Pyruvate Ratio Research Articles

Acute renal metabolic effect of metformin assessed with hyperpolarised MRI in rats.

Metformin inhibits hepatic mitochondrial glycerol phosphate dehydrogenase, thereby increasing cytosolic lactate and suppressing gluconeogenesis flux in the liver. This inhibition alters cytosolic and mitochondrial reduction-oxidation (redox) potential, which has been reported to protect organ function in several disease states including diabetes. In this study, we investigated the acute metabolic and functional changes induced by metformin in the kidneys of both healthy and insulinopenic Wistar rats used as a model of diabetes. Diabetes was induced by intravenous injection of streptozotocin, and kidney metabolism in healthy and diabetic animals was investigated 4weeks thereafter using hyperpolarised 13C-MRI, Clark-type electrodes and biochemical analysis. Metformin increased renal blood flow, but did not change total kidney oxygen consumption. In healthy rat kidneys, metformin increased [1-13C]lactate production and reduced mitochondrial [1-13C]pyruvate oxidation (decreased the 13C-bicarbonate/[1-13C]pyruvate ratio) within 30min of administration. Corresponding alterations to indices of mitochondrial, cytosolic and whole-cell redox potential were observed. Pyruvate oxidation was maintained in the diabetic rats, suggesting that the diabetic state abrogates metabolic reprogramming caused by metformin. This study demonstrates that metformin-induced acute metabolic alterations in healthy kidneys favoured anaerobic metabolism at the expense of aerobic metabolism. The results suggest that metformin directly alters the renal redox state, with elevated renal cytosolic redox states as well as decreased mitochondrial redox state. These findings suggest redox biology as a novel target to eliminate the renal complications associated with metformin treatment in individuals with impaired renal function.

Detection of metabolic pattern following decompressive craniectomy in severe traumatic brain injury: A microdialysis study

ABSTRACTObjective: The aim of the study was to detect mitochondrial dysfunction and ischaemia in severe traumatic brain injury and their relationship with outcome.Methods: Forty-one patients with severe traumatic brain injury (TBI) who underwent decompressive craniectomy were prospectively monitored with intracerebral microdialysis catheters (MD). Variables related to energy metabolism were studied using microdialysis.Results: Twentysix patients (63.4%) had a good outcome in terms of Glasgow outcome score (GOS) at 6 months while the rest (15 patients) had poor GOS at 6 months. Mitochondrial dysfunction was defined as Lactate Pyruvate ratio (LP ratio) > 25 and pyruvate <70 while ischaemia was defined as LP ratio > 25 and pyruvate >70. The poor outcome group showed significantly higher proportion of mitochondrial dysfunction 65.9% vs. 55.9% (p<0.001) and ischemia 13.9% vs. 7.2% (p<0.001)Conclusions: After decompressive craniectomy in severe TBI, patients with higher incidence of mitochondrial dysfunction and ischaemia were more likely to have poorer outcome with ischaemia having a more profound effect.Abbreviations: Traumatic brain injury (TBI), microdialysis (MD), lactate pyruvate ratio (LP ratio), Glasgow coma scale (GCS), Glasgow outcome scale (GOS), cerebral perfusion pressure (CPP), intracranial pressure (ICP), mitochondrial transition pore (MTP), non-contrast computed tomography (NCCT), traumatic axonal injury (TAI).

Abstract 2852: Monitoring the impact on metabolic flux in vivo of a newly developed LDH inhibitor using hyperpolarized 13C magnetic resonance spectroscopic imaging

Abstract [aim] Increased lactate production is a feature of many neoplasms, and Lactate Dehydrogenase A (LDH-A) plays a key role in conversion of pyruvate to lactate. LDHA inhibition, therefore, is considered to be a promising approach toward developing a new therapeutic strategy for cancer treatment focused on targeting cancer metabolism. Non-invasive imaging approaches able to monitor metabolic fluxes in vivo will be useful for this purpose. Hyperpolarized 13C Magnetic Resonance Imaging (MRI) has been well known as a valuable technology to investigate metabolic processes in tumor xenografts, allowing us to perform dynamic 13C-metabolic flux analysis in vivo. Use of [1-13C]pyruvate with this technology provides the ability to monitor LDHA activity in real time through dynamic observation of conversion of [1-13C]pyruvate to [1-13C]lactate. This study aimed to monitor drug efficacy of a newly developed LDH inhibitor (LDHI, obtained from National Cancer Institute Experimental Therapeutics Program, NExT) in a xenograft tumor model using 13C MRI technology with hyperpolarized 13C-labeled pyruvate. [Results] Hyperpolarized [1-13C]pyruvate MR studies were performed before and after LDHI administration to assess the impact on metabolic flux in vivo. Using hyperpolarized [1-13C]pyruvate MR Spectroscopy (MRS), we found that lactate production was significantly suppressed by LDHI administration in MiaPaca (a glycolytic pancreatic cancer cell line) tumors, as was the [1-13C]lactate to [1-13C]pyruvate ratio ([1-13C]-Lac/Pyr), which was calculated from the areas under the curves (AUC) using time-intensity data. This ratio decreased from 1.08 to 0.128 (88.1% decrease) 30 minutes after intravenous administration of the LDHI. In addition, hyperpolarized [1-13C]pyruvate MRS revealed that LDHI significantly suppressed lactate production in a dose dependent manner. Furthermore, Chemical Shift Imaging with 13C MRI demonstrated that the [1-13C]lactate signal in each voxel clearly decreased, compared to that before LDHI administration. The sum of [1-13C]lactate signals in the tumor region decreased after LDHI administration, resulting in a significant decrease in the tumor-specific [1-13C] Lac/Pyr ratio (1.463±0.31 before LDHI to 0.134±0.036 30 minutes after LDHI administration, a 90.67±2.56% decrease, n=3, p&amp;lt;0.01). [Conclusions] These results indicate that hyperpolarized 13C-MRI is a useful method to evaluate on-target efficacy of novel LDH inhibitors in vivo, and this technique can be used to determine optimum dose and exposure time of the LDHI in the tumor region. The current method can be of great value in providing an in vivo pharmacodynamic biomarker for this novel anti-cancer therapeutic targeting deregulated tumor metabolism. Citation Format: Nobu Oshima, Shun Kishimoto, Kristin Beebe, Keita Saito, Kazutoshi Yamamoto, Jeffery Brender, Anastasia Sowers, Ganesha Rai, Bryan T. Mott, David J. Maloney, James B. Mitchell, Murali K. Cherukuri, Leonard M. Neckers. Monitoring the impact on metabolic flux in vivo of a newly developed LDH inhibitor using hyperpolarized 13C magnetic resonance spectroscopic imaging [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 2852. doi:10.1158/1538-7445.AM2017-2852

Abstract 833: Alginate hydrogels for three-dimensional culture and real-time monitoring of cancer cell metabolism and radiation response

Abstract The purpose of this study was to characterize a three-dimensional calcium alginate culture system for real-time monitoring of cancer cell metabolism and radiation response. HCT116 colorectal cancer cells were encapsulated in 2.5% calcium alginate using a custom electrostatic rig adapted from a previously published system. Bead diameter was a function of voltage and alginate concentration: 4 x 107 cells/mL encapsulated at 4 and 8 kV resulted in mean diameters of 880 and 584 µm, respectively. The distribution of bead radii for all voltages showed positive excess kurtosis (4 kV = 1.6, 8 kV = 3.0) and 72.9% of 4 kV and 77.2% of 8 kV bead diameters fell within one standard deviation of the mean. Encapsulated cell viability was assessed by staining with Trypan Blue and clonogenic survival assays (CSAs). The baseline viability of cells immediately after encapsulation was 93.2%, which decreased 2 days after a single 10 Gy dose (90.7%, p&amp;lt;0.05). After two weeks of continuous culturing, 84.1% of non-irradiated and 73.8% of irradiated encapsulated cells remained viable (p&amp;lt;0.003). Pimonidazole staining demonstrated the presence of hypoxic cores proportional to bead size. CSAs of 8 kV beads showed a dose modifying factor (DMF) of 1.06 relative to cells grown in a monolayer, while larger 4 kV beads showed a DMF of 1.58. Mean oxygen consumption rate rose with increasing number of 8 kV beads/well: 1 bead, 35.5; 2 beads, 97.4; 3 beads, 175.5; 4 beads, 209.9; 2.0 x 104 monolayer cells, 92.6 (all pmol/min, ~8.2 x103 cells/bead, p&amp;lt;0.005). Hyperpolarized 13C-NMR spectroscopy of 400 uL of 8 kV beads containing 1.2 x 108 cells/mL showed a detectable conversion of [1-13C]-pyruvate to [1-13C]-lactate; 1 hour after a single 10 Gy dose, the lactate:pyruvate ratio decreased by 25%. Unpaired 2-tailed Student’s t-tests were used to determine significance between and one-way ANOVA was used to determine significance among groups with α set at p≤0.05. These data demonstrate the versatility of alginate hydrogels for real-time metabolic and radiation response studies which are non-invasive, higher throughput, and lower cost compared to in vivo systems. Future directions include additional metabolic flux analysis and hyperpolarized 13C-NMR spectroscopy to further investigate the cancer cell metabolic response to drug and/or radiation therapy. Citation Format: Aparna H. Kesarwala, Jenna L. Carter, Graham H. Read, Natsuko Miura, Kazutoshi Yamamoto, James B. Mitchell, Murali C. Krishna. Alginate hydrogels for three-dimensional culture and real-time monitoring of cancer cell metabolism and radiation response [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 833. doi:10.1158/1538-7445.AM2017-833

Analysis of extracellular brain chemistry during percutaneous dilational tracheostomy: A retrospective study of 19 patients

ObjectiveThe purpose of this study was to analyze changes in brain tissue chemistry around percutaneous dilational tracheostomy (PDT) in patients with acute brain injury (ABI) in a retrospective single-center analysis. Patients and methodsWe included 19 patients who had continuous monitoring of brain tissue chemistry and intracranial pressure (ICP) during a 20h period before and after PDT. Different microdialysis parameters (lactate, pyruvate, lactate pyruvate ratio (LPR), glycerol and glutamate) and values of ICP, cerebral perfusion pressure (CPP) and brain tissue oxygenation (PBrO2) were recorded per hour. Mean values were compared between a 10h period before PDT (prePDT) and after PDT (postPDT). ResultsMean values of cerebral lactate, pyruvate, LPR, glycerol and glutamate did not differ significantly between prePDT and postPDT. In addition, the rate of patients, which exceeded the known threshold was similar between prePDT and postPDT. Only one patient showed a strong increase of cerebral glycerol during the postPDT period, but analysis of subcutaneous glycerol could exclude an intracerebral event. ICP, CPP and PBrO2 did not exhibit significant changes. ConclusionsWe could exclude the occurrence of cerebral metabolic crisis and the excess release of cerebral glutamate and glycerol in a series of 19 patients. Our results support the safety of PDT in patients with ABI.

Metabolomic profling in gastric cancer tissues using time-of-flight mass spectrometry.

66 Background: To explore carcinogenic and prognostic biomarkers for gastric cancer, genomics, transcriptomics and proteomics approaches have been extensively applied;however, little has been investigated regarding the role of metabolomics profiles on progression and prognosis of gastric cancer. Therefore, in order to elucidate the role of metabolome on prognosis of gastric cancer, we investigated the metabolic profiles of gastric cancer tissue using time-of-flight mass spectrometry (TOFMS) Methods: A total of 162 patients with gastric cancer underwent gastrectomy from February 2010 to March 2013 were enrolled in this study. Cancer tissues (CA) and adjacent non-cancerous tissues (NC) were obtained from surgically resected sample and were snap-frozen in liquid nitrogen. The samples were then homogenized and then applied to capillary electrophoresis TOFMS (CE-TOFMS) The metabolomics dates were analyzed using principal component analysis (PCA) and hierachical clustering analysis(HCA) in order to compare the metabolic profiles of NC and CA. Metabolites date were further assessed according by the non-parametric Mann–Whitney U-test on the presence or absence of recurrence . Results: A total 96 metabolites were detected and quantified. PCA of the date well-distinguished CA from NC. In CA, lactate / pyruvate ratio was significantly higher, while adenylate energy charge was significantly lower than NC, which reaffirms the Warburug effect of cancer. Total glutathione and reducted glutathione/oxidized glutathione ratio in CA were significantly higher than in NC which possibly contributes to the homeostasis of redox status in CA. Intriguingly, in patients with recurrence, tumor concentrations of β-Ala, Asp, GDP and Gly were significantly lower than in those without recurrence. Conclusions: Metabolomic profiling clearly differentiated CA from NC. Considerably high lactate and amino acids levels expectedly highlighted the metabolome of tumors. Certain metabolite will be a candidate for biomarker in gastric cancer.

Lactate and Lactate: Pyruvate Ratio in the Diagnosis and Outcomes of Pediatric Acute Liver Failure

To assess the accuracy of blood lactate and lactate: pyruvate molar ratio (L:P) as a screen for mitochondrial, respiratory chain, or fatty acid oxidation disorders in children with pediatric acute liver failure (PALF); to determine whether serum lactate ≥ 2.5 mmol/L or L:P ≥ 25 correlated with biochemical variables of clinical severity; and to determine whether lactate or L:P is associated with clinical outcome at 21 days. Retrospective review of demographic, clinical, laboratory, and outcome data for PALF study group participants who had lactate and pyruvate levels collected on the same day. Of 986 participants, 110 had lactate and pyruvate levels collected on the same day. Of the 110, the etiology of PALF was a mitochondrial disorder in 8 (7%), indeterminate in 65 (59%), and an alternative diagnosis in 37 (34%). Lactate, pyruvate, and L:P were similar among the 3 etiologic groups. There was no significant association between the initial lactate or L:P and biochemical variables of clinical severity or clinical outcome at 21 days. A serum lactate ≥ 2.5 mmol/L and/or elevated L:P was common in all causes of PALF, not limited to those with a mitochondrial etiology, and did not predict 21-day clinical outcome. ClinicalTrials.gov: NCT00986648.

Lactate/pyruvate monitoring during carotid endarterectomy under general anaesthesia versus cervical plexus block: A randomised controlled study.

Background and Aims:Carotid endarterectomy (CEA) reduces the risk of stroke in patients with significant carotid stenosis and may be performed under general anaesthesia (GA) or regional anaesthesia (RA). This study aimed to compare RA and GA with regard the jugular venous bulb blood lactate and pyruvate levels.Methods:This randomised-controlled trial was done between October-2013 and September-2015. Thirty-six patients were randomised into either GA or RA groups, with six excluded after randomisation. In the RA group, combined deep and superficial cervical plexus blocks were performed. In the GA group, anaesthesia was induced with propofol and fentanyl. In both groups, monitoring of neurological function was done. Sampling of the contralateral jugular bulb blood was done. The main outcome measures were lactate and pyruvate in the jugular venous blood. For comparing categorical data, Chi-square test was used, and for the numerical variables, t-test was used.Results:Demographics were comparable in the two Groups. Serum lactate and pyruvate levels were higher in the GA group than RA group. At 120 min under anaesthesia, lactate and pyruvate levels under RA vs. GA, respectively were 0.76±0.03 mEq/L vs. 1.14±0.06, p-0.001 mEq/L, and 0.08± 0.00 mEq/L vs. 0.10±0.01 mEq/L, p=0.006. Lactate/ pyruvate ratios were normal in both groups. The mean blood pressure was significantly lower in the GA group during anaesthesia.Conclusion:In patients undergoing Carotid endarterectomy, serum levels of both lactate and pyruvate were higher under general versus regional anaesthesia.

линико-функциональные и метаболические нарушения и их коррекция у пациентов с дистальной сенсомоторной нейропатией

The clinical manifestations, biochemical and microhemocirculatory disorders, electroneuromographic (ENMG) changes in patients with distal sensomotor neuropathy and neuropathic pain syndrome including 16 persons of the comparison group (basic therapy) and 25 - of the main group (basic therapy + blood intravenous laser irradiation + nicergoline) at the time of hospitalization and after the course of treatment were studied. The patients showed a mixed, predominantly axonal type of nerve damage with the most pronounced changes in the ENMG indicators of sensory fibers in the lower limbs nerves. The use of blood intravenous laser irradiation and nicergoline contributes to a significant decrease in the activation of lipid peroxidation reactions and an increase in the activity of superoxide dismutase, while maintaining redox shifts characterized by lactic acid and an increase in the lactate / pyruvate ratio. Along with this, a significant reduction in the blood content of the substance P, was achieved in the patients of the main group. More effective correction of the disturbances in the vasomotor function of the endothelium was achieved. The obtained data confirm the pathogenetic validity of the use of blood intravenous laser irradiation and nicergoline for the treatment of patients with distal sensomotor neuropathy and neuropathic pain syndrome.

NON-INVASIVE ASSESSMENT OF METFORMIN INDUCED CHANGES IN CARDIAC AND HEPATIC REDOX STATE USING HYPERPOLARIZED [1-13C] PYRUVATE

Background and methods Metformin’s mechanism of action is controversial though may involve suppression of hepatic gluconeogenesis through perturbation of cellular redox state. Hyperpolarized magnetic resonance with [1-13C] pyruvate exploits huge increases in tracer signal to non-invasively assess both the redox coupled lactate dehydrogenase reaction and also pyruvate dehydrogenase activity. We performed hyperpolarized [1-13C] pyruvate magnetic resonance spectroscopic and imaging assessments of heart and liver metabolism in Wistar rats following either a single infusion of metformin or longer term oral treatment. Results Metformin infusion increased both cardiac and hepatic hyperpolarized [1-13C] lactate:[1-13C] pyruvate ratio (0.22 ± 0.04 versus 0.11 ± 0.01, P = 0.004 and 0.39 ± 0.04 versus 0.30 ± 0.02, P = 0.04 respectively) without changing pyruvate dehydrogenase flux. Metformin also increased both the cardiac and hepatic [lactate]:[pyruvate] ratio, a surrogate of cytosolic redox state (50 ± 8 versus 25 ± 7, P = 0.02 and 72 ± 12 versus 16 ± 8, P = 0.002 respectively). Longer term Metformin treatment for 4 weeks also increased the cardiac and hepatic hyperpolarized [1-13C] lactate:[1-13C] pyruvate ratio (0.27 ± 0.06 versus 0.10 ± 0.01, P = 0.005 and 0.87 ± 0.2 versus 0.36 ± 0.04, P = 0.003 respectively) without significantly changing pyruvate dehydrogenase flux and also increased the cardiac and hepatic [lactate]:[pyruvate] ratio (46 ± 6 versus 30 ± 6, P = 0.04 and 60 ± 9 versus 27 ± 3, P = 0.002 respectively). Conclusions Both acute and chronic metformin treatment significantly increased both the cardiac and hepatic hyperpolarized [1-13C] lactate:[1-13C] pyruvate ratio, reflecting an increase in cytosolic redox state. These findings 1) identify a novel cardiac effect of metformin, 2) demonstrate the sensitivity of hyperpolarized [1-13C] pyruvate to metformin induced changes in redox biology and 3) have implications for the design of upcoming human studies using hyperpolarized MR to study cardiac metabolism in diabetes.

Moderate Hypothermia Provides Better Protection of the Intestinal Barrier than Deep Hypothermia during Circulatory Arrest in a Piglet Model: A Microdialysis Study.

IntroductionThis study aimed to assess the effects of different temperature settings of hypothermic circulatory arrest (HCA) on intestinal barrier function in a piglet model.MethodsTwenty Wuzhishan piglets were randomly assigned to 40 min of HCA at 18°C (DHCA group, n = 5), 40 min of HCA at 24°C (MHCA group, n = 5), normothermic cardiopulmonary bypass (CPB group, n = 5) or sham operation (SO group, n = 5). Serum D-lactate (SDL) and lipopolysaccharide (LPS) levels were determined. Microdialysis parameters (glucose, lactate, pyruvate and glycerol) in the intestinal dialysate were measured. After 180 min of reperfusion, intestinal samples were harvested for real-time polymerase chain reaction and western blotting measurements for E-cadherin and Claudin-1.ResultsHigher levels of SDL and LPS were detected in the DHCA group than in the MHCA group (P < 0.001). Both MHCA and DHCA groups exhibited lower glucose levels, higher lactate and glycerol levels and a higher lactate to pyruvate (L/P) ratio compared with the CPB group (p<0.05); the DHCA group had higher lactate and glycerol levels and a higher L/P ratio (p<0.05) but similar glucose levels compared to the MHCA group. No significant differences in E-cadherin mRNA or protein levels were noted. Upregulation of claudin-1 mRNA levels was detected in both the DHCA and MHCA animals’ intestines (P < 0.01), but only the DHCA group exhibited a decrease in claudin-1 protein expression (P < 0.01).ConclusionHCA altered the energy metabolism and expression of epithelial junctions in the intestine. Moderate hypothermia (24°C) was less detrimental to the markers of normal functioning of the intestinal barrier than deep hypothermia (18°C).

Assessment of Metformin-Induced Changes in Cardiac and Hepatic Redox State Using Hyperpolarized[1-13C]Pyruvate.

Metformin improves cardiovascular outcomes in type 2 diabetes, but its exact mechanisms of action remain controversial. We used hyperpolarized [1-13C]pyruvate magnetic resonance spectroscopy to determine the effects of metformin treatment on heart and liver pyruvate metabolism in rats in vivo. Both oral treatment for 4 weeks and a single intravenous metformin infusion significantly increased the cardiac [1-13C]lactate:[1-13C]pyruvate ratio but had no effect on the [1-13C]bicarbonate + 13CO2:[1-13C]pyruvate ratio, an index of pyruvate dehydrogenase flux. These changes were paralleled by a significant increase in the heart and liver cytosolic redox state, estimated from the [lactate]:[pyruvate] ratio but not the whole-cell [NAD+]/[NADH] ratio. Hyperpolarized MRI localized the increase in cardiac lactate to the left ventricular myocardium, implying a direct myocardial effect, though metformin had no effect on systolic or diastolic cardiac function. These findings demonstrate the ability of hyperpolarized pyruvate magnetic resonance spectroscopy to detect metformin-induced changes in cytosolic redox biology, suggest that metformin has a previously unrecognized effect on cardiac redox state, and help to refine the design of impending hyperpolarized magnetic resonance studies in humans.

Cell-permeable succinate prodrugs bypass mitochondrial complex I deficiency.

Mitochondrial complex I (CI) deficiency is the most prevalent defect in the respiratory chain in paediatric mitochondrial disease. This heterogeneous group of diseases includes serious or fatal neurological presentations such as Leigh syndrome and there are very limited evidence-based treatment options available. Here we describe that cell membrane-permeable prodrugs of the complex II substrate succinate increase ATP-linked mitochondrial respiration in CI-deficient human blood cells, fibroblasts and heart fibres. Lactate accumulation in platelets due to rotenone-induced CI inhibition is reversed and rotenone-induced increase in lactate:pyruvate ratio in white blood cells is alleviated. Metabolomic analyses demonstrate delivery and metabolism of [13C]succinate. In Leigh syndrome patient fibroblasts, with a recessive NDUFS2 mutation, respiration and spare respiratory capacity are increased by prodrug administration. We conclude that prodrug-delivered succinate bypasses CI and supports electron transport, membrane potential and ATP production. This strategy offers a potential future therapy for metabolic decompensation due to mitochondrial CI dysfunction.

Mendeliome sequencing enables differential diagnosis and treatment of neonatal lactic acidosis.

BackgroundNeonatal lactic acidosis can be associated to severe inborn errors of metabolism. Rapid identification of the underlying disorder may improve the clinical management through reliable counseling of the parents and adaptation of the treatment.MethodsWe present the case of a term newborn with persistent hypoglycemia on postnatal day 1, who developed severe lactic acidosis, aggravating under intravenous glucose administration. Routine metabolic investigations revealed elevated pyruvate and lactate levels in urine, and magnetic resonance spectroscopy showed a lactic acid peak and decreased N-acetylaspartate levels. Mitochondrial disorders, e.g., pyruvate dehydrogenase (PDH) deficiency, were the major differential diagnoses. However, both hypoglycemia and the elevated lactate to pyruvate ratio in serum (=55.2) were not typical for PDH deficiency. We used “Mendeliome sequencing”, a next-generation sequencing approach targeting all genes which have been previously linked to single-gene disorders, to obtain the correct diagnosis.ResultsOn day 27 of life, we identified a homozygous stop mutation in the PDHX gene, causing pyruvate dehydrogenase E3-binding protein deficiency. After starting the ketogenic diet, the infant recovered and is showing delayed but progressive development.ConclusionsMendeliome sequencing was successfully used to disentangle the underlying cause of severe neonatal lactic acidosis. Indeed, it is one of several targeted sequencing approaches that allow rapid and reliable counseling of the parents, adaptation of the clinical management, and renunciation of unnecessary, potentially invasive and often costly diagnostic measures.Electronic supplementary materialThe online version of this article (doi:10.1186/s40348-016-0050-x) contains supplementary material, which is available to authorized users.

Combining hyperpolarized 13 C MRI with a liver-specific gadolinium contrast agent for selective assessment of hepatocyte metabolism.

Hyperpolarized 13 C MRI is a powerful tool for studying metabolism, but can lack tissue specificity. Gadoxetate is a gadolinium-based MRI contrast agent that is selectively taken into hepatocytes. The goal of this project was to investigate whether gadoxetate can be used to selectively suppress the hyperpolarized signal arising from hepatocytes, which could in future studies be applied to generate specificity for signal from abnormal cell types. Baseline gadoxetate uptake kinetics were measured using T1 -weighted contrast enhanced imaging. Relaxivity of gadoxetate was measured for [1-13 C]pyruvate, [1-13 C]lactate, and [1-13 C]alanine. Four healthy rats were imaged with hyperpolarized [1-13 C]pyruvate using a three-dimensional (3D) MRSI sequence prior to and 15 min following administration of gadoxetate. The lactate:pyruvate ratio and alanine:pyruvate ratios were measured in liver and kidney. Overall, the hyperpolarized signal decreased approximately 60% as a result of pre-injection of gadoxetate. In liver, the lactate:pyruvate and alanine:pyruvate ratios decreased 42% and 78%, respectively (P < 0.05) following gadoxetate administration. In kidneys, these ratios did not change significantly. Relaxivity of gadoxetate for [1-13 C]alanine was 12.6 times higher than relaxivity of gadoxetate for [1-13 C]pyruvate, explaining the greater selective relaxation effect on alanine. The liver-specific gadolinium contrast-agent gadoxetate can selectively suppress normal hepatocyte contributions to hyperpolarized 13 C MRI signals. Magn Reson Med 77:2356-2363, 2017. © 2016 International Society for Magnetic Resonance in Medicine.

Metabolomics of Human Amniotic Fluid and Maternal Plasma during Normal Pregnancy.

Metabolic profiles of amniotic fluid and maternal blood are sources of valuable information about fetus development and can be potentially useful in diagnosis of pregnancy disorders. In this study, we applied 1H NMR-based metabolic profiling to track metabolic changes occurring in amniotic fluid (AF) and plasma (PL) of healthy mothers over the course of pregnancy. AF and PL samples were collected in the 2nd (T2) and 3rd (T3) trimester, prolonged pregnancy (PP) until time of delivery (TD). A multivariate data analysis of both biofluids reviled a metabolic switch-like transition between 2nd and 3rd trimester, which was followed by metabolic stabilization throughout the rest of pregnancy probably reflecting the stabilization of fetal maturation and development. The differences were further tested using univariate statistics at α = 0.001. In plasma the progression from T2 to T3 was related to increasing levels of glycerol, choline and ketone bodies (3-hydroxybutyrate and acetoacetate) while pyruvate concentration was significantly decreased. In amniotic fluid, T2 to T3 transition was associated with decreasing levels of glucose, carnitine, amino acids (valine, leucine, isoleucine, alanine, methionine, tyrosine, and phenylalanine) and increasing levels of creatinine, succinate, pyruvate, choline, N,N-dimethylglycine and urocanate. Lactate to pyruvate ratio was decreased in AF and conversely increased in PL. The results of our study, show that metabolomics profiling can be used to better understand physiological changes of the complex interdependencies of the mother, the placenta and the fetus during pregnancy. In the future, these results might be a useful reference point for analysis of complicated pregnancies.

Neuron-Specific Enolase Is Correlated to Compromised Cerebral Metabolism in Patients Suffering from Acute Bacterial Meningitis; An Observational Cohort Study.

IntroductionPatients suffering from acute bacterial meningitis (ABM) with a decreased level of consciousness have been shown to have an improved clinical outcome if treated with an intracranial pressure (ICP) guided therapy. By using intracranial microdialysis (MD) to monitor cerebral metabolism in combination with serum samples of biomarkers indicating brain tissue injury, S100B and Neuron Specific Enolase (NSE), additional information might be provided. The aim of this study was to evaluate biomarkers in serum and MD parameters in patients with ABM.MethodsFrom a prior study on patients (n = 52) with a confirmed ABM and impaired consciousness (GCS ≤ 9, or GCS = 10 combined with lumbar spinal opening pressure > 400 mmH2O), a subgroup of patients (n = 21) monitored with intracerebral MD and biomarkers was included in the present study. All patients were treated in the NICU with intracranial pressure (ICP) guided therapy. Serum biomarkers were obtained at admission and every 12 hours. The MD parameters glucose, lactate, pyruvate and glycerol were analyzed. Outcome was assessed at 12–55 months after discharge from hospital. Mann-Whitney U-Test and Wilcoxon matched-pairs signed rank test were applied.ResultsThe included patients had a mean GCS of 8 (range, 3–10) on admission and increased ICP (>20 mmHg) was observed in 62% (n = 13/21) of the patients. Patients with a lactate:pyruvate ratio (LPR) >40 (n = 9/21, 43%) had significantly higher peak levels of serum NSE (p = 0.03), with similar, although non-significant observations made in patients with high levels of glycerol (>500 μmol/L, p = 0.11) and those with a metabolic crisis (Glucose <0.8 mmol/L, LPR >25, p = 0.09). No associations between serum S100B and MD parameters were found. Furthermore, median MD glucose levels decreased significantly between day 1 (0–24h) and day 3 (48–72h) after admission to the NICU (p = 0.0001). No correlation between MD parameters or biomarkers and outcome was found.ConclusionIn this observational cohort study, we were able to show that cerebral metabolism is frequently affected in patients with ABM. Furthermore, patients with high LPR (LPR>40) had significantly higher levels of NSE, suggesting ongoing deterioration in compromised cerebral tissue. However, the potential clinical impact of MD and biomarker monitoring in ABM patients will need to be further elaborated in larger clinical trials.

Transient neonatal renal failure and massive polyuria in MEGDEL syndrome

BackgroundMEGDEL (3-methylglutaconic aciduria with deafness, encephalopathy, and Leigh-like syndrome) syndrome is a mitochondrial disorder associated with recessive mutations in SERAC1. ObjectivesTo report transient neonatal renal findings in MEGDEL syndrome. ResultsThis 7year-old girl was the first child of consanguineous Turkish parents. She exhibited an acute neonatal deterioration with severe lactic acidosis and liver failure. Initial evaluation revealed massive polyuria and renal failure with 3-methylglutaconic aciduria. Symptoms and biological findings progressively improved with symptomatic treatment but lactic acidosis and high lactate to pyruvate ratio along with 3-methylglutaconic aciduria persisted. At 8months of age, a subacute neurological degradation occurred with severe hypotonia, dystonia with extrapyramidal movements and failure to thrive. Brain MRI revealed basal ganglia lesions suggestive of Leigh syndrome. At 3years of age, sensorineural deafness was documented. MEGDEL syndrome was further confirmed by the identification of an already reported homozygous mutation in SERAC1. ConclusionTransient neonatal polyuria and renal failure have not been reported to date in SERAC1 defective patients. Such neonatal kidney findings expand the clinical spectrum of MEGDEL syndrome.

Enhanced production of N-acetyl-d-neuraminic acid by whole-cell bio-catalysis of Escherichia coli

N-acetyl-d-neuraminic acid (Neu5Ac) has been considerably focused due to its promising potential applications in pharmaceuticals and dairy products. A whole-cell biocatalyst process is an important tool for synthesis of pharmaceutical intermediates and fine chemicals. In this study, a whole cell process using engineered Escherichia coli strain was developed and stepwise optimization was employed for Neu5Ac production. N-acetyl-D-glucosamine 2-epimerase and Neu5Ac aldolase were overexpressed in E. coli individually and the activity ratio was optimized by varying recombinant amounts of cell biomass for synthesis of Neu5Ac. Moreover, substrate concentrations and ratio of pyruvate and N-acetyl-D-glucosamine (GlcNAc) and detergent concentrations were optimized to increase product synthesis. The resulting process generated 237.4mM Neu5Ac with a yield of 40.0% mol/mol GlcNAc. Furthermore, transporter pathways involved in Neu5Ac and GlcNAc were engineered and their impact on the Neu5Ac synthesis was evaluated. Using a stepwise optimization, an overall whole-cell biocatalytic process was developed and a maximum titer of 260.0mM Neu5Ac (80.4g/L) with a conversion yield of 43.3% from GlcNAc was achieved. The process can be used for industrial large-scale production of Neu5Ac in terms of efficiency and economy.

Short-term prognostic value of perioperative coronary sinus-derived-serum cardiac troponin-I, creatine kinase-MB, lactate, pyruvate, and lactate-pyruvate ratio in adult patients undergoing open heart surgery.

Objectives:To investigate the release pattern of different cardiac metabolites and biomarkers directly from the coronary sinus (CS) and to establish the diagnostic discrimination limits of each marker protein and metabolites to evaluate perioperative myocardial injury in patients undergoing cardiac surgery under cardiopulmonary bypass (CPB).Patients and Methods:Sixty-eight patients undergoing first mitral and/or aortic valve replacements with/without coronary artery bypass grafting and Bentall procedure under CPB and blood cardioplegic arrest were studied. All cardiac metabolites and biomarkers were measured in serial CS-derived blood samples at pre-CPB, immediate post aortic declamping, 10 minutes post-CPB and 12 hrs post-CPB.Results:Receiver operating characteristic curve analysis of cardiac biomarkers indicated lactate-pyruvate ratio as the superior diagnostic discriminator of myocardial injury with an optimal “cut-off” value >10.8 immediately after aortic declamping (AUC, 0.92; 95% CI: 0.85-0.98). Lactate was the second best diagnostic discriminator of myocardial injury with an optimal “cut-off” value >2mmol/l at immediately after aortic declamping (AUC, 0.89; 95% CI: 0.80-0.96). Cardiac troponin-I was the third best diagnostic discriminator of myocardial injury with an optimal “cut-off” value >2.1ng/ml at immediately after aortic declamping (AUC, 0.88; 95% CI: 0.80-0.95). Creatine kinase-MB was the fourth best diagnostic discriminator of myocardial injury with an optimal “cut-off” value >58 log units/ml prior to decanulation (AUC, 0.85; 95% CI: 0.78-0.94).Conclusions:Measurable cardiac damage exists in all patients undergoing cardiac surgery under cardioplegic arrest. The degree of myocardial injury is more in patients with poor ventricular function and those requiring longer aortic clamp time. CS-derived lactate-pyruvate ratio, lactate, cTn-I served as superior diagnostic discriminators of peri-operative myocardial damage.