Exogenous Lactate Research Articles

Hypoxia-induced autophagy of stellate cells inhibits expression and secretion of lumican into microenvironment of pancreatic ductal adenocarcinoma.

Lumican is secreted by pancreatic stellate cells and inhibits cancer progression. Extracellular lumican inhibits cancer cell replication and restrains growth of early-stage pancreatic adenocarcinoma (PDAC) such that patients with tumors containing stromal lumican experience a three-fold longer survival after treatment. In the present study, patient tumor tissues, ex-vivo cultures of patient-derived xenografts (PDX), PDAC stellate and tumor cells were used to investigate whether hypoxia (1% O2) within the tumor microenvironment influences stromal lumican expression and secretion. We observed that hypoxia significantly reduced lumican expression and secretion from pancreatic stellate cells, but not cancer cells. Although hypoxia enhanced lactate dehydrogenase A (LDHA) expression and lactate secretion from all cells, neither hypoxia-induced nor exogenous lactate influenced lumican expression. Autophagy was induced by hypoxia in ex vivo cultures of PDX and pancreatic stellate cells, but not cancer cells cultured in 2D. Autophagic flux inhibitors, bafilomycin A1, chloroquine diphosphate salt, and ammonium chloride prevented hypoxia-mediated reduction in lumican expression in stellate cells. Furthermore, inhibition of AMP-regulated protein kinase (AMPK) phosphorylation or hypoxia-inducible factor (HIF)-1α expression within hypoxic stellate cells restored lumican expression levels. Hypoxia did not affect lumican mRNA expression, indicating that hypoxia-induced reduction of lumican occurs post-transcriptionally; in addition, AMPK inhibition prevented hypoxia-reduced phosphorylation of the mTOR/p70S6K/4EBP signaling pathway, a key contributor to protein synthesis. Taken together, these findings demonstrate that hypoxia reduces stromal lumican in PDAC through autophagy-mediated degradation and reduction in protein synthesis within pancreatic cancer stellate cells.

The Hyperinsulinemic Isoglycemic Hyperlactemic Clamp

The Cori Cycle (glucose-lactate interchange) dominates carbohydrate fluxes. Glucose-to-lactate flux follows peripheral disposal; lactate-to-glucose occurs during gluconeogenesis. As glucose flux can be assessed by the change in an exogenous glucose infusion rate (GIR) required to maintain a constant concentration (glucose clamp), so too can lactate flux be assessed from an exogenous lactate infusion rate (LIR) required to maintain isolactemia. Simultaneous assessment of both fluxes will enable estimation of lactate-derived gluconeogenesis (GNG), and its contribution to the rate of glucose appearance (Ra) without using radiolabeled tracers, the mainstay of flux measurement. Anesthetized Sprague-Dawley rats were intubated, catheterized via artery and vein, and partially pancreatectomized to eliminate endogenous glucagon secretion. Establishment of a hyperinsulinemic (100mU/kg/h), hyperglycemic (7mM) clamp was followed by a hyperlactemic (4mM) clamp via independent variable infusions of glucose/Na-lactate in response to 5-min measures of plasma concentrations (YSI 2950D Analyzer with glucose/lactate electrodes). Hyperlactemia (∼10x resting) rendered it the dominant gluconeogenic substrate. After at least 30 min of isolactemia, glucagon was infused for 60 min (0, 1, 3, 10 or 50 µg/kg/h) to enable dose/concentration response analysis. Glucagon administration dose-dependently reduced the GIR and increased the LIR necessary to maintain isoglycemia/isolactemia (respective ED50’s 1.33, 0.98 µg/kg/h). δ-GIR of up to -12.2 mmol/kg/h potentially represented increased glucose Ra. δ-LIR of up to +8.6 mmol/kg/h potentially represented GNG. Since 2 lactate moieties are required per glucosyl, GNG contributed ∼35% of glucagon-stimulated glucose Ra (∼65% being glycogenolysis). In conclusion, the hyperinsulinemic isoglycemic hyperlactemic clamp enables a quantitative assessment of hepatic processes, complementing that of peripheral processes revealed with the glucose clamp. Disclosure D. Zaretsky: Employee; Self; Intarcia Therapeutics, Inc. A.A. Young: Employee; Self; Intarcia Therapeutics, Inc.. Stock/Shareholder; Self; GlaxoSmithKline plc..

The Warburg Effect and Lactate Signaling Augment Fgf Signaling to Promote Sensory‐neural Development in the Otic Vesicle

In a forward ENU screen in zebrafish, we recovered a novel mutation that specifically impairs development of sensory hair cells and neurons in the otic vesicle. Whole‐genome sequencing identified a null mutation in the gene encoding the glycolytic enzyme Phosphoglycerate Kinase‐1 (Pgk1). Targeted disruption of Pgk1 using CRISPR/Cas9 yields an identical phenotype. Surprisingly, analysis of genetic mosaics showed that Pgk1 acts non‐autonomously to promote efficient Fgf signaling at a distance, as required for specification of neurons and hair cells of the inner ear. Expression of known Fgf ligands appears normal but Fgf‐reporters Pea3 and Erm are dramatically under‐expressed in mutant embryos. Moreover, transient overexpression of Fgf8 from an inducible transgene does not rescue the phenotype, suggesting a problem with signal transduction. These and other data led us to the discovery that otic cells require metabolic reprogramming to enhance glycolysis and “aerobic fermentation” – similar to the “Warburg Effect” exhibited by tumor cells – to promote synthesis and secretion of lactate. Wild‐type embryos treated with drugs that block synthesis or secretion of lactate mimic the phenotype of Pgk1 mutants, whereas Pgk1 mutants are fully rescued by treatment with exogenous lactate. Lactate treatment of wild‐type embryos elevates expression of Pea3 and Erm even when Fgf signaling is blocked. Thus, by raising steady‐state levels of Pea3 and Erm (critical effectors of the Fgf‐MAPK pathway), lactate signaling primes the pathway and renders cells more responsive to dynamic changes in Fgf signaling required for sensory‐neural development in the inner ear.Support or Funding InformationNIH‐NIDCD grant R01‐DC03806This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Abstract B021: Inhibition of β-catenin impairs triple-negative breast cancer (TNBC) cell migration and invasion by modulating aerobic glycolysis components

Abstract In the present study, we investigated the role of β-catenin on aerobic glycolysis in breast cancer cells and further evaluated its effects on triple-negative breast cancer (TNBC) cell migration and invasion. We used two metastatic TNBC cell lines, i.e., MDA-MB-468 and MDA-MB-231, and treated them with β-catenin antagonists (WNT5A and XAV939). Evaluation of glycolytic markers in WNT5A-expressing cell lines showed selective downregulation in the expression of β-catenin, platelet type-phosphofructokinase (PFKP), and lactate secretion, without affecting cell proliferation. Of the three phosphofructokinase isoform, we found that only PFKP expression was associated with decreased survival in breast cancer patients. We also found that the cause of PKFP and lactate downregulation was mediated by inhibition of β-catenin signaling, which is in accordance with the finding that β-catenin signaling positively regulates lactate production. Functionally, we showed that specific downregulation of β-catenin resulted in reduced lactate production accompanied by impaired TNBC cells migration and invasion. Finally, we demonstrated that apart from its inhibitory effect on lactate production, inhibition of β-catenin can also impair TNBC cell migration induced by exogenous lactate via downregulating monocarboxylate transporter 1 (MCT1). Overall, we demonstrated that inhibition of β-catenin-PFKP signaling axis impaired lactate production as well as its uptake from tumor microenvironment (via MCT1), thereby impairing TNBC cells' migration. Citation Format: Chandra P. Prasad, Tommy Andersson. Inhibition of β-catenin impairs triple-negative breast cancer (TNBC) cell migration and invasion by modulating aerobic glycolysis components [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2017 Oct 26-30; Philadelphia, PA. Philadelphia (PA): AACR; Mol Cancer Ther 2018;17(1 Suppl):Abstract nr B021.

Lactate-mediated mitoribosomal defects impair mitochondrial oxidative phosphorylation and promote hepatoma cell invasiveness

Impaired mitochondrial oxidative phosphorylation (OXPHOS) capacity, accompanied by enhanced glycolysis, is a key metabolic feature of cancer cells, but its underlying mechanism remains unclear. Previously, we reported that human hepatoma cells that harbor OXPHOS defects exhibit high tumor cell invasiveness via elevated claudin-1 (CLN1). In the present study, we show that OXPHOS-defective hepatoma cells (SNU354 and SNU423 cell lines) exhibit reduced expression of mitochondrial ribosomal protein L13 (MRPL13), a mitochondrial ribosome (mitoribosome) subunit, suggesting a ribosomal defect. Specific inhibition of mitoribosomal translation by doxycycline, chloramphenicol, or siRNA-mediated MRPL13 knockdown decreased mitochondrial protein expression, reduced oxygen consumption rate, and increased CLN1-mediated tumor cell invasiveness in SNU387 cells, which have active mitochondria. Interestingly, we also found that exogenous lactate treatment suppressed MRPL13 expression and oxygen consumption rate and induced CLN1 expression. A bioinformatic analysis of the open RNA-Seq database from The Cancer Genome Atlas (TCGA) liver hepatocellular carcinoma (LIHC) cohort revealed a significant negative correlation between MRPL13 and CLN1 expression. Moreover, in patients with low MRPL13 expression, two oxidative metabolic indicators, pyruvate dehydrogenase B expression and the ratio of lactate dehydrogenase type B to type A, significantly and negatively correlated with CLN1 expression, indicating that the combination of elevated glycolysis and deficient MRPL13 activity was closely linked to CLN1-mediated tumor activity in LIHC. These results suggest that OXPHOS defects may be initiated and propagated by lactate-mediated mitoribosomal deficiencies and that these deficiencies are critically involved in LIHC development.

Astrocytic l-Lactate Signaling Facilitates Amygdala-Anterior Cingulate Cortex Synchrony and Decision Making in Rats

Human brain imaging studies have revealed the anterior cingulate cortex (ACC) as a key brain region for mediating visceral-pain-cognitive interactions. Recently, we characterized impairments of long-term potentiation and spike-field coherence in the basolateral amygdala (BLA)-ACC network in association with a decision-making deficit in rats with visceral hypersensitivity (VH). Now, by combining integrative neurobiological approaches, we show that ACC-reactive astrogliosis and activity-dependent impairment of lactate release occur in VH rats. Exogenous lactate supply rescues chronic-visceral-pain-caused impairments of ACC phase locking and decision making, which can be mimicked by optogenetic activation of ACC astrocytes. Large-scale electrophysiological recordings in free-moving animals during a decision-making task indicate that optogenetic astrocytic activation improves decision-making performance and engages ACC phase locking and BLA-to-ACC information flow. Collectively, these observations support the idea of an "astrocyte-neuron l-lactate shuttle" and suggest that targeting astrocytes may help with cognitive dysfunctions under chronic visceral pain.

Reduced production and uptake of lactate are essential for the ability of WNT5A signaling to inhibit breast cancer cell migration and invasion.

Here we investigated the impact of WNT5A signaling on aerobic glycolysis and evaluated its effects on breast cancer cell migration/invasion. WNT5A signaling reduced migration and lactate production and caused selective down-regulation of the glycolytic enzyme phosphofructokinase platelet-type (PFKP). These events occurred in parallel with a WNT5A-induced inhibition of β-catenin signaling. Support for essential involvement of β-catenin and PFKP in lactate production and migration/invasion was obtained by siRNA knockdown of their expression. To also explore the effect of non-tumor cell-derived lactate, we added exogenous lactate to the cells and noted an increase in migration that was significantly impaired by recombinant WNT5A in parallel with a down-regulation of the lactate transporter monocarboxylate transporter 1 (MCT1). Interestingly enough, the drug-candidate Foxy5 (WNT5A-mimic hexapeptide) also inhibited breast cancer cell migration in the presence of exogenous lactate, suggesting a therapeutic potential for Foxy5 in managing breast tumors with high glycolytic activity. Overall, we demonstrated that WNT5A signaling (via a β-catenin-PFKP axis) reduces lactate production and lowers the expression of MCT1, a carrier mediating the uptake of lactate from the tumor microenvironment. These effects of WNT5A are essential for its ability to impair breast cancer migration/invasion even in an environment with elevated lactate levels.

Lactate metabolism is associated with mammalian mitochondria.

It is well established that lactate secreted by fermenting cells can be oxidized or used as a gluconeogenic substrate by other cells and tissues. Within the fermenting cell itself, however, it is generally assumed that lactate is produced to replenish NAD+ and then is secreted. Here we explored the possibility that cytosolic lactate is metabolized by the mitochondria of fermenting mammalian cells. We found that fermenting HeLa and H460 cells utilize exogenous lactate carbon to synthesize a large percentage of their lipids. With high-resolution mass spectrometry, we found that both 13C and 2-2H labels from enriched lactate enter the mitochondria. The lactate dehydrogenase (LDH) inhibitor oxamate decreased respiration of isolated mitochondria incubated in lactate, but not isolated mitochondria incubated in pyruvate. Additionally, transmission electron microscopy (TEM) showed that LDHB localizes to the mitochondria. Taken together, our results demonstrate a link between lactate metabolism and the mitochondria of fermenting mammalian cells.

Effects of dexmedetomidine and esmolol on systemic hemodynamics and exogenous lactate clearance in early experimental septic shock.

BackgroundPersistent hyperlactatemia during septic shock is multifactorial. Hypoperfusion-related anaerobic production and adrenergic-driven aerobic generation together with impaired lactate clearance have been implicated. An excessive adrenergic response could contribute to persistent hyperlactatemia and adrenergic modulation might be beneficial. We assessed the effects of dexmedetomidine and esmolol on hemodynamics, lactate generation, and exogenous lactate clearance during endotoxin-induced septic shock.MethodsEighteen anesthetized and mechanically ventilated sheep were subjected to a multimodal hemodynamic/perfusion assessment including hepatic and portal vein catheterizations, total hepatic blood flow, and muscle microdialysis. After monitoring, all received a bolus and continuous infusion of endotoxin. After 1 h they were volume resuscitated, and then randomized to endotoxin-control, endotoxin-dexmedetomidine (sequential doses of 0.5 and 1.0 μg/k/h) or endotoxin-esmolol (titrated to decrease basal heart rate by 20 %) groups. Samples were taken at four time points, and exogenous lactate clearance using an intravenous administration of sodium L-lactate (1 mmol/kg) was performed at the end of the experiments.ResultsDexmedetomidine and esmolol were hemodynamically well tolerated. The dexmedetomidine group exhibited lower epinephrine levels, but no difference in muscle lactate. Despite progressive hypotension in all groups, both dexmedetomidine and esmolol were associated with lower arterial and portal vein lactate levels. Exogenous lactate clearance was significantly higher in the dexmedetomidine and esmolol groups.ConclusionsDexmedetomidine and esmolol were associated with lower arterial and portal lactate levels, and less impairment of exogenous lactate clearance in a model of septic shock. The use of dexmedetomidine and esmolol appears to be associated with beneficial effects on gut lactate generation and lactate clearance and exhibits no negative impact on systemic hemodynamics.

Exogenous lactate infusion improved neurocognitive function of patients with mild traumatic brain injury.

Background:Many studies showed a better recovery of cognitive function after administration of exogenous lactate during moderate-severe traumatic brain injury. However, the study evaluating lactate effect on mild traumatic brain injury is still limited.Aims:To evaluate the effect of exogenous lactate on cognitive function in mild traumatic brain injury patients.Settings and Design:Prospective, single blind, randomized controlled study on 60 mild traumatic brain injury patients who were undergoing neurosurgery.Materials and Methods:Subjects were randomly assigned into hyperosmolar sodium lactate (HSL) group or hyperosmolar sodium chloride (HSS) group. Patients in each group received either intravenous infusion of HSL or NaCl 3% at 1.5 ml/KgBW within 15 min before neurosurgery. During the surgery, patients in both groups received maintenance infusion of NaCl 0.9% at 1.5 ml/KgBW/hour.Statistical Analysis:Cognitive function, as assessed by Mini-Mental State Examination (MMSE) score at 24 h, 30 and 90 days post-surgery, was analyzed by Anova repeated measures test.Results:The MMSE score improvement was significantly better in HSL group than HSS group (P < 0.001). In HSL group the MMSE score improved from 16.00 (13.75-18.00) at baseline to 21.00 (18.75-22.00); 25.00 (23.75-26.00); 28.00 (27.00-29.00) at 24 h, 30, 90 days post-surgery, respectively. In contrast, in HSS group the MMSE score almost unchanged at 24 h and only slightly increased at 30 and 90 days post-surgery.Conclusions:Hyperosmolar sodium lactate infusion during mild traumatic brain injury improved cognitive function better than sodium chloride 3%.

Physiological Concentration of Exogenous Lactate Reduces Antimycin A Triggered Oxidative Stress in Intestinal Epithelial Cell Line IPEC-1 and IPEC-J2 In Vitro.

Weaning triggers an adaptation of the gut function including luminal lactate generation by lactobacilli, depending on gastrointestinal site. We hypothesized that both lactobacilli and lactate influence porcine intestinal epithelial cells. In vivo experiments showed that concentration of lactate was significantly higher in gastric, duodenal and jejunal chyme of suckling piglets compared to their weaned counterparts. In an in vitro study we investigated the impact of physiological lactate concentration as derived from the in vivo study on the porcine intestinal epithelial cells IPEC-1 and IPEC-J2. We detected direct adherence of lactobacilli on the apical epithelial surface and a modulated F-actin structure. Application of lactobacilli culture supernatant alone or lactate (25 mM) at low pH (pH 4) changed the F-actin structure in a similar manner. Treatment of IPEC cultures with lactate at near neutral pH resulted in a significantly reduced superoxide-generation in Antimycin A-challenged cells. This protective effect was nearly completely reversed by inhibition of cellular lactate uptake via monocarboxylate transporter. Lactate treatment enhanced NADH autofluorescence ratio (Fcytosol/Fnucleus) in non-challenged cells, indicating an increased availability of reduced nucleotides, but did not change the overall ATP content of the cells. Lactobacilli-derived physiological lactate concentration in intestine is relevant for alleviation of redox stress in intestinal epithelial cells.

Abstract A47: A microRNA signature identifies subtypes of triple-negative breast cancer and reveals miR-342-3p as regulator of a lactate metabolic pathway through silencing monocarboxylate transporter 1

Abstract Triple negative breast cancer (TNBC) represents a challenging tumor type due to their poor prognosis and limited treatment options. It is well recognize that clinical and molecular heterogeneity of TNBC is driven in part by post-transcriptional regulators such as miRNAs. To stratify TNBCs, we profiled 1050 miRNAs in 132 adjuvant TNBC tumors and 40 tumors from other immunophenotypes using an Affymetrix microarray platform. A NMF clustering analysis allowed us to identify 4 TNBC subtypes featuring unique miRNA expression patterns, disease free and overall survival rates and particular gene ontology enrichments (performed with GSEA algorithm). Our agglomerative approach was cross-validated by using two other clustering algorithms (k-means and consensus clustering). TNBC miRNAs subgroups were also correlated with the Lehmann intrinsic subtypes, finding a significant enrichment of immmnomodulartory and basal 1 subtypes in our high risk miRNA subgroups. 3 cell line models (MDA MB 468, MDA MB 231 and HS578T) were classified according to our miRNA signature, recapitulating two different miRNA subgroups. The TNBC tumors were compared against other phenotypes identifying differentially expressed miRNAs that together with the altered miRNAs within the subgroups allowed us to define interesting miRNAs for further functional analysis. We found low expression levels of miR-342-3p in TNBC tumors compared with other breast cancer phenotypes, and this down-regulation characterizes one of our miRNA subgroups with high risk to relapse. To characterize the functional its functional role, miR-342-3p was transiently transfected in the cell line MDA-MB-468, showing a decrease in cell proliferation, viability and migration rates. A gene expression profile (Human gene st, Affymetrix) revealed 140 altered mRNAs, from which 35 are potential direct targets of miR-342-3p defined by an in-silico analysis. The monocarboxylate transporter 1(MCT1), was confirmed as one target of miR-342-3p by a luciferase assay and western blot analysis. MCT1 repression by the miRNA promotes lactate efflux changes in the tumor cells, reflected in the accumulation of exogenous lactate and the increase in levels of extracellular endogenous lactate together with a decrease level of intra and inter cellular glucose concentration. We also explored other mechanisms that can contribute to the modulation of lactate and glucose levels such as the expression of MCT4, HIF1alpha and Glut1, but not significant changes were observed, pointing out the important role of the regulatory circuit of miR-342-3p and MCT1 in a particular TNBC tumor cells metabolism. These data suggest a metabolic change that favors a more glycolytic environment, which lead to a glucose deprivation context that may contribute to the reduction in proliferation, viability and migration capabilities already described. Furthermore, to define the main transcriptional modulator of the intronic miR-342-3p in TNBC, we evaluated the correlation between the expression of estrogen receptor, miR-342-3p and its host gene EVL in TNBC tumors from TCGA data. A significant correlation between these transcripts was observed. These results, together with information from other studies and Chip-seq analysis suggest that the ER activates the transcription of EVL and consequently the intronic miR-342-3p. We validated these correlations in our cellular model by transiently silencing of the ER. Our data provides evidence that miRNAs sub-classification signature can recapitulate the biological and clinical heterogeneity of TNBC. Moreover, show that miRNAs regulate important oncogenic pathways in TNBC cells such as: proliferation, cell movement and apoptosis in the different subgroups. Citation Format: Sandra L. Romero-Cordoba, Sergio Rodriguez-Cuevas, Rosa Rebollar-Vega, Veronica Bautista-Pina, Antonio Maffuz-Aziz, Elda Tagliabue, Marilena Iorio, Elvira D'Ippolito, Sara Baroni, Ilaria Plantamura, Alfredo Hidalgo-Miranda. A microRNA signature identifies subtypes of triple-negative breast cancer and reveals miR-342-3p as regulator of a lactate metabolic pathway through silencing monocarboxylate transporter 1. [abstract]. In: Proceedings of the AACR Special Conference on Noncoding RNAs and Cancer: Mechanisms to Medicines ; 2015 Dec 4-7; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2016;76(6 Suppl):Abstract nr A47.

Cerebral Lactate Metabolism After Traumatic Brain Injury.

Cerebral energy dysfunction has emerged as an important determinant of prognosis following traumatic brain injury (TBI). A number of studies using cerebral microdialysis, positron emission tomography, and jugular bulb oximetry to explore cerebral metabolism in patients with TBI have demonstrated a critical decrease in the availability of the main energy substrate of brain cells (i.e., glucose). Energy dysfunction induces adaptations of cerebral metabolism that include the utilization of alternative energy resources that the brain constitutively has, such as lactate. Two decades of experimental and human investigations have convincingly shown that lactate stands as a major actor of cerebral metabolism. Glutamate-induced activation of glycolysis stimulates lactate production from glucose in astrocytes, with subsequent lactate transfer to neurons (astrocyte-neuron lactate shuttle). Lactate is not only used as an extra energy substrate but also acts as a signaling molecule and regulator of systemic and brain glucose use in the cerebral circulation. In animal models of brain injury (e.g., TBI, stroke), supplementation with exogenous lactate exerts significant neuroprotection. Here, we summarize the main clinical studies showing the pivotal role of lactate and cerebral lactate metabolism after TBI. We also review pilot interventional studies that examined exogenous lactate supplementation in patients with TBI and found hypertonic lactate infusions had several beneficial properties on the injured brain, including decrease of brain edema, improvement of neuroenergetics via a "cerebral glucose-sparing effect," and increase of cerebral blood flow. Hypertonic lactate represents a promising area of therapeutic investigation; however, larger studies are needed to further examine mechanisms of action and impact on outcome.

Conversion of l-lactate into n-caproate by a continuously fed reactor microbiome

Conversion of lactate to n-caproate had been described for the type strain Megasphaera elsdenii in batch systems. Recently, investigators have also described production of n-caproate from endogenous or exogenous lactate with batch-fed reactor microbiome systems. However, no reports exist of lactate to n-caproate conversion within a continuously fed bioreactor. Since continuously fed systems are advantageous for biotechnology production platforms, our objective was to develop such a system. Here, we demonstrated continuous lactate to n-caproate conversion for more than 165 days. The volumetric n-caproate production rate (productivity) was improved when we decreased the operating pH from 5.5 to 5.0, and was again improved when we utilized in-line product recovery via pertraction (membrane-based liquid–liquid extraction). We observed a maximum n-caproate productivity of 6.9 g COD/L-d for a period of 17 days at an l-lactate loading rate of 9.1 g COD/L-d, representing the highest sustained lactate to n-caproate conversion rate ever reported. We had to manage two competing lactate conversion pathways: 1) the reverse β-oxidation pathway to n-caproate; and 2) the acrylate pathway to propionate. We found that maintaining a low residual lactate concentration in the bioreactor broth was necessary to direct lactate conversion towards n-caproate instead of propionate. These findings provide a foundation for the development of new resource recovery processes to produce higher-value liquid products (e.g., n-caproate) from carbon-rich wastewaters containing lactate or lactate precursors (e.g., carbohydrates).

Impairment of exogenous lactate clearance in experimental hyperdynamic septic shock is not related to total liver hypoperfusion.

IntroductionAlthough the prognostic value of persistent hyperlactatemia in septic shock is unequivocal, its physiological determinants are controversial. Particularly, the role of impaired hepatic clearance has been underestimated and is only considered relevant in patients with liver ischemia or cirrhosis. Our objectives were to establish whether endotoxemia impairs whole body net lactate clearance, and to explore a potential role for total liver hypoperfusion during the early phase of septic shock.MethodsAfter anesthesia, 12 sheep were subjected to hemodynamic/perfusion monitoring including hepatic and portal catheterization, and a hepatic ultrasound flow probe. After stabilization (point A), sheep were alternatively assigned to lipopolysaccharide (LPS) (5 mcg/kg bolus followed by 4 mcg/kg/h) or sham for a three-hour study period. After 60 minutes of shock, animals were fluid resuscitated to normalize mean arterial pressure. Repeated series of measurements were performed immediately after fluid resuscitation (point B), and one (point C) and two hours later (point D). Monitoring included systemic and regional hemodynamics, blood gases and lactate measurements, and ex-vivo hepatic mitochondrial respiration at point D. Parallel exogenous lactate and sorbitol clearances were performed at points B and D. Both groups included an intravenous bolus followed by serial blood sampling to draw a curve using the least squares method.ResultsSignificant hyperlactatemia was already present in LPS as compared to sham animals at point B (4.7 (3.1 to 6.7) versus 1.8 (1.5 to 3.7) mmol/L), increasing to 10.2 (7.8 to 12.3) mmol/L at point D. A significant increase in portal and hepatic lactate levels in LPS animals was also observed. No within-group difference in hepatic DO2, VO2 or O2 extraction, total hepatic blood flow (point D: 915 (773 to 1,046) versus 655 (593 to 1,175) ml/min), mitochondrial respiration, liver enzymes or sorbitol clearance was found. However, there was a highly significant decrease in lactate clearance in LPS animals (point B: 46 (30 to 180) versus 1,212 (743 to 2,116) ml/min, P <0.01; point D: 113 (65 to 322) versus 944 (363 to 1,235) ml/min, P <0.01).ConclusionsEndotoxemia induces an early and severe impairment in lactate clearance that is not related to total liver hypoperfusion.Electronic supplementary materialThe online version of this article (doi:10.1186/s13054-015-0928-3) contains supplementary material, which is available to authorized users.

Lactate does not activate NF-κB in oxidative tumor cells.

The lactate anion is currently emerging as an oncometabolite. Lactate, produced and exported by glycolytic and glutaminolytic cells in tumors, can be recycled as an oxidative fuel by oxidative tumors cells. Independently of hypoxia, it can also activate transcription factor hypoxia-inducible factor-1 (HIF-1) in tumor and endothelial cells, promoting angiogenesis. These protumoral activities of lactate depend on lactate uptake, a process primarily facilitated by the inward, passive lactate-proton symporter monocarboxylate transporter 1 (MCT1); the conversion of lactate and NAD+ to pyruvate, NADH and H+ by lactate dehydrogenase-1 (LDH-1); and a competition between pyruvate and α-ketoglutarate that inhibits prolylhydroxylases (PHDs). Endothelial cells do not primarily use lactate as an oxidative fuel but, rather, as a signaling agent. In addition to HIF-1, lactate can indeed activate transcription factor nuclear factor-κB (NF-κB) in these cells, through a mechanism not only depending on PHD inhibition but also on NADH alimenting NAD(P)H oxidases to generate reactive oxygen species (ROS). While NF-κB activity in endothelial cells promotes angiogenesis, NF-κB activation in tumor cells is known to stimulate tumor progression by conferring resistance to apoptosis, stemness, pro-angiogenic and metastatic capabilities. In this study, we therefore tested whether exogenous lactate could activate NF-κB in oxidative tumor cells equipped for lactate signaling. We report that, precisely because they are oxidative, HeLa and SiHa human tumor cells do not activate NF-κB in response to lactate. Indeed, while lactate-derived pyruvate is well-known to inhibit PHDs in these cells, we found that NADH aliments oxidative phosphorylation (OXPHOS) in mitochondria rather than NAD(P)H oxidases in the cytosol. These data were confirmed using oxidative human Cal27 and MCF7 tumor cells. This new information positions the malate-aspartate shuttle as a key player in the oxidative metabolism of lactate: similar to glycolysis that aliments OXPHOS with pyruvate produced by pyruvate kinase and NADH produced by glyceraldehyde-3-phosphate dehydrogenase (GAPDH), oxidative lactate metabolism aliments OXPHOS in oxidative tumor cells with pyruvate and NADH produced by LDH1.

Dexmedetomidine ameliorates gut lactate production and impairment of exogenous lactate clearance in an endotoxic sheep model

The mechanisms of persistent hyperlactemia during endotoxic shock are probably multifactorial. Both hypoperfusion-related anaerobic production and adrenergic-driven aerobic generation have been implicated. More recently an early and severe impairment in exogenous lactate clearance has also been described [1]. Theoretically, an excessive adrenergic response could influence all these mechanisms and thus aggravate the problem.

Lactate and glucose concomitant consumption as a self-regulated pH detoxification mechanism in HEK293 cell cultures.

One of the most important limitations of mammalian cell-based processes is the secretion and accumulation of lactate as a by-product of their metabolism. Among the cell lines commonly used in industrial bioprocesses, HEK293 has been gaining importance over the last years. Up recently, HEK293 cells were known to consume lactate in late stages of cell culture usually when glucose and/or glutamine were depleted from media. Remarkably, in both scenarios, no significant cell growth was reported. However, we have observed a different metabolic behavior regarding lactate production and consumption in HEK293 cultures. HEK293 cells were able to co-metabolize glucose and lactate simultaneously, even in exponentially growing cell cultures. Our deep study of the effects of environmental conditions on lactate metabolism revealed that pH was the key to trigger the metabolic shift from lactate production to lactate and glucose concomitant consumption. Remarkably, this shift could be triggered at will when pH was set at 6.8. Even more interesting was the fact that lowering pH to 6.6 and supplementing media with exogenous lactate resulted in co-consumption of glucose and lactate from the beginning of cell culture, without affecting cell growth or protein productivity. On the contrary, cell growth was clearly hampered at this low pH if extracellular lactate was lacking. From our results, we hypothesize that HEK293 cells metabolize extracellular lactate as a strategy for pH detoxification, by means of co-transporting extracellular protons together with lactate into the cytosol. This novel hypothesis for unraveling lactate metabolism in HEK293 cells could open a door to re-direct genetic engineering strategies in order to obtain more efficient cell lines and also to further develop animal cell technology applications.

Ketone bodies effectively compete with glucose for neuronal acetyl-CoA generation in rat hippocampal slices.

Ketone bodies can be used for cerebral energy generation in situ, when their availability is increased as during fasting or ingestion of a ketogenic diet. However, it is not known how effectively ketone bodies compete with glucose, lactate, and pyruvate for energy generation in the brain parenchyma. Hence, the contributions of exogenous 5.0 mM [1-(13)C]glucose and 1.0 mM [2-(13)C]lactate + 0.1 mM pyruvate (combined [2-(13)C]lactate + [2-(13)C]pyruvate) to acetyl-CoA production were measured both without and with 5.0 mM [U-(13)C]3-hydroxybutyrate in superfused rat hippocampal slices by (13)C NMR non-steady-state isotopomer analysis of tissue glutamate and GABA. Without [U-(13)C]3-hydroxybutyrate, glucose, combined lactate + pyruvate, and unlabeled endogenous sources contributed (mean ± SEM) 70 ± 7%, 10 ± 2%, and 20 ± 8% of acetyl-CoA, respectively. With [U-(13)C]3-hydroxybutyrate, glucose contributions significantly fell from 70 ± 7% to 21 ± 3% (p < 0.0001), combined lactate + pyruvate and endogenous contributions were unchanged, and [U-(13)C]3-hydroxybutyrate became the major acetyl-CoA contributor (68 ± 3%)--about three-times higher than glucose. A direct analysis of the GABA carbon 2 multiplet revealed that [U-(13)C]3-hydroxybutyrate contributed approximately the same acetyl-CoA fraction as glucose, indicating that it was less avidly oxidized by GABAergic than glutamatergic neurons. The appearance of superfusate lactate derived from glycolysis of [1-(13)C]glucose did not decrease significantly in the presence of 3-hydroxybutyrate, hence total glycolytic flux (Krebs cycle inflow + exogenous lactate formation) was attenuated by 3-hydroxybutyrate. This indicates that, under these conditions, 3-hydroxybutyrate inhibited glycolytic flux upstream of pyruvate kinase.

Effect of fipronil on energy metabolism in the perfused rat liver

Fipronil is an insecticide used to control pests in animals and plants that can causes hepatotoxicity in animals and humans, and it is hepatically metabolized to fipronil sulfone by cytochrome P-450. The present study aimed to characterize the effects of fipronil (10–50μM) on energy metabolism in isolated perfused rat livers. In fed animals, there was increased glucose and lactate release from glycogen catabolism, indicating the stimulation of glycogenolysis and glycolysis. In the livers of fasted animals, fipronil inhibited glucose and urea production from exogenous l-alanine, whereas ammonia and lactate production were increased. In addition, fipronil at 50μM concentration inhibited the oxygen uptake and increased the cytosolic NADH/NAD+ ratio under glycolytic conditions. The metabolic alterations were found both in livers from normal or proadifen-pretreated rats revealing that fipronil and its reactive metabolites contributed for the observed activity. The effects on oxygen uptake indicated that the possible mechanism of toxicity of fipronil involves impairment on mitochondrial respiratory activity, and therefore, interference with energy metabolism. The inhibitory effects on oxygen uptake observed at the highest concentration of 50μM was abolished by pretreatment of the rats with proadifen indicating that the metabolites of fipronil, including fipronil sulfone, acted predominantly as inhibitors of respiratory chain. The hepatoxicity of both the parent compound and its reactive metabolites was corroborated by the increase in the activity of lactate dehydrogenase in the effluent perfusate in livers from normal or proadifen-pretreated rats.