13 C]dehydroascorbic acid (DHA) reduction, which can be measured in vivo using non-invasive 13 C magnetic resonance spectroscopic imaging, is increased in tumors and that this is correlated with the levels of reduced glutathione. We show here that the rate of hyperpolarized [1- 13 C]DHA reduction is increased in tumors that have been oxidatively prestressed by depleting the glutathione pool by buthionine sulfoximine treatment. This increase was associated with a corresponding increase in pentose phosphate pathway flux, assessed using 13 C-labeled glucose, and an increase in glutaredoxin activity, which catalyzes the glutathione-dependent reduction of DHA. These results show that the rate of DHA reduction depends not only on the level of reduced glutathione, but also on the rate of NADPH production, contradicting the conclusions of some previous studies. Hyperpolarized [1- 13 C]DHA can be used, therefore, to assess the capacity of tumor cells to resist oxidative stress in vivo However, DHA administration resulted in transient respiratory arrest and cardiac depression, which may prevent translation to the clinic., (© 2017 by The American Society for Biochemistry and Molecular Biology, Inc.)

Changes in tumor metabolism may accompany disease progression and can occur following treatment, often before there are changes in tumor size. We focus here on imaging methods that can be used to image various aspects of tumor metabolism, with an emphasis on methods that can be used for tumor grading, assessing disease progression, and monitoring treatment response. Clin Cancer Res; 22(21); 5196-203. ©2016 AACR., (©2016 American Association for Cancer Research.)

Purpose: Dissolution dynamic nuclear polarization can increase the sensitivity of the (13) C magnetic resonance spectroscopy experiment by at least four orders of magnitude and offers a novel approach to the development of MRI gene reporters based on enzymes that metabolize (13) C-labeled tracers. We describe here a gene reporter based on the enzyme pyruvate decarboxylase (EC 4.1.1.1), which catalyzes the decarboxylation of pyruvate to produce acetaldehyde and carbon dioxide., Methods: Pyruvate decarboxylase from Zymomonas mobilis (zmPDC) and a mutant that lacked enzyme activity were expressed using an inducible promoter in human embryonic kidney (HEK293T) cells. Enzyme activity was measured in the cells and in xenografts derived from the cells using (13) C MRS measurements of the conversion of hyperpolarized [1-(13) C] pyruvate to H(13) CO3-., Results: Induction of zmPDC expression in the cells and in the xenografts derived from them resulted in an approximately two-fold increase in the H(13) CO3-/[1-(13) C] pyruvate signal ratio following intravenous injection of hyperpolarized [1-(13) C] pyruvate., Conclusion: We have demonstrated the feasibility of using zmPDC as an in vivo reporter gene for use with hyperpolarized (13) C MRS. Magn Reson Med 76:391-401, 2016. © 2015 The Authors. Magnetic Resonance in Medicine published by Wiley Periodicals, Inc. on behalf of International Society for Magnetic Resonance in Medicine. This is an open access article under the terms of the Creative Commons Attribution License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited., (© 2015 The Authors. Magnetic Resonance in Medicine published by Wiley Periodicals, Inc. on behalf of International Society for Magnetic Resonance in Medicine.)

Purpose: A resonance at ∼181 ppm in the (13) C spectra of tumors injected with hyperpolarized [U-(2) H, U-(13) C]glucose was assigned to 6-phosphogluconate (6PG), as in previous studies in yeast, whereas in breast cancer cells in vitro this resonance was assigned to 3-phosphoglycerate (3PG). These peak assignments were investigated here using measurements of 6PG and 3PG (13) C-labeling using liquid chromatography tandem mass spectrometry (LC-MS/MS) METHODS: Tumor-bearing mice were injected with (13) C6 glucose and the (13) C-labeled and total 6PG and 3PG concentrations measured. (13) C MR spectra of glucose-6-phosphate dehydrogenase deficient (zwf1Δ) and wild-type yeast were acquired following addition of hyperpolarized [U-(2) H, U-(13) C]glucose and again (13) C-labeled and total 6PG and 3PG were measured by LC-MS/MS RESULTS: Tumor (13) C-6PG was more abundant than (13) C-2PG/3PG and the resonance at ∼181 ppm matched more closely that of 6PG. (13) C MR spectra of wild-type and zwf1Δ yeast cells showed a resonance at ∼181 ppm after labeling with hyperpolarized [U-(2) H, U-(13) C]glucose, however, there was no 6PG in zwf1Δ cells. In the wild-type cells 3PG was approximately four-fold more abundant than 6PG CONCLUSION: The resonance at ∼181 ppm in (13) C MR spectra following injection of hyperpolarized [U-(2) H, U-(13) C]glucose originates predominantly from 6PG in EL4 tumors and 3PG in yeast cells., (© 2014 Wiley Periodicals, Inc.)

Purpose: Aldehyde dehydrogenase (ALDH2) is an emerging drug target for the treatment of heart disease, cocaine and alcohol dependence, and conditions caused by genetic polymorphisms in ALDH2. Noninvasive measurement of ALDH2 activity in vivo could inform the development of these drugs and accelerate their translation to the clinic., Methods: [1-(13) C, U-(2) H5 ] ethanol was hyperpolarized using dynamic nuclear polarization, injected into mice and its oxidation in the liver monitored using (13) C MR spectroscopy and spectroscopic imaging., Results: Oxidation of [1-(13) C, U-(2) H5 ] ethanol to [1-(13) C] acetate was observed. Saturation of the acetaldehyde resonance, which was below the level of detection in vivo, demonstrated that acetate was produced via acetaldehyde. Irreversible inhibition of ALDH2 activity with disulfiram resulted in a proportional decrease in the amplitude of the acetate resonance., Conclusion: (13) C magnetic resonance spectroscopy measurements of hyperpolarized [1-(13) C, U-(2) H5 ] ethanol oxidation allow real-time assessment of ALDH2 activity in liver in vivo., (© 2014 The Authors. Magnetic Resonance in Medicine Published by Wiley Periodicals, Inc. on behalf of International Society of Medicine in Resonance.)

Purpose: To assess the potential of a gene reporter system, based on a urea transporter (UTB) and hyperpolarized [(13) C]urea., Methods: Mice were implanted subcutaneously with either unmodified control cells or otherwise identical cells expressing UTB. After injection of hyperpolarized [(13) C]urea, a spin echo sequence was used to measure urea concentration, T1 , and diffusion in control and UTB-expressing tissue., Results: The apparent diffusion coefficient of hyperpolarized urea was 21% lower in tissue expressing UTB, in comparison with control tissue (P < 0.05, 1-tailed t-test, n = 6 in each group). No difference in water apparent diffusion coefficient or cellularity between these tissues was found, indicating that they were otherwise similar in composition., Conclusion: Expression of UTB, by mediating cell uptake of urea, lowers the apparent diffusion coefficient of hyperpolarized (13) C urea in tissue and thus the transporter has the potential to be used as a magnetic resonance-based gene reporter in vivo. Magn Reson Med 73:1401-1406, 2015. © 2014 Wiley Periodicals, Inc., (© 2014 Wiley Periodicals, Inc.)

Hyperpolarized NMR is a promising technique for non-invasive imaging of tissue metabolism in vivo. However, the pathways that can be studied are limited by the fast T1 decay of the nuclear spin order. In metabolites containing pairs of coupled nuclear spins-1/2, the spin order may be maintained by exploiting the non-magnetic singlet (spin-0) state of the pair. This may allow preservation of the hyperpolarization in vivo during transport to tissues of interest, such as tumors, or to detect slower metabolic reactions. We show here that in human blood and in a mouse in vivo at millitesla fields the (13)C singlet lifetime of [1,2-(13)C2]pyruvate was significantly longer than the (13)C T1, although it was shorter than the T1 at field strengths of several tesla. We also examine the singlet-derived NMR spectrum observed for hyperpolarized [1,2-(13)C2]lactate, originating from the metabolism of [1,2-(13)C2]pyruvate., (© 2013 The Authors. NMR in Biomedicine published by John Wiley & Sons, Ltd.)