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Cataracts are a disease that reduces vision due to opacity formation of the lens. Diabetic cataracts occur at young age and progress relatively quickly, so the development of effective treatment has been awaited. Several studies have shown that pyruvate inhibits oxidative stress and glycation of lens proteins, which contribute to onset of diabetic cataracts. However, detailed molecular mechanisms have not been revealed. In this study, we attempted to reduce galactose‐induced opacity by pyruvate with rat ex vivo model. Rat lenses were extracted and cultured in galactose‐containing medium to induce lens opacity. After opacity had developed, continued culturing with pyruvate in the medium resulted in a reduction of lens opacity. Subsequently, we conducted microarray analysis to investigate the genes that contribute to the therapeutic effect. We performed quantitative expression measurements using RT‐qPCR for extracted genes that were upregulated in cataract‐induced lenses and downregulated in pyruvate‐treated lenses, resulting in the identification of 34 candidate genes. Functional analysis using the STRING database suggests that metallothionein‐related factors (Mt1a, Mt1m, and Mt2A) and epithelial‐mesenchymal transition‐related factors (Acta2, Anxa1, Cd81, Mki67, Timp1, and Tyms) contribute to the therapeutic effect of cataracts. [ABSTRACT FROM AUTHOR]

The addition of antioxidants to cryopreservation media reportedly improves sperm post‐thaw quality and reproductive performance after artificial insemination. Therefore, the objectives of this study were to evaluate if the addition of L‐carnitine and pyruvate to freezing media, or their addition to samples after thawing, improves the post‐thaw quality of equine spermatozoa. Thus, in Experiment 1, stallion semen samples were cryopreserved in: (1) EDTA–glucose–based extender with 20% egg yolk and 5% dimethylformamide (EDTA control); (2) skim milk–based extender with 20% egg yolk and 5% dimethylformamide (milk control); (3) Extender 1 supplemented with 50 mM L‐carnitine and 10 mM pyruvate (EDTA‐carnitine‐pyruvate); and (4) Extender 2 supplemented with 50 mM L‐carnitine and 10 mM pyruvate (milk‐carnitine‐pyruvate). In Experiment 2, 50 mM L‐carnitine and 10 mM pyruvate were added post‐thaw to samples cryopreserved with extenders 1 and 2 (EDTA control and milk control). Sperm kinematic parameters, DNA fragmentation, membrane lipid peroxidation, acrosome status and viability were evaluated after thawing. No significant differences (p > 0.05) were observed for most of the kinematic parameters, DNA fragmentation, membrane lipid peroxidation, acrosome status and viability of spermatozoa, between the samples frozen in the presence or absence of L‐carnitine and pyruvate, nor between the samples after the post‐thaw addition of these components. A higher (p < 0.05) mean velocity and higher (p < 0.05) amplitude of lateral head displacement were observed in the samples frozen in the milk‐based extender with the addition of L‐carnitine and pyruvate after thawing. The addition of 50 mM L‐carnitine and 10 mM pyruvate, either to the freezing extenders or after thawing, was not deleterious for sperm; however, it did not improve equine sperm motility, viability, acrosome and DNA integrity, nor decrease membrane lipid peroxidation after thawing. [ABSTRACT FROM AUTHOR]

Purpose: We aimed to incorporate a deep learning prior with k‐space data fidelity for accelerating hyperpolarized carbon‐13 MRSI, demonstrated on synthetic cancer datasets. Methods: A two‐site exchange model, derived from the Bloch equation of MR signal evolution, was firstly used in simulating training and testing data, that is, synthetic phantom datasets. Five singular maps generated from each simulated dataset were used to train a deep learning prior, which was then employed with the fidelity term to reconstruct the undersampled MRI k‐space data. The proposed method was assessed on synthetic human brain tumor images (N = 33), prostate cancer images (N = 72), and mouse tumor images (N = 58) for three undersampling factors and 2.5% additive Gaussian noise. Furthermore, varied levels of Gaussian noise with SDs of 2.5%, 5%, and 10% were added on synthetic prostate cancer data, and corresponding reconstruction results were evaluated. Results: For quantitative evaluation, peak SNRs were approximately 32 dB, and the accuracy was generally improved for 5 to 8 dB compared with those from compressed sensing with L1‐norm regularization or total variation regularization. Reasonable normalized RMS error were obtained. Our method also worked robustly against noise, even on a data with noise SD of 10%. Conclusion: The proposed singular value decomposition + iterative deep learning model could be considered as a general framework that extended the application of deep learning MRI reconstruction to metabolic imaging. The morphology of tumors and metabolic images could be measured robustly in six times acceleration using our method. [ABSTRACT FROM AUTHOR]

A 3‐year‐old male neutered domestic shorthair cat presented with lethargy, hyporexia, and pyrexia of unknown origin. Biochemical analysis using a Beckman Coulter AU480 demonstrated marked increases in creatine kinase and aspartate aminotransferase, indicative of severe muscle injury, with concurrent presumptive myoglobinuria on urinalysis. A marked, non‐physiologic increase in measured bicarbonate and resultant negative anion gap was documented; however, calculated bicarbonate obtained via a point‐of‐care blood gas analyzer was within normal limits. Laboratory error due to interference by lactate dehydrogenase was suspected and supported by the results of subsequent biochemical testing. Artifactual increases in bicarbonate have been documented in cases of rhabdomyolysis in horses, cows, and a bird. However, to the best of the authors' knowledge, this is the first report to demonstrate this spurious change in a cat. [ABSTRACT FROM AUTHOR]

Background: Pancreatic ductal adenocarcinoma (PDA) is the third leading cause of cancer‐related death in the United States. However, early response assessment using the current approach of measuring changes in tumor size on computed tomography (CT) or MRI is challenging. Purpose: To investigate the feasibility of hyperpolarized (HP) [1‐13C]pyruvate MRI to quantify metabolism in the normal appearing pancreas and PDA, and to assess changes in PDA metabolism following systemic chemotherapy. Study Type: Prospective. Subjects: Six patients (65.0 ± 7.6 years, 2 females) with locally advanced or metastatic PDA enrolled prior to starting a new line of systemic chemotherapy. Field Strength/Sequence: 3‐T, T1‐weighted gradient echo, metabolite‐selective 13C echoplanar imaging. Assessment: Time‐resolved HP [1‐13C]pyruvate data were acquired before (N = 6) and 4‐weeks after (N = 3) treatment initiation. Pyruvate metabolism, as quantified by pharmacokinetic modeling and metabolite area‐under‐the‐curve ratios, was assessed in manually segmented PDA and normal appearing pancreas ROIs (N = 5). The change in tumor metabolism before and 4‐weeks after treatment initiation was assessed in primary PDA (N = 2) and liver metastases (N = 1), and was compared to objective tumor response defined by response evaluation criteria in solid tumors (RECIST) on subsequent CTs. Statistical Tests: Descriptive tests (mean ± standard deviation), model fit error for pharmacokinetic rate constants. Results: Primary PDA showed reduced alanine‐to‐lactate ratios when compared to normal pancreas, due to increased lactate‐to‐pyruvate or reduced alanine‐to‐pyruvate ratios. Of the three patients who received HP [1‐13C]pyruvate MRI before and 4‐weeks after treatment initiation, one patient had a primary tumor with early metabolic response (increase in alanine‐to‐lactate) and subsequent partial response according to RECIST, one patient had a primary tumor with relatively stable metabolism and subsequent stable disease by RECIST, and one patient had metastatic PDA with increase in lactate‐to‐pyruvate of the liver metastases and corresponding progressive disease according to RECIST. Data Conclusion: Altered pyruvate metabolism with increased lactate or reduced alanine was observed in the primary tumor. Early metabolic response assessed at 4‐weeks after treatment initiation correlated with subsequent objective tumor response assessed using RECIST. Level of Evidence: 2 Technical Efficacy: Stage 2 [ABSTRACT FROM AUTHOR]

Background: Most commerce of equine seminal doses is carried out using commercial extenders under refrigeration at 5°C. Objectives: To determine if 10 mm pyruvate in a 67 mm glucose extender and storage at 22°C could be the basis of an alternative storage method to cooling to 5°C. Material and methods: Stallion ejaculates were extendedin: INRA96 (67 mm glucose, non‐pyruvate control), modified Tyrode's (67 mm glucose–10 mm pyruvate), supplemented with 0, 10, 50, and 100 μM itaconate. As itaconate was vehiculated in DMSO, a control vehicle was also included. Sperm motility, viability, mitochondrial membrane potential, and production of reactive oxygen species were measured after collection and again after 48 and 96 h of storage at 22°C. To disclose molecular metabolic changes, spermatozoa were incubated up to 3 h in modified Tyrode's 67 mm glucose–10 mm pyruvate and modified Tyrode's 67 mm glucose, and metabolic analysis conducted. Results: After 96 h of storage aliquots stored in the control, INRA96 had a very poor total motility of 5.6% ± 2.3%, while in the 67 mm glucose–10 mm pyruvate/10 μm itaconate extender, total motility was 34.7% ± 3.8% (p = 0.0066). After 96 h, viability was better in most pyruvate‐based media, and the mitochondrial membrane potential in spermatozoa extended in INRA96 was relatively lower (p < 0.0001). Metabolomics revealed that in the spermatozoa incubated in the high pyruvate media, there was an increase in the relative amounts of NAD+, pyruvate, lactate, and ATP. Discussion and conclusions: Aliquots stored in a 67 mm glucose–10 mm pyruvate‐based medium supplemented with 10 μM itaconate, maintained a 35% total motility after 96 h of storage at 22°C, which is considered the minimum acceptable motility for commercialization. Improvements may be related to the conversion of pyruvate to lactate and regeneration of NAD+. [ABSTRACT FROM AUTHOR]

MRI with hyperpolarized (HP) 13C agents, also known as HP 13CMRI, can measure processes such as localized metabolism that is altered in numerous cancers, liver, heart, kidney diseases, and more. It has been translated into human studies during the past 10 years, with recent rapid growth in studies largely based on increasing availability of HP agent preparation methods suitable for use in humans. This paper aims to capture the current successful practices for HP MRI human studies with [1-13C]pyruvate--by far the most commonly used agent, which sits at a keymetabolic junction in glycolysis. The paper is divided into four major topic areas: (1) HP 13C-pyruvate preparation; (2) MRI system setup and calibrations; (3) data acquisition and image reconstruction; and (4) data analysis and quantification. In each area, we identified the key components for a successful study, summarized both published studies and current practices, and discuss evidence gaps, strengths, and limitations. This paper is the output of the "HP 13C MRI Consensus Group" as well as the ISMRM Hyperpolarized Media MR and Hyperpolarized Methods and Equipment study groups. It further aims to provide a comprehensive reference for future consensus, building as the field continues to advance human studies with this metabolic imaging modality. [ABSTRACT FROM AUTHOR]

Purpose: Improving the quality and maintaining the fidelity of large coverage abdominal hyperpolarized (HP) 13C MRI studies with a patch based global–local higher‐order singular value decomposition (GL‐HOVSD) spatiotemporal denoising approach. Methods: Denoising performance was first evaluated using the simulated [1‐13C]pyruvate dynamics at different noise levels to determine optimal kglobal and klocal parameters. The GL‐HOSVD spatiotemporal denoising method with the optimized parameters was then applied to two HP [1‐13C]pyruvate EPI abdominal human cohorts (n = 7 healthy volunteers and n = 8 pancreatic cancer patients). Results: The parameterization of kglobal = 0.2 and klocal = 0.9 denoises abdominal HP data while retaining image fidelity when evaluated by RMSE. The kPX (conversion rate of pyruvate‐to‐metabolite, X = lactate or alanine) difference was shown to be <20% with respect to ground‐truth metabolic conversion rates when there is adequate SNR (SNRAUC > 5) for downstream metabolites. In both human cohorts, there was a greater than nine‐fold gain in peak [1‐13C]pyruvate, [1‐13C]lactate, and [1‐13C]alanine apparent SNRAUC. The improvement in metabolite SNR enabled a more robust quantification of kPL and kPA. After denoising, we observed a 2.1 ± 0.4 and 4.8 ± 2.5‐fold increase in the number of voxels reliably fit across abdominal FOVs for kPL and kPA quantification maps. Conclusion: Spatiotemporal denoising greatly improves visualization of low SNR metabolites particularly [1‐13C]alanine and quantification of [1‐13C]pyruvate metabolism in large FOV HP 13C MRI studies of the human abdomen. [ABSTRACT FROM AUTHOR]

Despite the significant role of pyruvic acid in biotransformations, the utilization of pyruvates in the field of catalytic asymmetric synthesis has been unexpectedly limited. This statement also applies to enamine catalysis, in which pyruvates seem to be challenging substrates. Recently, the first illustration of the asymmetric addition of pyruvic acid to β‐nitrostyrenes, catalyzed by a NahE enzyme using the enamine mechanism, has appeared in the literature. Now, we demonstrate pyrrolidine‐based catalyst can mimick enamine mechanism used by this enzyme to activate pyruvates in the asymmetric addition to structurally diverse nitroalkenes to produce highly enantioenriched Michael adducts without the limitations in substrate structures associated with biocatalysts. In addition to confirming that optically pure pyrrolidines can serve as ultimately small organic catalysts in pyruvate activation, we present an exceedingly simple and versatile method for the synthesis of optically pure γ‐nitro acids, which are precursors for well‐known active pharmaceutical ingredients namely GABA derivatives [ABSTRACT FROM AUTHOR]

Introduction: The healthy heart has remarkable metabolic flexibility that permits rapid switching between mitochondrial glucose oxidation and fatty acid oxidation to generate ATP. Loss of metabolic flexibility has been implicated in the genesis of contractile dysfunction seen in cardiomyopathy. Metabolic flexibility has been imaged in experimental models, using hyperpolarized (HP) [2‐13C]pyruvate MRI, which enables interrogation of metabolites that reflect tricarboxylic acid (TCA) cycle flux in cardiac myocytes. This study aimed to develop methods, demonstrate feasibility for [2‐13C]pyruvate MRI in the human heart for the first time, and assess cardiac metabolic flexibility. Methods: Good manufacturing practice [2‐13C]pyruvic acid was polarized in a 5 T polarizer for 2.5–3 h. Following dissolution, quality control parameters of HP pyruvate met all safety and sterility criteria for pharmacy release, prior to administration to study subjects. Three healthy subjects each received two HP injections and MR scans, first under fasting conditions, followed by oral glucose load. A 5 cm axial slab‐selective spectroscopy approach was prescribed over the left ventricle and acquired at 3 s intervals on a 3 T clinical MRI scanner. Results: The study protocol, which included HP substrate injection, MR scanning, and oral glucose load, was performed safely without adverse events. Key downstream metabolites of [2‐13C]pyruvate metabolism in cardiac myocytes include the glycolytic derivative [2‐13C]lactate, TCA‐associated metabolite [5‐13C]glutamate, and [1‐13C]acetylcarnitine, catalyzed by carnitine acetyltransferase (CAT). After glucose load, 13C‐labeling of lactate, glutamate, and acetylcarnitine from 13C‐pyruvate increased by an average of 39.3%, 29.5%, and 114% respectively in the three subjects, which could result from increases in lactate dehydrogenase, pyruvate dehydrogenase, and CAT enzyme activity as well as TCA cycle flux (glucose oxidation). Conclusions: HP [2‐13C]pyruvate imaging is safe and permits noninvasive assessment of TCA cycle intermediates and the acetyl buffer, acetylcarnitine, which is not possible using HP [1‐13C]pyruvate. Cardiac metabolite measurement in the fasting/fed states provides information on cardiac metabolic flexibility and the acetylcarnitine pool. [ABSTRACT FROM AUTHOR]

The goal of this study was to investigate the origin of brain lactate (Lac) signal in the healthy anesthetized rat after injection of hyperpolarized (HP) [1‐13C]pyruvate (Pyr). Dynamic two‐dimensional spiral chemical shift imaging with flow‐sensitizing gradients revealed reduction in both vascular and brain Pyr, while no significant dependence on the level of flow suppression was detected for Lac. These results support the hypothesis that the HP metabolites predominantly reside in different compartments in the brain (i.e., Pyr in the blood and Lac in the parenchyma). Data from high‐resolution metabolic imaging of [1‐13C]Pyr further demonstrated that Lac detected in the brain was not from contributions of vascular signal attributable to partial volume effects. Additionally, metabolite distributions and kinetics measured with dynamic imaging after injection of HP [1‐13C]Lac were similar to Pyr data when Pyr was used as the substrate. These data do not support the hypothesis that Lac observed in the brain after Pyr injection was generated in other organs and then transported across the blood–brain barrier (BBB). Together, the presented results provide further evidence that even in healthy anesthetized rats, the transport of HP Pyr across the BBB is sufficiently fast to permit detection of its metabolic conversion to Lac within the brain. [ABSTRACT FROM AUTHOR]

d‐Serine plays vital physiological roles in the functional regulation of the mammalian brain, where it is produced from l‐serine by serine racemase and degraded by d‐amino acid oxidase. In the present study, we identified a new d‐serine metabolizing activity of serine hydroxymethyltransferase (SHMT) in bacteria as well as mammals. SHMT is known to catalyze the conversion of l‐serine and tetrahydrofolate (THF) to glycine and 5,10‐methylenetetrahydrofolate, respectively. In addition, we found that human and Escherichia coli SHMTs have d‐serine dehydratase activity, which degrades d‐serine to pyruvate and ammonia. We characterized this enzymatic activity along with canonical SHMT activity. Intriguingly, SHMT required THF to catalyze d‐serine dehydration and did not exhibit dehydratase activity toward l‐serine. Furthermore, SHMT did not use d‐serine as a substrate in the canonical hydroxymethyltransferase reaction. The d‐serine dehydratase activities of two isozymes of human SHMT were inhibited in the presence of a high concentration of THF, whereas that of E. coli SHMT was increased. The pH and temperature profiles of d‐serine dehydratase and serine hydroxymethyltransferase activities of these three SHMTs were partially distinct. The catalytic efficiency (kcat/Km) of dehydratase activity was lower than that of hydroxymethyltransferase activity. Nevertheless, the d‐serine dehydratase activity of SHMT was physiologically important because d‐serine inhibited the growth of an SHMT deletion mutant of E. coli, ∆glyA, more than that of the wild‐type strain. Collectively, these results suggest that SHMT is involved not only in l‐ but also in d‐serine metabolism through the degradation of d‐serine. [ABSTRACT FROM AUTHOR]

In the present study, cryo‐electron tomography was used to investigate the localization of 2‐oxoacid dehydrogenase complexes (OADCs) in cardiac mitochondria and mitochondrial inner membrane samples. Two classes of ordered OADC inner cores with different symmetries were distinguished and their quaternary structures modeled. One class corresponds to pyruvate dehydrogenase complexes and the other to dehydrogenase complexes of α‐ketoglutarate and branched‐chain α‐ketoacids. OADCs were shown to be localized in close proximity to membrane‐embedded respirasomes, as observed both in densely packed lamellar cristae of cardiac mitochondria and in ruptured mitochondrial samples where the dense packing is absent. This suggests the specificity of the OADC–respirasome interaction, which allows localized NADH/NAD+ exchange between OADCs and complex I of the respiratory chain. The importance of this local coupling is based on OADCs being the link between respiration, glycolysis and amino acid metabolism. The coupling of these basic metabolic processes can vary in different tissues and conditions and may be involved in the development of various pathologies. The present study shows that this important and previously missing parameter of mitochondrial complex coupling can be successfully assessed using cryo‐electron tomography. [ABSTRACT FROM AUTHOR]

Enhancing magnetic resonance signal via hyperpolarization techniques enables the real‐time detection of metabolic transformations even in vivo. The use of para‐hydrogen to enhance 13C‐enriched metabolites has opened a rapid pathway for the production of hyperpolarized metabolites, which usually requires specialized equipment. Metabolite precursors that can be hyperpolarized and converted into metabolites at any given field would open up opportunities for many labs to make use of this technology because already existing hardware could be used. We report here on the complete synthesis and hyperpolarization of suitable precursor molecules of the side‐arm hydrogenation approach. The better accessibility to such side‐arms promises that the para‐hydrogen approach can be implemented in every lab with existing two channel NMR spectrometers for 1H and 13C independent of the magnetic field. [ABSTRACT FROM AUTHOR]

Purpose: To investigate high‐resolution hyperpolarized (HP) 13C pyruvate MRI for measuring cerebral perfusion in the human brain. Methods: HP [1‐13C]pyruvate MRI was acquired in five healthy volunteers with a multi‐resolution EPI sequence with 7.5 × 7.5 mm2 resolution for pyruvate. Perfusion parameters were calculated from pyruvate MRI using block‐circulant singular value decomposition and compared to relative cerebral blood flow calculated from arterial spin labeling (ASL). To examine regional perfusion patterns, correlations between pyruvate and ASL perfusion were performed for whole brain, gray matter, and white matter voxels. Results: High resolution 7.5 × 7.5 mm2 pyruvate images were used to obtain relative cerebral blood flow (rCBF) values that were significantly positively correlated with ASL rCBF values (r = 0.48, 0.20, 0.28 for whole brain, gray matter, and white matter voxels respectively). Whole brain voxels exhibited the highest correlation between pyruvate and ASL perfusion, and there were distinct regional patterns of relatively high ASL and low pyruvate normalized rCBF found across subjects. Conclusion: Acquiring HP 13C pyruvate metabolic images at higher resolution allows for finer spatial delineation of brain structures and can be used to obtain cerebral perfusion parameters. Pyruvate perfusion parameters were positively correlated to proton ASL perfusion values, indicating a relationship between the two perfusion measures. This HP 13C study demonstrated that hyperpolarized pyruvate MRI can assess cerebral metabolism and perfusion within the same study. [ABSTRACT FROM AUTHOR]

Parahydrogen‐induced polarization (PHIP) followed by polarization transfer to 13C is a rapidly developing technique for the generation of 13C‐hyperpolarized substrates. Chirality plays an essential role in living systems and differential metabolism of enantiomeric pairs of metabolic substrates is well documented. Inspired by asymmetric hydrogenation, here we report stereoPHIP, which involves the addition of parahydrogen to a prochiral substrate with a chiral catalyst followed by polarization transfer to 13C spins. We demonstrate that parahydrogen could be rapidly added to the prochiral precursor to both enantiomers of lactic acid (D and L), with both the (R,R) and (S,S) enantiomers of a chiral rhodium(I) catalyst to afford highly 13C‐hyperpolarized (over 20 %) L‐ and D‐lactate ester derivatives, respectively, with excellent stereoselectivity. We also show that the hyperpolarized 1H signal decays obtained with the (R,R) and (S,S) catalysts were markedly different. StereoPHIP expands the scope of conventional PHIP to the production of 13C hyperpolarized chiral substrates with high stereoselectivity. [ABSTRACT FROM AUTHOR]

Previous research suggests potential mitochondrial dysfunction and changes in fatty acid metabolism in a subgroup of individuals with autism spectrum disorder (ASD), indicated by higher lactate, pyruvate levels, and mitochondrial disorder prevalence. This study aimed to further investigate potential mitochondrial dysfunction in ASD by assessing blood metabolite levels linked to mitochondrial metabolism. Blood levels of creatine kinase (CK), alanine aminotransferase (ALT), aspartate aminotransferase (AST), lactate, pyruvate, free and total carnitine, as well as acylcarnitines were obtained in 73 adults with ASD (47 males, 26 females) and compared with those of 71 neurotypical controls (NTC) (44 males, 27 females). Correlations between blood parameters and psychometric ASD symptom scores were also explored. Lower CK (pcorr = 0.045) levels were found exclusively in males with ASD compared to NTC, with no such variation in females. ALT and AST levels did not differ significantly between both groups. After correction for antipsychotic and antidepressant medication, CK remained significant. ASD participants had lower serum lactate levels (pcorr = 0.036) compared to NTC, but pyruvate and carnitine concentrations showed no significant difference. ASD subjects had significantly increased levels of certain acylcarnitines, with a decrease in tetradecadienoyl‐carnitine (C14:2), and certain acylcarnitines correlated significantly with autistic symptom scores. We found reduced serum lactate levels in ASD, in contrast to previous studies suggesting elevated lactate or pyruvate. This difference may reflect the focus of our study on high‐functioning adults with ASD, who are likely to have fewer secondary genetic conditions associated with mitochondrial dysfunction. Our findings of significantly altered acylcarnitine levels in ASD support the hypothesis of altered fatty acid metabolism in a subset of ASD patients. Lay Summary: This study examined potential differences in energy metabolism in adults with autism spectrum disorder (ASD) and neurotypical controls (NTC), focusing on the function of mitochondria, the "powerhouses" of our cells. In contrast to previous research that reported elevated levels of lactate, an indicator of mitochondrial dysfunction, the study found lower levels of lactate in individuals with ASD compared to NTC, possibly due to a focus on adults with high‐functioning ASD. The study also reported alterations in fatty acid metabolism in ASD, as evidenced by specific changes in compounds called acylcarnitines, but more research is needed to understand the causes and implications of these changes. [ABSTRACT FROM AUTHOR]

Hyperpolarization techniques increase nuclear spin polarization by more than four orders of magnitude, enabling metabolic MRI. Even though hyperpolarization has shown clear value in clinical studies, the complexity, cost and slowness of current equipment limits its widespread use. Here, a polarization procedure of [1‐13C]pyruvate based on parahydrogen‐induced polarization by side‐arm hydrogenation (PHIP‐SAH) in an automated polarizer is demonstrated. It is benchmarked in a study with 48 animals against a commercial dissolution dynamic nuclear polarization (d‐DNP) device. Purified, concentrated (≈70–160 mM) and highly hyperpolarized (≈18%) solutions of pyruvate are obtained at physiological pH for volumes up to 2 mL within 85 s in an automated process. The safety profile, image quality, as well as the quantitative perfusion and lactate‐to‐pyruvate ratios, are equivalent for PHIP and d‐DNP, rendering PHIP a viable alternative to established hyperpolarization techniques. [ABSTRACT FROM AUTHOR]

Metabolic magnetic resonance imaging (MRI) using hyperpolarized (HP) pyruvate is becoming a non‐invasive technique for diagnosing, staging, and monitoring response to treatment in cancer and other diseases. The clinically established method for producing HP pyruvate, dissolution dynamic nuclear polarization, however, is rather complex and slow. Signal Amplification By Reversible Exchange (SABRE) is an ultra‐fast and low‐cost method based on fast chemical exchange. Here, for the first time, we demonstrate not only in vivo utility, but also metabolic MRI with SABRE. We present a novel routine to produce aqueous HP [1‐13C]pyruvate‐d3 for injection in 6 minutes. The injected solution was sterile, non‐toxic, pH neutral and contained ≈30 mM [1‐13C]pyruvate‐d3 polarized to ≈11 % (residual 250 mM methanol and 20 μM catalyst). It was obtained by rapid solvent evaporation and metal filtering, which we detail in this manuscript. This achievement makes HP pyruvate MRI available to a wide biomedical community for fast metabolic imaging of living organisms. [ABSTRACT FROM AUTHOR]

Purpose: To develop a high spatiotemporal resolution 3D dynamic pulse sequence for preclinical imaging of hyperpolarized [1‐13C]pyruvate‐to‐[1‐13C]lactate metabolism at 7T. Methods: A standard 3D balanced SSFP (bSSFP) sequence was modified to enable alternating‐frequency excitations. RF pulses with 2.33 ms duration and 900 Hz FWHM were placed off‐resonance of the target metabolites, [1‐13C]pyruvate (by approximately −245 Hz) and [1‐13C]lactate (by approximately 735 Hz), to selectively excite those resonances. Relatively broad bandwidth (compared to those metabolites' chemical shift offset) permits a short TR of 6.29 ms, enabling higher spatiotemporal resolution. Bloch equation simulations of the bSSFP response profile guided the sequence parameter selection to minimize spectral contamination between metabolites and preserve magnetization over time. Results: Bloch equation simulations, phantom studies, and in vivo studies demonstrated that the two target resonances could be cleanly imaged without substantial bSSFP banding artifacts and with little spectral contamination between lactate and pyruvate and from pyruvate hydrate. High spatiotemporal resolution 3D images were acquired of in vivo pyruvate‐lactate metabolism in healthy wild‐type and endogenous pancreatic tumor‐bearing mice, with 1.212 s acquisition time per single‐metabolite image and (1.75 mm)3 isotropic voxels with full mouse abdomen 56 × 28 × 21 mm3 FOV and fully‐sampled k‐space. Kidney and tumor lactate/pyruvate ratios of two consecutive measurements in one animal, 1 h apart, were consistent. Conclusion: Spectrally selective bSSFP using off‐resonant RF excitations can provide high spatio‐temporal resolution 3D dynamic images of pyruvate‐lactate metabolic conversion. [ABSTRACT FROM AUTHOR]

Purpose: Hyperpolarized [1‐13C]pyruvate MRI is an emerging clinical tool for metabolic imaging. It has the potential for absolute quantitative metabolic imaging. However, the method itself is not quantitative, limiting comparison of images across both time and between individuals. Here, we propose a simple signal normalization to the whole‐body oxidative metabolism to overcome this limitation. Theory and Methods: A simple extension of the model‐free ratiometric analysis of hyperpolarized [1‐13C]pyruvate MRI is presented, using the expired 13CO2 in breath for normalization. The proposed framework was investigated in two porcine cohorts (N = 11) subjected to local renal hypoperfusion defects and subsequent [1‐13C]pyruvate MRI. A breath sample was taken before the [1‐13C]pyruvate injection and 5 min after. The raw MR signal from both the healthy and intervened kidney in the two cohorts was normalized using the 13CO2 in the expired air. Results: 13CO2 content in the expired air was significantly different between the two cohorts. Normalization to this reduced the coefficients of variance in the aerobic metabolic sensitive pathways by 25% for the alanine/pyruvate ratio, and numerical changes were observed in the bicarbonate/pyruvate ratio. The lactate/pyruvate ratio was largely unaltered (<2%). Conclusion: Our results indicate that normalizing the hyperpolarized 13C‐signal ratios by the 13CO2 content in expired air can reduce variation as well as improve specificity of the method by normalizing the metabolic readout to the overall metabolic status of the individual. The method is a simple and cheap extension to the hyperpolarized 13C exam. [ABSTRACT FROM AUTHOR]

Purpose: Model‐constrained reconstruction with Fourier‐based undersampling (MoReFUn) is introduced to accelerate the acquisition of dynamic MRI using hyperpolarized [1‐13C]‐pyruvate. Methods: The MoReFUn method resolves spatial aliasing using constraints introduced by a pharmacokinetic model that describes the signal evolution of both pyruvate and lactate. Acceleration was evaluated on three single‐channel data sets: a numerical digital phantom that is used to validate the accuracy of reconstruction and model parameter restoration under various SNR and undersampling ratios, prospectively and retrospectively sampled data of an in vitro dynamic multispectral phantom, and retrospectively undersampled imaging data from a prostate cancer patient to test the fidelity of reconstructed metabolite time series. Results: All three data sets showed successful reconstruction using MoReFUn. In simulation and retrospective phantom data, the restored time series of pyruvate and lactate maintained the image details, and the mean square residual error of the accelerated reconstruction increased only slightly (< 10%) at a reduction factor up to 8. In prostate data, the quantitative estimation of the conversion‐rate constant of pyruvate to lactate was achieved with high accuracy of less than 10% error at a reduction factor of 2 compared with the conversion rate derived from unaccelerated data. Conclusion: The MoReFUn technique can be used as an effective and reliable imaging acceleration method for metabolic imaging using hyperpolarized [1‐13C]‐pyruvate. [ABSTRACT FROM AUTHOR]

BACKGROUND: The demand for value‐added compounds such as amylase, pyruvate and phenolic compounds produced by biological methods has prompted the rapid development of advanced technologies for their enhanced production. Nanobiohybrids (NBs) make use of both the microbial properties of whole‐cell microorganisms and the light‐harvesting efficiency of semiconductors. Photosynthetic NBs were constructed that link the biosynthetic pathways of Aspergillus niger with CuS nanoparticles. RESULTS: In this work, NB formation was confirmed by negative values of the interaction energy, i.e., 2.31 × 108 to −5.52 × 108 kJ mol−1 for CuS–Che NBs, whereas for CuS–Bio NBs the values were −2.31 × 108 to −4.62 × 108 kJ mol−1 for CuS–Bio NBs with spherical nanoparticle interaction. For CuS–Bio NBs with nanorod interaction, it ranged from −2.3 × 107 to −3.47 × 107 kJ mol−1. Further, the morphological changes observed by scanning electron microscopy showed the presence of the elements Cu and S in the energy‐dispersive X‐ray spectra and the presence of CuS bonds in Fourier transform infrared spectroscopy indicate NB formation. In addition, the quenching effect in photoluminescence studies confirmed NB formation. Production yields of amylase, phenolic compounds and pyruvate amounted to 11.2 μmol L−1, 52.5 μmol L−1 and 28 nmol μL−1, respectively, in A. niger–CuS Bio NBs on the third day of incubation in the bioreactor. Moreover, A niger cells–CuS Bio NBs had amino acids and lipid yields of 6.2 mg mL−1 and 26.5 mg L−1, respectively. Furthermore, probable mechanisms for the enhanced production of amylase, pyruvate and phenolic compounds are proposed. CONCLUSION: Aspergillus niger–CuS NBs were used for the production of the amylase enzyme and value‐added compounds such as pyruvate and phenolic compounds. Aspergillus niger–CuS Bio NBs showed a greater efficiency compared to A. niger–CuS Che NBs as the biologically produced CuS nanoparticles had a higher compatibility with A. niger cells. © 2022 The Authors. Journal of Chemical Technology and Biotechnology published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry (SCI). [ABSTRACT FROM AUTHOR]

Parahydrogen‐induced polarization is a hyperpolarization method that exploits the spin order of hydrogen enriched in the para‐isomer, by means of a chemical reaction. Recently, its field of application has been extended significantly, through the introduction of non‐hydrogenative PHIP (i. e. SABRE) and of innovative h‐PHIP strategies that allowed to increase the intensity of the MR signals in molecules relevant for biological applications. This Concept article aims to show the potentialities of this hyperpolarization method in the field of diagnostics, through the discussion of some of the reported applications of parahydrogen polarized substrates. A section is also dedicated to the methods that have been introduced for the purification of parahydrogen polarized products, in order to make them suitable for biological studies. [ABSTRACT FROM AUTHOR]

Cachexia is a systemic disease associated with several pathologies, including cancer, that leads to excessive weight loss due to enhanced protein degradation. Previously, we showed that cachectic features in myotubes are provoked by a metabolic shift toward lactic fermentation. Our previous results led us to hyphotesise that increasing pyruvate concentration could impede the metabolic modifications responsible for induction of cachexia in myotubes. Here, we demonstrated that the addition of sodium pyruvate in conditioned media from CT26 colon cancer cells (CM CT26) prevents the onset of either phenotypic and metabolic cachectic features. Myotubes treated with CM CT26 containing sodium pyruvate show a phenotype similar to the healthy counterpart and display lactate production, oxygen consumption, and pyruvate dehydrogenase activity as control myotubes. The use of the Mitochondrial Pyruvate Carrier inhibitor UK5099, highlights the importance of mitochondrial pyruvate amount in the prevention of cachexia. Indeed, UK5099‐treated myotubes show cachectic features as those observed in myotubes treated with CM CT26. Finally, we found that sodium pyruvate is able to decrease STAT3 phosphorylation level, a signaling pathway involved in the induction of cachexia in myotubes. Collectively, our results show that cachexia in myotubes could be prevented by the utilization of sodium pyruvate which impedes the metabolic modifications responsible for the acquisition of the cachectic features. [ABSTRACT FROM AUTHOR]

Purpose: To investigate multi‐resolution hyperpolarized (HP) 13C pyruvate MRI for measuring kinetic conversion rates in the human brain. Methods: HP [1‐13C]pyruvate MRI was acquired in 6 subjects with a multi‐resolution EPI sequence at 7.5 × 7.5 mm2 resolution for pyruvate and 15 × 15 mm2 resolution for lactate and bicarbonate. With the same lactate data, 2 quantitative maps of pyruvate‐to‐lactate conversion (kPL) maps were generated: 1 using 7.5 × 7.5 mm2 resolution pyruvate data and the other using synthetic 15 × 15 mm2 resolution pyruvate data to simulate a standard constant resolution acquisition. To examine local kPL values, 4 voxels were manually selected in each study representing brain tissue near arteries, brain tissue near veins, white matter, and gray matter. Results: High resolution 7.5 × 7.5 mm2 pyruvate images increased the spatial delineation of brain structures and decreased partial volume effects compared to coarser resolution 15 × 15 mm2 pyruvate images. Voxels near arteries, veins and in white matter exhibited higher calculated kPL for multi‐resolution images. Conclusion: Acquiring HP 13C pyruvate metabolic data with a multi‐resolution approach minimized partial volume effects from vascular pyruvate signals while maintaining the SNR of downstream metabolites. Higher resolution pyruvate images for kinetic fitting resulted in increased kinetic rate values, particularly around the superior sagittal sinus and cerebral arteries, by reducing extracellular pyruvate signal contributions from adjacent blood vessels. This HP 13C study showed that acquiring pyruvate with finer resolution improved the quantification of kinetic rates throughout the human brain. [ABSTRACT FROM AUTHOR]

Purpose: To investigate the potential effects of [1‐13C]lactate RF saturation pulses on [13C]bicarbonate detection in hyperpolarized [1‐13C]pyruvate MRI of the brain. Methods: Thirteen healthy rats underwent MRI with hyperpolarized [1‐13C]pyruvate of either the brain (n = 8) or the kidneys, heart, and liver (n = 5). Dynamic, metabolite‐selective imaging was used in a cross‐over experiment in which [1‐13C]lactate was excited with either 0° or 90° flip angles. The [13C]bicarbonate SNR and apparent [1‐13C]pyruvate‐to‐[13C]bicarbonate conversion (kPB) were determined. Furthermore, simulations were performed to identify the SNR optimal flip‐angle scheme for detection of [1‐13C]lactate and [13C]bicarbonate. Results: In the brain, the [13C]bicarbonate SNR was 64% higher when [1‐13C]lactate was not excited (5.8 ± 1.5 vs 3.6 ± 1.3; 1.2 to 3.3–point increase; p = 0.0027). The apparent kPB decreased 25% with [1‐13C]lactate saturation (0.0047 ± 0.0008 s−1 vs 0.0034 ± 0.0006 s−1; 95% confidence interval, 0.0006–0.0019 s−1 increase; p = 0.0049). These effects were not present in the kidneys, heart, or liver. Simulations suggest that the optimal [13C]bicarbonate SNR with a TR of 1 s in the brain is obtained with [13C]bicarbonate, [1‐13C]lactate, and [1‐13C]pyruvate flip angles of 60°, 15°, and 10°, respectively. Conclusions: Radiofrequency saturation pulses on [1‐13C]lactate limit [13C]bicarbonate detection in the brain specifically, which could be due to shuttling of lactate from astrocytes to neurons. Our results have important implications for experimental design in studies in which [13C]bicarbonate detection is warranted. [ABSTRACT FROM AUTHOR]

Sepsis is a heterogeneous syndrome resulting from a dysregulated host response to infection. It is considered as a global major health priority. Sepsis is characterized by significant metabolic perturbations, leading to increased circulating metabolites such as lactate. In mammals, pyruvate is the primary substrate for lactate production. It plays a critical role in metabolism by linking glycolysis, where it is produced, with the mitochondrial oxidative phosphorylation pathway, where it is oxidized. Here, we provide an overview of all cytosolic and mitochondrial enzymes involved in pyruvate metabolism and how their activities are disrupted in sepsis. Based on the available data, we also discuss potential therapeutic strategies targeting these pyruvate-related enzymes leading to enhanced survival., (© 2024. The Author(s).)

Enantioselective catalytic formation of polyfunctional molecules, such as non‐natural amino acids, is important for the efficient production of many chiral compounds. To this end, we present here the synthesis and evaluation of the catalytic activity of bifunctional peptide–thiourea organocatalysts. These hybrid organocatalysts consist of Pro‐Pro dipeptide and thiourea moiety connected via a 1,2‐diaminocyclohexane unit. These catalysts promoted challenging Mannich reaction between α‐imino esters and pyruvates, providing orthogonally protected oxo‐glutamate derivatives. The N‐tosyl‐protected imino ester, as the most active imine, was required to compensate for the poor reactivity of pyruvates. DFT calculations, NMR, and CD spectroscopy help elucidate the mode of action of these catalysts. Configuration of the dipeptide Pro‐Pro moiety is responsible for the sense of the stereoinduction of the Mannich reaction. [ABSTRACT FROM AUTHOR]

Hyperpolarization methods in magnetic resonance enhance the signals by several orders of magnitude, opening new windows for real‐time investigations of dynamic processes in vitro and in vivo. Here, we propose a field‐independent para‐hydrogen‐based pulsed method to produce rapidly hyperpolarized 13C‐labeled substrates. We demonstrate the method by polarizing the carboxylic carbon of the pyruvate moiety in a purposely designed precursor to 24 % at ≈22 mT. Following a fast purification procedure, we measure 8 % polarization on free [1‐13C]pyruvate in clean water solutions at physiological conditions at 7 T. The enhanced signals allow real‐time monitoring of the pyruvate‐lactate conversion in cancer cells, demonstrating the potential of the method for biomedical applications in combination with existing or developing magnetic resonance technologies. [ABSTRACT FROM AUTHOR]

A novel electrochemical immunosensor for lactate dehydrogenase (LDH) detection was proposed based on analyte‐driven catalytic reaction by attaching LDH antibodies on multi‐walled carbon nanotubes (MWCNTs) and gold nanoparticles (AuNPs) modified glassy carbon electrodes (GCE). As LDH was captured by the antibodies on electrode surface, it catalyzed the formation of pyruvate and the reduced form of nicotinamide adenine dinucleotide (NADH), thus a sensitive electrochemical signal obtained from the above redox reaction was recorded by differential pulse voltammetry (DPV). Under optimum conditions, the developed immunosensor exhibits high sensitivity for LDH quantification ranging from 0.001 μg/mL to 0.5 μg/mL with a low detection limit at 0.39 ng/mL. This developed immunosensor reveals ideal accuracy and feasibility for LDH detection in Streptococcus uberis (S. uberis) induced bovine mammary epithelial cells (MECs) samples by comparison with conventional commercial kit, which shows remarkably application potential in diseases diagnosis. [ABSTRACT FROM AUTHOR]

High volumetric productivities can be achieved when perfusion processes are operated at high cell densities. Yet it is fairly challenging to keep high cell density cultures in a steady state over an extended period. Aiming for robust processes, cultures were operated at a constant biomass specific perfusion rate (BSPR) in this study. The cell density was monitored with a capacitance probe and a continuous bleed maintained the targeted viable cell volume. Despite our tightly controlled BSPR, a gradual accumulation of ammonium and changes in cell diameter were observed during the production phase for three different monoclonal antibodies. Although a lot of efforts in media optimization have been made to reduce ammonium in fed‐batch process, less examples are known about how media components impact the cellular metabolism and thus the quality of monoclonal antibodies in continuous processes. In this study, we show that a continuous Na‐pyruvate feed (2 g/L/day) strongly reduced ammonium production and stabilized fucosylation, sialylation and high mannose content for three different mAbs. [ABSTRACT FROM AUTHOR]

DNA repair confers the resistance of tumor cells to DNA‐damaging anticancer therapies, while how reprogrammed metabolism in tumor cells contributes to such process remains poorly understood. Pyruvate kinase M2 isoform (PKM2) catalyzes the conversion of phosphoenolpyruvate to pyruvate and regulates the last rate‐limiting step of glycolysis. Here it is shown that the glycolytic metabolite pyruvate enhances DNA damage repair by facilitating chromatin loading of γH2AX, thereby promoting the radiation resistance of glioma cells. Mechanistically, PKM2 is phosphorylated at serine (S) 222 upon DNA damage and interacts with FACT complex, a histone chaperone comprising SPT16 and SSRP1 subunit. The pyruvate produced by PKM2 directly binds to SSRP1, which increases the association of FACT complex with γH2AX and subsequently facilitates FACT‐mediated chromatin loading of γH2AX, ultimately promoting DNA repair and tumor cell survival. Intriguingly, the supplementation of exogenous pyruvate can also sufficiently enhance FACT‐mediated chromatin loading of γH2AX and promotes tumor cell survival upon DNA damage. The levels of PKM2 S222 phosphorylation correlate with the malignancy and prognosis of human glioblastoma. The finding demonstrates a novel mechanism by which PKM2‐produced pyruvate promotes DNA repair by regulating γH2AX loading to chromatin and establishes a critical role of this mechanism in glioblastoma radiation resistance. [ABSTRACT FROM AUTHOR]

Parkinson's disease (PD) is a progressive neurodegenerative disorder induced by the loss of dopaminergic neurons in midbrain. The mechanism of neurodegeneration is associated with aggregation of misfolded proteins, oxidative stress, and mitochondrial dysfunction. Considering this, the process of removal of unwanted organelles or proteins by autophagy is vitally important in neurons, and activation of these processes could be protective in PD. Short‐time acidification of the cytosol can activate mitophagy and autophagy. Here, we used sodium pyruvate and sodium lactate to induce changes in intracellular pH in human fibroblasts with PD mutations (Pink1, Pink1/Park2, α‐synuclein triplication, A53T). We have found that both lactate and pyruvate in millimolar concentrations can induce a short‐time acidification of the cytosol in these cells. This induced activation of mitophagy and autophagy in control and PD fibroblasts and protected against cell death. Importantly, application of lactate to acute brain slices of WT and Pink1 KO mice also induced a reduction of pH in neurons and astrocytes that increased the level of mitophagy. Thus, acidification of the cytosol by compounds, which play an important role in cell metabolism, can also activate mitophagy and autophagy and protect cells in the familial form of PD. [ABSTRACT FROM AUTHOR]

CRISPR interference (CRISPRi) was applied to enable the aerobic production of pyruvate in Escherichia coli MG1655 under glucose excess conditions by targeting the promoter regions of aceE or pdhR. Knockdown strains were cultivated in aerobic shaking flasks and the influence of inducer concentration and different sgRNA binding sites on the production of pyruvate was measured. Targeting the promoter regions of aceE or pdhR triggered pyruvate production during the exponential phase and reduced expression of aceE. In lab‐scale bioreactor fermentations, an aceE silenced strain successfully produced pyruvate under fully aerobic conditions during the exponential phase, but loss of productivity occurred during a subsequent nitrogen‐limited phase. Targeting the promoter region of pdhR enabled pyruvate production during the growth phase of cultivations, and a continued low‐level accumulation during the nitrogen‐limited production phase. Combinatorial targeting of the promoter regions of both aceE and pdhR in E. coli MG1655 pdCas9 psgRNA_aceE_234_pdhR_329 resulted in the stable aerobic production of pyruvate with non‐growing cells at YP/S = 0.36 ± 0.029 gPyruvate/gGlucose in lab‐scale bioreactors throughout an extended nitrogen‐limited production phase. [ABSTRACT FROM AUTHOR]

Purpose: We focused on the metabolism of oocytes in pre‐culture and insemination and compared these results between our existing fertilization medium, GEMS Fertilisation Medium (GEMS group) (Merck BioPharma) and Continuous Single Culture Medium—NX Complete (CSCM‐NXC group) (FUJIFILM Irvine Scientific). Methods: Patients under 42 years of age were received controlled ovarian stimulation and oocytes were retrieved. Those were pre‐cultured and fertilized with either GEMS fertilization medium or CSCM‐NXC. After fertilization was confirmed, embryos were cultured using CSCM‐NXC in both groups. The embryos were cryopreserved at blastocyst stage (3BB or more, Gardner classification) and then transferred in HRT cycles. Results: The fertilization rate of both groups was the same, but the 3PN rate was significantly lower in the CSCM‐NXC group. In terms of embryo culture results, the CSCM‐NXC group had a significantly higher rate of good quality blastocysts, high‐grade embryos, and embryos with a high degree of expansion. Conclusions: The use of CSCM‐NXC, a low lactate embryo culture medium, from pre‐culture and for insemination, increases the energy metabolic efficiency of oocytes and cumulus cells, making it possible to supply sufficient energy ATP for fertilization and early division, which is thought to promote good embryonic development.The use of CSCM‐NXC, a low lactate embryo culture medium, from pre‐culture and for insemination promotes good embryonic development. [ABSTRACT FROM AUTHOR]

Purpose: The combined hyperpolarized (HP) 13C pyruvate and urea MRI has provided a simultaneous assessment of glycolytic metabolism and tissue perfusion for improved cancer diagnosis and therapeutic evaluation in preclinical studies. This work aims to translate this dual‐probe HP imaging technique to clinical research. Methods: A co‐polarization system was developed where [1‐13C]pyruvic acid (PA) and [13C, 15N2]urea in water solution were homogeneously mixed and polarized on a 5T SPINlab system. Physical and chemical characterizations and toxicology studies of the combined probe were performed. Simultaneous metabolic and perfusion imaging was performed on a 3T clinical MR scanner by alternatively applying a multi‐slice 2D spiral sequence for [1‐13C]pyruvate and its downstream metabolites and a 3D balanced steady‐state free precession (bSSFP) sequence for [13C, 15N2]urea. Results: The combined PA/urea probe has a glass‐formation ability similar to neat PA and can generate nearly 40% liquid‐state 13C polarization for both pyruvate and urea in 3‐4 h. A standard operating procedure for routine on‐site production was developed and validated to produce 40 mL injection product of approximately 150 mM pyruvate and 35 mM urea. The toxicology study demonstrated the safety profile of the combined probe. Dynamic metabolite‐specific imaging of [1‐13C]pyruvate, [1‐13C]lactate, [1‐13C]alanine, and [13C, 15N2]urea was achieved with adequate spatial (2.6 mm × 2.6 mm) and temporal resolution (4.2 s), and urea images showed reduced off‐resonance artifacts due to the JCN coupling. Conclusion: The reported technical development and translational studies will lead to the first‐in‐human dual‐agent HP MRI study and mark the clinical translation of the first HP 13C MRI probe after pyruvate. [ABSTRACT FROM AUTHOR]

Background: Alterations in glycolysis are central to the increasing incidence of non‐alcoholic fatty liver disease (NAFLD), highlighting a need for in vivo, non‐invasive technologies to understand the development of hepatic metabolic aberrations. Purpose: To use hyperpolarized magnetic resonance spectroscopy (MRS) and proton density fat fraction (PDFF) magnetic resonance imaging (MRI) techniques to investigate the effects of a chronic, life‐long exposure to the Western diet (WD) in an animal model resulting in NAFLD; to investigate the hypothesis that exposure to the WD will result in NAFLD in association with altered pyruvate metabolism. Study Type: Prospective. Animal Model: Twenty‐eight male guinea pigs weaned onto a control diet (N = 14) or WD (N = 14). Field Strength/Sequence: 3 T; T1‐weighted gradient echo, T2‐weighted spin‐echo, three‐dimensional gradient multi‐echo fat‐water separation (IDEAL‐IQ), and broadband point‐resolved spectroscopy (PRESS) chemical‐shift sequences. Assessment: Median PDFF was calculated in the liver and hind limbs. [1‐13C]pyruvate dynamic MRS in the liver was quantified by the time‐to‐peak (TTP) for each metabolite. Animals were euthanized and tissue was analyzed for lipid and cholesterol concentration and enzyme level and activity. Statistical Tests: Unpaired Student's t‐tests were used to determine differences in measurements between the two diet groups. The Pearson correlation coefficient was calculated to determine correlations between measurements. Results: Life‐long WD consumption resulted in significantly higher liver PDFF and elevated triglyceride content in the liver. The WD group exhibited a decreased TTP for lactate production, and ex vivo analysis highlighted increased liver lactate dehydrogenase (LDH) activity. Data Conclusion: PDFF MRI results suggest differential fat deposition patterns occurring in animals fed a life‐long WD characteristic of lean, or lacking excessive subcutaneous fat, NAFLD. The decreased liver lactate TTP and increased ex vivo LDH activity suggest lipid accumulation occurs in association with a shift from oxidative metabolism to anaerobic glycolytic metabolism in WD‐exposed livers. Level of Evidence: 2 Technical Efficacy Stage: 1 [ABSTRACT FROM AUTHOR]

BACKGROUND: Optimal treatment selection for localized renal tumors is challenging because of their variable biologic behavior and limitations in the preoperative assessment of tumor aggressiveness. The authors investigated the emerging hyperpolarized (HP) 13C magnetic resonance imaging (MRI) technique to noninvasively assess tumor lactate production, which is strongly associated with tumor aggressiveness. METHODS: Eleven patients with renal tumors underwent HP 13C pyruvate MRI before surgical resection. Tumor 13C pyruvate and 13C lactate images were acquired dynamically. Five patients underwent 2 scans on the same day to assess the intrapatient reproducibility of HP 13C pyruvate MRI. Tumor metabolic data were compared with histopathology findings. RESULTS: Eight patients had tumors with a sufficient metabolite signal‐to‐noise ratio for analysis; an insufficient tumor signal‐to‐noise ratio was noted in 2 patients, likely caused by poor tumor perfusion and, in 1 patient, because of technical errors. Of the 8 patients, 3 had high‐grade clear cell renal cell carcinoma (ccRCC), 3 had low‐grade ccRCC, and 2 had chromophobe RCC. There was a trend toward a higher lactate‐to‐pyruvate ratio in high‐grade ccRCCs compared with low‐grade ccRCCs. Both chromophobe RCCs had relatively high lactate‐to‐pyruvate ratios. Good reproducibility was noted across the 5 patients who underwent 2 HP 13C pyruvate MRI scans on the same day. CONCLUSIONS: The current results demonstrate the feasibility of HP 13C pyruvate MRI for investigating the metabolic phenotype of localized renal tumors. The initial data indicate good reproducibility of metabolite measurements. In addition, the metabolic data indicate a trend toward differentiating low‐grade and high‐grade ccRCCs, the most common subtype of renal cancer. LAY SUMMARY: Renal tumors are frequently discovered incidentally because of the increased use of medical imaging, but it is challenging to identify which aggressive tumors should be treated.A new metabolic imaging technique was applied to noninvasively predict renal tumor aggressiveness.The imaging results were compared with tumor samples taken during surgery and showed a trend toward differentiating between low‐grade and high‐grade clear cell renal cell carcinomas, which are the most common type of renal cancers. A new metabolic imaging technique, hyperpolarized 13C pyruvate magnetic resonance imaging, is applied to predict renal tumor aggressiveness. The results demonstrate feasibility, reproducibility, and promising trends for risk stratifying renal tumors noninvasively. [ABSTRACT FROM AUTHOR]

Purpose: To compare carbon‐13 (13C) MRSI of hyperpolarized [1‐13C]pyruvate metabolism in a murine tumor model with mass spectrometric (MS) imaging of the corresponding tumor sections in order to cross validate these metabolic imaging techniques and to investigate the effects of pyruvate delivery and tumor lactate concentration on lactate labeling. Methods: [1‐13C]lactate images were obtained from tumor‐bearing mice, following injection of hyperpolarized [1‐13C]pyruvate, using a single‐shot 3D 13C spectroscopic imaging sequence in vivo and using desorption electrospray ionization MS imaging of the corresponding rapidly frozen tumor sections ex vivo. The images were coregistered, and levels of association were determined by means of Spearman rank correlation and Cohen kappa coefficients as well as linear mixed models. The correlation between [1‐13C]pyruvate and [1‐13C]lactate in the MRS images and between [12C] and [1‐13C]lactate in the MS images were determined by means of Pearson correlation coefficients. Results: [1‐13C]lactate images generated by MS imaging were significantly correlated with the corresponding MRS images. The correlation coefficient between [1‐13C]lactate and [1‐13C]pyruvate in the MRS images was higher than between [1‐13C]lactate and [12C]lactate in the MS images. Conclusion: The inhomogeneous distribution of labeled lactate observed in the MRS images was confirmed by MS imaging of the corresponding tumor sections. The images acquired using both techniques show that the rate of 13C label exchange between the injected pyruvate and endogenous tumor lactate pool is more correlated with the rate of pyruvate delivery to the tumor cells and is less affected by the endogenous lactate concentration. [ABSTRACT FROM AUTHOR]

Hyperpolarized [1‐13C] pyruvate can be used to examine the metabolic state of cancer cells, highlighting a key metabolic characteristic of cancer: the upregulated metabolic flux to lactate, even in the presence of oxygen (Warburg effect). Thus, the rate constant of 13C exchange of pyruvate to lactate, kPL, can serve as a metabolic biomarker of cancer presence, aggressiveness and therapy response. Established in vitro hyperpolarized experiments dissolve the probe for each cell sample independently, an inefficient process that consumes excessive time and resources. Expanding on our previous development of a microcoil with greatly increased detection sensitivity (103‐fold) compared with traditional in vitro methods, we present a novel microcoil equipped with a 10‐μL vertical reservoir and an experimental protocol utilizing deuterated dissolution buffer to measure metabolic flux in multiple mass‐limited cell suspension samples using a single dissolution. This method increases efficiency and potentially reduces the methodological variability associated with hyperpolarized experiments. This technique was used to measure pyruvate‐to‐lactate flux in melanoma cells to assess BRAF‐inhibition treatment response. There was a significant reduction of kPL in BRAFV600E cells following 24 and 48 hours of treatment with 2 μM vemurafenib (P ≤.05). This agrees with significant changes observed in the pool sizes of extracellular lactate (P ≤.05) and glucose (P ≤.001) following 6 and 48 hours of treatment, respectively, and a significant reduction in cell proliferation following 72 hours of treatment (P ≤.01). BRAF inhibition had no significant effect on the metabolic flux of BRAFWT cells. These data demonstrate a 6‐8–fold increase in efficiency for the measurement of kPL in cell suspension samples compared with traditional hyperpolarized in vitro methods. [ABSTRACT FROM AUTHOR]

Purpose: To ameliorate tradeoffs between a fixed spatial resolution and signal‐to‐noise ratio (SNR) for hyperpolarized 13C MRI. Methods: In MRI, SNR is proportional to voxel volume but retrospective downsampling or voxel averaging only improves SNR by the square root of voxel size. This can be exploited with a metabolite‐selective imaging approach that independently encodes each compound, yielding high‐resolution images for the injected substrate and coarser resolution images for downstream metabolites, while maintaining adequate SNR for each. To assess the efficacy of this approach, hyperpolarized [1‐13C]pyruvate data were acquired in healthy Sprague‐Dawley rats (n = 4) and in two healthy human subjects. Results: Compared with a constant resolution acquisition, variable‐resolution data sets showed improved detectability of metabolites in pre‐clinical renal studies with a 3.5‐fold, 8.7‐fold, and 6.0‐fold increase in SNR for lactate, alanine, and bicarbonate data, respectively. Variable‐resolution data sets from healthy human subjects showed cardiac structure and neuro‐vasculature in the higher resolution pyruvate images (6.0 × 6.0 mm2 for cardiac and 7.5 × 7.5 mm2 for brain) that would otherwise be missed due to partial‐volume effects and illustrates the level of detail that can be achieved with hyperpolarized substrates in a clinical setting. Conclusion: We developed a variable‐resolution strategy for hyperpolarized 13C MRI using metabolite‐selective imaging and demonstrated that it mitigates tradeoffs between a fixed spatial resolution and SNR for hyperpolarized substrates, providing both high resolution pyruvate and coarse resolution metabolite data sets in a single exam. This technique shows promise to improve future studies by maximizing metabolite SNR while minimizing partial‐volume effects from the injected substrate. [ABSTRACT FROM AUTHOR]

Streptococcus pneumoniae is a major human pathogen that must adapt to unique nutritional environments in several host niches. The pneumococcus can metabolize a range of carbohydrates that feed into glycolysis ending in pyruvate, which is catabolized by several enzymes. We investigated how the pneumococcus utilizes these enzymes to metabolize different carbohydrates and how this impacts survival in the host. Loss of ldh decreased bacterial burden in the nasopharynx and enhanced bacteremia in mice. Loss of spxB, pdhC or pfl2 decreased bacteremia and increased host survival. In glucose or galactose, loss of ldh increased capsule production, whereas loss of spxB and pdhC reduced capsule production. The pfl2 mutant exhibited reduced capsule production only in galactose. In glucose, pyruvate was metabolized primarily by LDH to generate lactate and NAD+ and by SpxB and PDHc to generate acetyl‐CoA. In galactose, pyruvate metabolism was shunted toward acetyl‐CoA production. The majority of acetyl‐CoA generated by PFL was used to regenerate NAD+ with a subset used in capsule production, while the acetyl‐CoA generated by SpxB and PDHc was utilized primarily for capsule biosynthesis. These data suggest that the pneumococcus can alter flux of pyruvate metabolism dependent on the carbohydrate present to succeed in distinct host niches. [ABSTRACT FROM AUTHOR]

Purpose: Imaging tumor metabolism in vivo using hyperpolarized [1‐13C]pyruvate is a promising technique for detecting disease, monitoring disease progression, and assessing treatment response. However, the transient nature of the hyperpolarization and its depletion following excitation limits the available time for imaging. We describe here a single‐shot multi spin echo sequence, which improves on previously reported sequences, with a shorter readout time, isotropic point spread function (PSF), and better signal‐to‐noise ratio. Methods: The sequence uses numerically optimized spectrally selective excitation pulses set to the resonant frequencies of pyruvate and lactate and a hyperbolic secant adiabatic refocusing pulse, all applied in the absence of slice selection gradients. The excitation pulses were designed to be resistant to the effects of B0 and B1 field inhomogeneity. The gradient readout uses a 3D cone trajectory composed of 13 cones, all fully refocused and distributed among 7 spin echoes. The maximal gradient amplitude and slew rate were set to 4 G/cm and 20 G/cm/ms, respectively, to demonstrate the feasibility of clinical translation. Results: The pulse sequence gave an isotropic PSF of 2.8 mm. The excitation profiles of the optimized pulses closely matched simulations and a 46.10 ± 0.04% gain in image SNR was observed compared to a conventional Shinnar–Le Roux excitation pulse. The sequence was demonstrated with dynamic imaging of hyperpolarized [1‐13C]pyruvate and [1‐13C]lactate in vivo. Conclusion: The pulse sequence was capable of dynamic imaging of hyperpolarized 13C labeled metabolites in vivo with relatively high spatial and temporal resolution and immunity to system imperfections. [ABSTRACT FROM AUTHOR]

Purpose: The balanced steady‐state free precession sequence has been previously explored to improve the efficient use of nonrecoverable hyperpolarized 13C magnetization, but suffers from poor spectral selectivity and long acquisition time. The purpose of this study was to develop a novel metabolite‐specific 3D bSSFP ("MS‐3DSSFP") sequence with stack‐of‐spiral readouts for improved lactate imaging in hyperpolarized [1-13C]pyruvate studies on a clinical 3T scanner. Methods: Simulations were performed to evaluate the spectral response of the MS‐3DSSFP sequence. Thermal 13C phantom experiments were performed to validate the MS‐3DSSFP sequence. In vivo hyperpolarized [1-13C], pyruvate studies were performed to compare the MS‐3DSSFP sequence with metabolite‐specific gradient echo ("MS‐GRE") sequences for lactate imaging. Results: Simulations, phantom, and in vivo studies demonstrate that the MS‐3DSSFP sequence achieved spectrally selective excitation on lactate while minimally perturbing other metabolites. Compared with MS‐GRE sequences, the MS‐3DSSFP sequence showed approximately a 2.5‐fold SNR improvement for lactate imaging in rat kidneys, prostate tumors in a mouse model, and human kidneys. Conclusions: Improved lactate imaging using the MS‐3DSSFP sequence in hyperpolarized [1-13C]pyruvate studies was demonstrated in animals and humans. The MS‐3DSSFP sequence could be applied for other clinical applications such as in the brain or adapted for imaging other metabolites such as pyruvate and bicarbonate. [ABSTRACT FROM AUTHOR]

Purpose: To improve the SNR of hyperpolarized carbon‐13 MRI of [1‐13C]pyruvate using a multispectral variable flip angle (msVFA) scheme in which the spectral profile and flip angle vary dynamically with time. Methods: Each image acquisition in a time‐resolved imaging experiment used a unique spectrally varying RF pulse shape for msVFA. Therefore, the flip angle for every acquisition was optimized for pyruvate and each of its metabolites to yield the highest SNR across the acquisition. Multispectral VFA was compared with a spectrally varying constant flip‐angle excitation model through simulations and in vivo. A modified broadband chemical shift‐encoded gradient‐echo sequence was used for in vivo experiments on six pregnant guinea pigs. Regions of interest placed in the placentae, maternal liver, and maternal kidneys were used as areas for SNR measurement. Results: In vivo experiments showed significant increases in SNR for msVFA relative to constant flip angle of up to 250% for multiple metabolites. Conclusion: Hyperpolarized carbon‐13 imaging with msVFA excitation produces improved SNR for all metabolites in organs of interest. [ABSTRACT FROM AUTHOR]

Aim: Little information exists on the peripheral metabolite levels in individuals with at‐risk mental state who meet the criteria for a high‐risk state of psychosis. Here, we aimed to investigate serum levels of glucose, pyruvate and d‐ and l‐lactate, which may act as a signalling molecule for learning and memory in neuronal cells. Methods: High performance liquid chromatography or commercial kits were used to assess serum metabolites in individuals with attenuated psychosis symptoms of at‐risk mental state (n = 24, men = 12) who were not receiving antipsychotics. The metabolite levels of these individuals were compared with those of age‐ and sex‐matched healthy individuals (controls, n = 23, men = 11). Correlations between the metabolites and clinical symptoms of at‐risk mental state were also examined. Results: Individuals with at‐risk mental state had higher serum glucose levels than did controls (P = 2.18 × 10−3), while no significant difference in pyruvate levels were observed between the groups. Individuals with at‐risk mental state had significantly lower serum l‐lactate levels than did controls (P = 6.31 × 10−5), while no differences in d‐lactate levels were observed. Furthermore, a negative correlation was identified between serum l‐lactate levels and Positive and Negative Syndrome Scale negative symptoms scores (r = −0.5651, P = 4.01 × 10−3) in individuals with at‐risk mental state. Conclusions: We found that, compared with controls, individuals with at‐risk mental state have reduced serum l‐lactate levels, which may predate psychosis onset, and may be involved in the related negative symptoms. [ABSTRACT FROM AUTHOR]

The direct asymmetric aldol reaction of α‐ketoesters catalyzed by chiral tertiary amines is reported. The described methodology is characterized by mild reaction conditions and distinct product selectivity determined by the starting materials. In the developed transformation pyruvates undergo highly selective self‐condensation reaction, whereas cross‐aldol reaction take place predominantly for their carbon chain homologues in presence of aldehyde acceptors. Notably, addition of bulky phenol moiety into pyruvate inhibit the spontaneous lactonization of products and enhance the enantioselectivity. [ABSTRACT FROM AUTHOR]

Pyruvate is an important pharmaceutical intermediate and is widely used in food, nutraceuticals, and pharmaceuticals. However, high environmental pollution caused by chemical synthesis or complex separation process of microbial fermentation methods constrain the supply of pyruvate. Here, one‐step pyruvate and d‐alanine production from d,l‐alanine by whole‐cell biocatalysis was investigated. First, l‐amino acid deaminase (Pm1) from Proteus mirabilis was expressed in Escherichia coli, resulting in pyruvate titer of 12.01 g/L. Then, N‐terminal coding sequences were introduced to the 5′‐end of the pm1 gene to enhance the expression of Pm1 and the pyruvate titer increased to 15.13 g/L. Next, product utilization by the biocatalyst was prevented by knocking out the pyruvate uptake transporters (cstA, btsT) and the pyruvate metabolic pathway genes pps, poxB, pflB, ldhA, and aceEF using CRISPR/Cas9, yielding 30.88 g/L pyruvate titer. Finally, by optimizing the reaction conditions, the pyruvate titer was further enhanced to 43.50 g/L in 8 H with a 79.99% l‐alanine conversion rate; meanwhile, the resolution of d‐alanine reached 84.0%. This work developed a whole‐cell biocatalyst E. coli strain for high‐yield, high‐efficiency, and low‐pollution pyruvate and d‐alanine production, which has great potential for the commercial application in the future. [ABSTRACT FROM AUTHOR]

Background: Infants with hypoxic‐ischemic injury often require cardiopulmonary resuscitation. Mitochondrial failure to generate adenosine triphosphate (ATP) during hypoxic‐ischemic reperfusion injury contributes to cellular damage. Current postnatal strategies to improve outcome in hypoxic‐ischemic injury need sophisticated equipment to perform servo‐controlled cooling. Administration of intravenous pyruvate, an antioxidant with favorable effects on mitochondrial bioenergetics, is a simple intervention that can have a global impact. We hypothesize that the administration of pyruvate following the return of spontaneous circulation (ROSC) would improve cardiac function, systemic hemodynamics, and oxygen utilization in the brain in newborn lambs with cardiac arrest (CA). Methods: Term lambs were instrumented, delivered by C‐section and asphyxia induced by umbilical cord occlusion along with clamping of the endotracheal tube until asystole; Lambs resuscitated following 5 min of CA; upon ROSC, lambs were randomized to receive pyruvate or saline infusion over 90 min and ventilated for 150 min postinfusion. Pulmonary and systemic hemodynamics and arterial gases monitored. We measured plasma pyruvate, tissue lactate, and ATP levels (heart and brain) in both groups. Results: Time to ROSC was not different between the two groups. Systolic and diastolic blood pressures, stroke volume, arterial oxygen content, and cerebral oxygen delivery were similar between the two groups. The cerebral metabolic rate of oxygen was higher following pyruvate infusion; higher oxygen consumption in the brain was associated with lower plasma levels but higher brain ATP levels compared to the saline group. Conclusions: Pyruvate promotes energy generation accompanied by efficient oxygen utilization in the brain and may facilitate additional neuroprotection in the presence of hypoxic‐ischemic injury. [ABSTRACT FROM AUTHOR]