Lactate Release Research Articles

Salbutamol Increases Leg Glucose Uptake and Metabolic Rate but not Muscle Glycogen Resynthesis in Recovery From Exercise.

Exercise blunts the effect of beta2-agonists on peripheral glucose uptake and energy expenditure. Whether such attenuation extends into recovery is unknown. To examine the effect of a beta2-agonist on leg glucose uptake and metabolic rate in recovery from exercise. Using leg arteriovenous balance technique and analyses of thigh muscle biopsies, we investigated the effect of a beta2-agonist (24 mg of oral salbutamol) vs placebo on leg glucose, lactate, and oxygen exchange before and during quadriceps exercise, and 0.5 to 5 hours in recovery from quadriceps exercise, as well as on muscle glycogen resynthesis and activity in recovery. Twelve healthy, lean, young men participated. Before exercise, leg glucose uptake was 0.42 ± 0.12 and 0.20 ± 0.02 mmol × min-1 (mean ± SD) for salbutamol and placebo (P = .06), respectively, while leg oxygen consumption was around 2-fold higher (P < .01) for salbutamol than for placebo (25 ± 3 vs 14 ± 1 mL × min-1). No treatment differences were observed in leg glucose uptake, lactate release, and oxygen consumption during exercise. But in recovery, cumulated leg glucose uptake, lactate release, and oxygen consumption was 21 mmol (95% CI 18-24, P = .018), 19 mmol (95% CI 16-23, P < .01), and 1.8 L (95% CI 1.6-2.0, P < .01) higher for salbutamol than for placebo, respectively. Muscle glycogen content was around 30% lower (P < .01) for salbutamol than for placebo in recovery, whereas no treatment differences were observed in muscle glycogen resynthesis or glycogen synthase activity. Exercise blunts the effect of beta2-agonist salbutamol on leg glucose uptake, but this attenuation diminishes in recovery. Salbutamol increases leg lactate release in recovery, which may relate to glycolytic trafficking due to excessive myocellular glucose uptake.

Macrosphelide A Exhibits a Specific Anti-Cancer Effect by Simultaneously Inactivating ENO1, ALDOA, and FH.

Aerobic glycolysis in cancer cells, also known as the Warburg effect, is an indispensable hallmark of cancer. This metabolic adaptation of cancer cells makes them remarkably different from normal cells; thus, inhibiting aerobic glycolysis is an attractive strategy to specifically target tumor cells while sparing normal cells. Macrosphelide A (MSPA), an organic small molecule, is a potential lead compound for the design of anti-cancer drugs. However, its role in modulating cancer metabolism remains poorly understood. MSPA target proteins were screened using mass spectrometry proteomics combined with affinity chromatography. Direct and specific interactions of MSPA with its candidate target proteins were confirmed by in vitro binding assays, competition assays, and simulation modeling. The siRNA-based knockdown of MSPA target proteins indirectly confirmed the cytotoxic effect of MSPA in HepG2 and MCF-7 cancer cells. In addition, we showed that MSPA treatment in the HEPG2 cell line significantly reduced glucose consumption and lactate release. MSPA also inhibited cancer cell proliferation and induced apoptosis by inhibiting critical enzymes involved in the Warburg effect: aldolase A (ALDOA), enolase 1 (ENO1), and fumarate hydratase (FH). Among these enzymes, the purified ENO1 inhibitory potency of MSPA was further confirmed to demonstrate the direct inhibition of enzyme activity to exclude indirect/secondary factors. In summary, MSPA exhibits anti-cancer effects by simultaneously targeting ENO1, ALDOA, and FH.

LINC00035 Transcriptional Regulation of SLC16A3 via CEBPB Affects Glycolysis and Cell Apoptosis in Ovarian Cancer.

Objective Ovarian cancer (OC) represents the most lethal gynecologic malignancy globally. Over the decades, lncRNAs have been considered as study focuses due to their genome-wide expression through multiple mechanisms in which regulation of target gene transcription through interaction with transcription factors or epigenetic proteins is proven. In the present work, we focus on the functional role of LINC00035 in OC and its regulation mechanism on gene expression. Methods We collected OC tissues and adjacent tumor-free tissues surgically resected from 67 OC patients. Cultured human OC cell lines SKOV3 and A2780 were assayed for their viability, migration, invasion, apoptosis in vitro using CCK-8 assays, transwell assays, and flow cytometric analysis. OC cell tumorigenesis in vivo was evaluated by mouse xenograft experiments. Glycolysis was evaluated by glucose uptake, lactate release, and ATP production assays. Luciferase activity assay, RNA immunoprecipitation (RIP), and RNA pull-down were performed to confirm the interactions among LINC00035, CEBPB, and SLC16A3. Results LINC00035 was upregulated in OC tissues. LINC00035 knockdown was shown to repress SKOV3 and A2780 cell viability, migration, invasion, induce their apoptosis, and reduce glucose uptake, lactate release, and ATP production. LINC00035 could recruit CEBPB into the SLC16A3 promoter region, thus increasing the SLC16A3 transcription. SLC16A3 was upregulated in OC tissues. SLC16A3 knockdown exerted similar effects on SKOV3 and A2780 cells as LINC00035 knockdown. Rescue experiments found SLC16A3 overexpression resisting to LINC00035 knockdown on SKOV3 and A2780 cell viability, migration, invasion, apoptosis, glucose uptake, lactate release, and ATP production. Results also showed LINC00035 knockdown could inhibit OC cell tumorigenesis in vivo. Conclusion The study reveals that LINC00035 promotes OC progression by regulating glycolysis and cell apoptosis through CEBPB-mediated transcriptional promotion of SLC16A3.

Zhoushi Qi Ling decoction represses docetaxel resistance and glycolysis of castration-resistant prostate cancer via regulation of SNHG10/miR-1271-5p/TRIM66 axis

Docetaxel resistance developed in half of castration-resistant prostate cancer (CRPC) patients hinders its long-term clinical application. The current study was designed to investigate the effects of Chinese medicine Zhoushi Qi Ling decoction on the docetaxel resistance of prostate cancer as well as elucidate the underlying molecular mechanism. In our study, Qi Ling significantly decreased viability and colony formation as well as increased apoptosis of docetaxel-resistant (DR) CRPC cells. Qi Ling-treated DR cells exhibited decreased glucose consumption, lactate release and pyruvate production. Moreover, lncRNA SNHG10 was upregulated in DR tissues of CRPC patients and was negatively correlated with the progression-free survival. Bioinformatics analysis indicated miR-1271-5p as the associated miRNA possibly binding with SNHG10. miR-1271-5p up-regulation dramatically decreased the luciferase activity of SNHG10 in DR cells. SNHG10 knockdown sharply increased the expression of miR1271-5p in DR cells. Targetscan predicted TRIM66 as one of the downstream targets of miR-1271-5p. miR-1271-5p up-regulation drastically reduced luciferase activity as well as TRIM66 expression in DR cells. Also, the knockdown of SNHG10 remarkably suppressed the expression of TRIM66 in DR cells. Additionally, Qi Ling treatment reduced SNHG10 and TRIM66, while increased miR1271-5p, in DR cells. In summary, Qi Ling inhibited docetaxel resistance and glycolysis of CRPC possibly via SNHG10/miR-1271-5p/TRIM66 pathway.

Exercise-Induced Lactate Release Mediates Mitochondrial Biogenesis in the Hippocampus of Mice via Monocarboxylate Transporters.

Regular exercise training induces mitochondrial biogenesis in the brain via activation of peroxisome proliferator-activated receptor gamma-coactivator 1α (PGC-1α). However, it remains unclear whether a single bout of exercise would increase mitochondrial biogenesis in the brain. Therefore, we first investigated whether mitochondrial biogenesis in the hippocampus is affected by a single bout of exercise in mice. A single bout of high-intensity exercise, but not low- or moderate-intensity, increased hippocampal PGC-1α mRNA and mitochondrial DNA (mtDNA) copy number at 12 and 48h. These results depended on exercise intensity, and blood lactate levels observed immediately after exercise. As lactate induces mitochondrial biogenesis in the brain, we examined the effects of acute lactate administration on blood and hippocampal extracellular lactate concentration by in vivo microdialysis. Intraperitoneal (I.P.) lactate injection increased hippocampal extracellular lactate concentration to the same as blood lactate level, promoting PGC-1α mRNA expression in the hippocampus. However, this was suppressed by administering UK5099, a lactate transporter inhibitor, before lactate injection. I.P. UK5099 administration did not affect running performance and blood lactate concentration immediately after exercise but attenuated exercise-induced hippocampal PGC-1α mRNA and mtDNA copy number. In addition, hippocampal monocarboxylate transporters (MCT)1, MCT2, and brain-derived neurotrophic factor (BDNF) mRNA expression, except MCT4, also increased after high-intensity exercise, which was abolished by UK5099 administration. Further, injection of 1,4-dideoxy-1,4-imino-D-arabinitol (glycogen phosphorylase inhibitor) into the hippocampus before high-intensity exercise suppressed glycogen consumption during exercise, but hippocampal lactate, PGC-1α, MCT1, and MCT2 mRNA concentrations were not altered after exercise. These results indicate that the increased blood lactate released from skeletal muscle may induce hippocampal mitochondrial biogenesis and BDNF expression by inducing MCT expression in mice, especially during short-term high-intensity exercise. Thus, a single bout of exercise above the lactate threshold could provide an effective strategy for increasing mitochondrial biogenesis in the hippocampus.

The thermogenic activity of adjacent adipocytes fuels the progression of ccRCC and compromises anti-tumor therapeutic efficacy

Clear cell renal cell carcinoma (ccRCC) preferentially invades into perinephric adipose tissue (PAT), a process associated with poor prognosis. However, the detailed mechanisms underlying this interaction remain elusive. Here, we describe a bi-directional communication between ccRCC cells and the PAT. We found that ccRCC cells secrete parathyroid-hormone-related protein (PTHrP) to promote the browning of PAT by PKA activation, while PAT-mediated thermogenesis results in the release of excess lactate to enhance ccRCC growth, invasion, and metastasis. Further, tyrosine kinase inhibitors (TKIs) extensively used in the treatment of ccRCC enhanced this vicious cycle of ccRCC-PAT communication by promoting the browning of PAT. However, if this cross-communication was short circuited by the pharmacological suppression of adipocyte browning via H89 or KT5720, the anti-tumor efficacy of the TKI, sunitinib, was enhanced. These results suggest that ccRCC-PAT cross-communication has important clinical relevance, and use of combined therapy holds great promise in enhancing the efficacy of TKIs.

Aerobic exercise improves LPS-induced sepsis via regulating the Warburg effect in mice

We investigated the impact of aerobic exercise (AE) on multiple organ dysfunction syndrome (MODS), aortic injury, pathoglycemia, and death during sepsis. ICR mice were randomized into four groups: Control (Con), Lipopolysaccharide (LPS), Exercise (Ex), and Exercise + LPS (Ex + LPS) groups. Mice were trained with low-intensity for 4 weeks. LPS and Ex + LPS mice received 5 mg/kg LPS intraperitoneally for induction of sepsis. Histopathological micrographs showed the organ morphology and damage. This study examined the effects of AE on LPS-induced changes in systemic inflammation, pulmonary inflammation, lung permeability, and bronchoalveolar lavage fluid (BALF) cell count, oxidative stress-related indicators in the lung, blood glucose levels, plasma lactate levels, serum insulin levels, plasma high-mobility group box 1 (HMGB1) levels, glucose transporter 1 (Glut1) and HMGB1, silent information regulator 1 (Sirt-1), and nuclear factor erythroid 2-related factor 2 (Nrf-2) mRNA expression levels in lung tissue. AE improved sepsis-associated multiple organ dysfunction syndrome (MODS), aortic injury, hypoglycemia, and death. AE prominently decreased pulmonary inflammation, pulmonary edema, and modulated redox balance during sepsis. AE prominently decreased neutrophil content in organ. AE prominently downregulated CXCL-1, CXCL-8, IL-6, TNF-α, Glu1, and HMGB1 mRNA expression but activated IL-1RN, IL-10, Sirt-1, and Nrf-2 mRNA expression in the lung during sepsis. AE decreased the serum levels of lactate and HMGB1 but increased blood glucose levels and serum insulin levels during sepsis. A 4-week AE improves sepsis-associated MODS, aortic injury, pathoglycemia, and death. AE impairs LPS-induced lactate and HMGB1 release partly because AE increases serum insulin levels and decreases the levels of Glut1. AE is a novel therapeutic strategy for sepsis targeting aerobic glycolysis.

Hemin attenuates response of primary rat adipocytes to adrenergic stimulation.

Hemin is an activator of heme oxygenase-1 (HO-1), an enzyme catalyzing heme degradation. Up-regulation of HO-1 is observed in response to various pathological conditions. Moreover, pharmacological activation of HO-1 is associated with numerous beneficial effects in the organism. Hemin was shown to exert, among other, anti-diabetic and anti-obesity properties. These effects are strongly linked with adipose tissue. However, the direct influence of hemin on metabolism of the fat cells have not been explored. The present study aimed to determine the short-term effects of hemin on metabolism of the primary rat adipocytes. We focused on processes directly related to lipid accumulation, such as lipogenesis and lipolysis. For this purpose, the isolated cells were subjected for 2 h to 40 µM hemin, and effects of this compound on insulin-stimulated glucose conversion to lipids, lactate release, lipolysis induced by various stimuli, and also on the antilipolytic action of insulin were determined. It was shown that hemin did not affect insulin-induced lipogenesis and lactate release. However, hemin significantly decreased lipolysis stimulated by epinephrine. The inhibitory effect of hemin on epinephrine-induced lipolysis was not abolished in the presence of SnMP, an inhibitor of HO-1, which suggests hemin action irrespective of this enzyme. Similar inhibitory effects on epinephrine-induced lipolysis were observed in the presence of 3 and 12 mM glucose. Moreover, hemin was shown to reduce epinephrine-induced lipolysis also when glucose was replaced by alanine or by succinate. Apart from changes in epinephrine action, it was found that the lipolytic response of the adipocytes to isoproterenol was also diminished by hemin. However, hemin failed to affect lipolysis stimulated by dibutyryl-cAMP (a direct activator of protein kinase A), forskolin (an activator of adenylate cyclase), and also by DPCPX (an adenosine A1 receptor antagonist). Additionally, epinephrine-induced lipolysis was shown to be decreased by insulin, and this effect was deepened in the presence of hemin. These results indicate that short-term exposure of the adipocytes to hemin does not affect processes related to glucose metabolism, such as lipogenesis and lactate release. However, hemin was found to decrease the lipolytic response to adrenergic stimulation, which is associated with reduced lipid release from adipocytes. Moreover, our results indicate that hemin is also capable of diminishing the exaggerated lipolysis, which occurs in the presence of supraphysiological concentrations of glucose.

Astrocyte-neuron lactate transport in the ACC contributes to the occurrence of long-lasting inflammatory pain in male mice

Lactate transport is an important means of communication between astrocytes and neurons and is implicated in a variety of neurobiological processes. However, the connection between astrocyte-neuron lactate transport and nociceptive modulation has not been well established. Here, we found that Complete Freund’s adjuvant (CFA)-induced inflammation pain leads to a significant increase in extracellular lactate levels in the anterior cingulate cortex (ACC). Inhibition of glycogenolysis and lactate release in the ACC disrupted the persistent, but not acute, inflammation pain induced by CFA, and this effect was reversed by exogenous L-lactate administration. Knocking down the expression of lactate transporters (MCT1, MCT4, or MCT2) also disrupted the long lasting inflammation pain induced by CFA. Moreover, glycogenolysis in the ACC is critical for the induction of molecular changes related to neuronal plasticity, including the induction of phospho- (p-) ERK, p-CREB, and Fos. Taken together, our findings indicate that astrocyte-neuron lactate transport in the ACC is critical for the occurrence of persistent inflammation pain, suggesting a novel mechanism underlying chronic pain.

Impact of Inhibition of the Mitochondrial Pyruvate Carrier on the Tumor Extracellular pH as Measured by CEST-MRI.

Simple SummaryDysregulated metabolism is a key hallmark of cancer cells and many solid tumors are acidic. Acidosis is responsible for cancer aggressiveness and for resistance to several treatments. In the present study, we evaluated to which extent tumor acidosis was influenced upon inhibition of the import of pyruvate into the mitochondria, the powerhouse of the cell. Using advanced molecular imaging to measure non-invasively the acidity in a model of breast cancer, we found that while some tumor regions became much more acidic, others did not show any change. This study highlights the capacity of this advanced technology to reveal the heterogeneity of response to the treatment.(1) Background: The acidosis of the tumor micro-environment may have profound impact on cancer progression and on the efficacy of treatments. In the present study, we evaluated the impact of a treatment with UK-5099, a mitochondrial pyruvate carrier (MPC) inhibitor on tumor extracellular pH (pHe); (2) Methods: glucose consumption, lactate secretion and extracellular acidification rate (ECAR) were measured in vitro after exposure of cervix cancer SiHa cells and breast cancer 4T1 cells to UK-5099 (10 µM). Mice bearing the 4T1 tumor model were treated daily during four days with UK-5099 (3 mg/kg). The pHe was evaluated in vivo using either chemical exchange saturation transfer (CEST)-MRI with iopamidol as pHe reporter probe or 31P-NMR spectroscopy with 3-aminopropylphosphonate (3-APP). MR protocols were applied before and after 4 days of treatment; (3) Results: glucose consumption, lactate release and ECAR were increased in both cell lines after UK-5099 exposure. CEST-MRI showed a significant decrease in tumor pHe of 0.22 units in UK-5099-treated mice while there was no change over time for mice treated with the vehicle. Parametric images showed a large heterogeneity in response with 16% of voxels shifting to pHe values under 7.0. In contrast, 31P-NMR spectroscopy was unable to detect any significant variation in pHe; (4) Conclusions: MPC inhibition led to a moderate acidification of the extracellular medium in vivo. CEST-MRI provided high resolution parametric images (0.44 µL/voxel) of pHe highlighting the heterogeneity of response within the tumor when exposed to UK-5099.

Single Impact Injury of Vertebral Endplates Without Structural Disruption, Initiates Disc Degeneration Through Piezo1 Mediated Inflammation and Metabolism Dysfunction.

In vitro experimental study. To establish an axial impact injury model of intervertebral disc (IVD) and to investigate if a single impact injury without endplate structural disruption could initiate intervertebral disc degeneration (IDD), and what is the roles of Piezo1 in this process. Although IDD process has been confirmed to be associated with structural failures such as endplate fractures, whether a single impact injury of the endplates without structural disruption could initiate IDD remains controversial. Previous studies reported that Piezo1 mediated inflammation participated in the progression of IDD induced by mechanical stretch; however, the roles of Piezo1 in IVD impact injury remain unknown. Rats spinal segments were randomly assigned into Control, Low, and High Impact groups, which were subjected to pure axial impact loading using a custom-made apparatus, and cultured for 14 days. The degenerative process was investigated by using histomorphology, real-time Polymerase Chain Reaction(PCR), western-blot, immunofluorescence, and energy metabolism of IVD cell. The effects of Piezo1 were investigated by using siRNA transfection, real-time PCR, western-blot, and immunofluorescence. The discs in both of the impact groups presented degenerative changes after 14 days, which showed significant up-regulation of Piezo1, NLRP3 inflammasome, the catabolic (MMP-9, MMP-13), and pro-inflammatory gene (IL-1β) expression than that of the control group (P < 0.05), accompanied by significantly increased release of ATP, lactate, nitric oxide (NO), and glucose consumption of IVD cells at first 7 days. Silencing Piezo1 reduced the activation of NLRP3 inflammasome and IL-1β expression in the nucleus pulposus induced by impact injury. It demonstrated that not only fracture of the endplate but also a single impact injury without structural impairment could also initiate IDD, which might be mediated by activation of Piezo1 induced inflammation and abnormal energy metabolism of IVD cells.Level of Evidence: N/A.

Microglia-Derived Small Extracellular Vesicles Reduce Glioma Growth by Modifying Tumor Cell Metabolism and Enhancing Glutamate Clearance through miR-124.

Brain homeostasis needs continuous exchange of intercellular information among neurons, glial cells, and immune cells, namely microglial cells. Extracellular vesicles (EVs) are active players of this process. All the cells of the body, including the brain, release at least two subtypes of EVs, the medium/large EVs (m/lEVs) and small EVs (sEVs). sEVs released by microglia play an important role in brain patrolling in physio-pathological processes. One of the most common and malignant forms of brain cancer is glioblastoma. Altered intercellular communications constitute a base for the onset and the development of the disease. In this work, we used microglia-derived sEVs to assay their effects in vitro on murine glioma cells and in vivo in a glioma model on C57BL6/N mice. Our findings indicated that sEVs carry messages to cancer cells that modify glioma cell metabolism, reducing lactate, nitric oxide (NO), and glutamate (Glu) release. sEVs affect Glu homeostasis, increasing the expression of Glu transporter Glt-1 on astrocytes. We demonstrated that these effects are mediated by miR-124 contained in microglia-released sEVs. The in vivo benefit of microglia-derived sEVs results in a significantly reduced tumor mass and an increased survival of glioma-bearing mice, depending on miR-124.

The combinatorial inhibition of Wnt signaling and Akt kinase is beneficial for reducing the survival and glycolytic activity of tongue cancer cells.

Wnt signaling is important in the development of head and neck squamous cell carcinomas (HNSCC); however, Wnt pathway inhibitors lack satisfactory potency when used in monotherapy. The aim of this study was to assess the effects of the combinations of Wnt-signaling inhibitors and the inhibitor of Akt kinase on the survival and glycolytic activity of tongue carcinoma cells. CAL27, SCC-25, and BICR22 tongue cancer cell lines were used. Cells were treated with Wnt signaling (PRI-724 and IWP-O1) and Akt-kinase inhibitors. The effect of the chemicals on cell viability and cytotoxicity were evaluated by MTS and CellTox Green assays, respectively. Cell cycle distribution was analyzed cytometrically after propidium iodide staining. Annexin V binding to externalized phosphatidylserine and analysis of mitochondrial potential allowed the assessment of apoptosis. Glucose uptake and lactate release were evaluated luminometrically. Additionally, the viability of cells in spheroids was analyzed based on ATP content. The Akt-kinase inhibitor showed significant cytotoxicity toward primary cancer cells. Moreover, its pro-apoptotic effects were enhanced by Wnt-pathway inhibitors. The activity of Akt inhibitor was even higher (by twofold) in 3D spheroids in comparison to cells grown in monolayer. The synergistic reduction in the growth of spheroids was observed between Akt inhibitor and IWP-O1. Reduced glucose consumption may play a part in the combinatorial effects of these chemicals. The results point to the therapeutic potential of the combinatorial use of Wnt inhibitors together with Akt inhibitors in HNSCC.

Expression of Microbial Enzymes in Mammalian Astrocytes to Modulate Lactate Release.

Astrocytes support and modulate neuronal activity through the release of L-lactate. The suggested roles of astrocytic lactate in the brain encompass an expanding range of vital functions, including central control of respiration and cardiovascular performance, learning, memory, executive behaviour and regulation of mood. Studying the effects of astrocytic lactate requires tools that limit the release of lactate selectively from astrocytes. Here, we report the validation in vitro of novel molecular constructs derived from enzymes originally found in bacteria, that when expressed in astrocytes, interfere with lactate handling. When lactate 2-monooxygenase derived from M. smegmatis was specifically expressed in astrocytes, it reduced intracellular lactate pools as well as lactate release upon stimulation. D-lactate dehydrogenase derived from L. bulgaricus diverts pyruvate towards D-lactate production and release by astrocytes, which may affect signalling properties of lactate in the brain. Together with lactate oxidase, which we have previously described, this set of transgenic tools can be employed to better understand astrocytic lactate release and its role in the regulation of neuronal activity in different behavioural contexts.

Gingival-Derived Mesenchymal Stem Cells Protect Against Sepsis and Its Complications.

ObjectiveIn the present study, we separated and characterized mouse gingival-derived mesenchymal stem cells (GMSCs) and investigated whether GMSCs can improve lipopolysaccharide (LPS)-induced sepsis and its complications.MethodsNinety-six ICR mice were randomly divided into the following groups: the control (Sham), LPS, and LPS + MSC groups. Mice received 5 mg/kg LPS intraperitoneally to induce sepsis. Histopathological micrographs illustrated organ injury. We detected systemic inflammation, blood glucose levels, and serum levels of high-mobility group box 1 (HMGB1) and lactate. In addition, pulmonary inflammation, lung permeability, and oxidative stress-related indicators in lung tissue were measured.ResultsWe successfully separated a novel population of MSCs from mouse gingiva. These cells had MSC-associated properties, such as a typical fibroblast-like morphology, multiple differentiation potential, and certain phenotypes. Cell-based therapy using GMSCs significantly improved the survival rate, systemic inflammation, hypoglycemia, multiple organ dysfunction syndrome (MODS), and aortic injury during sepsis. GMSCs administration reduced pulmonary inflammation, lung permeability, and oxidative stress injury. GMSCs administration reduced neutrophil infiltration partly because GMSCs inhibited neutrophil chemoattractants tumor necrosis factor (TNF-α), C-X-C motif chemokine ligand (CXCL-1), and Interleukin (IL-8). GMSCs impaired LPS-induced HMGB1 and lactate release during sepsis.ConclusionGMSCs administration is a novel therapeutic strategy targeting aerobic glycolysis for the treatment of sepsis because GMSCs impair LPS-induced HMGB1 and lactate release. GMSCs alleviate lung injury partly because GMSCs exert immune effects, inhibit neutrophilic inflammation, and reduce oxidative stress injury.

Stressed axons craving for glial sugar: links to regeneration?

Stressed axons craving for glial sugar: links to regeneration?

Abstract 1974: C3G down-regulation in glioblastoma induces a pro-invasive and glycolytic phenotype, accompanied by RTKs dysregulation

Abstract C3G is a guanine-nucleotide exchange factor (GEF) for Rap1, although it can act through GEF-independent mechanisms. C3G plays a dual role in cancer, acting as either a tumor suppressor or inducer depending on the tumor type/stage. It regulates different key aspects of the tumorigenic process such as invasion, apoptosis or proliferation. In colon carcinoma, C3G represses invasion through down-regulation of p38αMAPK activity and promotes tumor growth. We have now analyzed the role of C3G in glioblastoma (GBM), a tumor characterized by its aggressiveness and disseminative capacity. To do it, we have used different experimental approaches: permanent gene silencing, knock-out using CRISPR/Cas9 technology and transient overexpression in U87 cell line and patient-derived GBM cells. We found that C3G down-regulation enhances invasion through the induction of an epithelial/glial to mesenchymal transition-like process. C3G deficiency also facilitates foci generation in anchorage-dependent and independent growth assays, but with lower cell density as a consequence of a reduced proliferation. In in vivo analyses, cells with C3G knock-down generated larger tumors in xenografts and chick chorioallantoic membrane xenografts assays with increased angiogenesis and α-SMA+ fibroblasts, but a lower proliferation. Mechanistically, C3G down-regulation impairs EGF/EGFR signaling by decreasing EGFR cell membrane localization, leading to a reduction in EGF/EGFR-induced invasiveness. In contrast, C3G down-regulation promotes the activation of several receptor tyrosine kinases (RTKs) that might promote invasion. In particular, the enhanced FGF2/FGFR1 signaling increases invasion in C3G-silenced cells, through a mechanism likely dependent on ERKs. Moreover, a proteomic analysis revealed that C3G down-regulation increases the levels of different key glycolytic enzymes, such as aldolase, phosphoglycerate kinase, enolase and pyruvate kinase (PK). The activity of PK and lactate dehydrogenase as well as lactate release to the extracellular environment were also increased upon C3G silencing. In conclusion, our data demonstrate that C3G inhibits invasion of GBM cells, while increasing their proliferative capacity. Moreover, we show a distinct dependency on C3G for EGF/EGFR signaling versus other RTKs, suggesting that assessing C3G levels may discriminate GBM patient responders to different RTK inhibitors. In addition, our results indicate that C3G regulates not only GBM growth and invasiveness, but it also contributes to reprogram its glycolytic metabolism. Hence, low levels of C3G correlate with a more mesenchymal and glycolytic phenotype with enhanced aggressiveness and worse patient prognosis. This promising role of C3G as a key player in GBM should be further characterized to define its prognostic value and its potential relevance as a predictor of the response to therapy. Citation Format: Sara Manzano, Óscar Herranz, Paloma Bragado, Patricia Jáuregui, María Rodrigo, Celia Sequera, Cristina Baquero, Nerea Palao, Ignacio Rubio, Álvaro Gutierrez-Uzquiza, Carmen Guerrero, Almudena Porras. C3G down-regulation in glioblastoma induces a pro-invasive and glycolytic phenotype, accompanied by RTKs dysregulation [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1974.

Lactate Release By Bone Marrow Neutrophils Promotes Their Inflammatory Mobilization Via Endothelial GPR81 Signaling

Neutrophils are best known as glycolytic cells and an essential first host defense line of the innate immune system during inflammation. Activation of pro-inflammatory signaling by immune cells accelerates metabolic pathways, leading to increased lactate production. While lactate levels are the best clinical marker for sepsis diagnosis, the mechanisms underlying lactate production by bone marrow (BM) neutrophils and its role during inflammation remain poorly understood. Here, we report that LPS exposure in mice (mimicking bacterial inflammation) activated BM neutrophils leading to their mobilization to peripheral blood (PB) and recruitment to the liver. This happened concomitantly with an increase in BM neutrophil metabolism, reflected by high glucose uptake, high generation of reactive oxygen species (ROS) and increase in HIF-1α expression. Importantly, we identified that BM neutrophils highly express lactate dehydrogenase A (LDHA), an enzyme that converts pyruvate to lactate, as well as the lactate efflux transporter MCT4. On the other hand BM neutrophils express low levels of the lactate influx transporter MCT1 and the lactate receptor GPR81. In addition, we found that BM neutrophil depletion resulted in lower levels of BM derived lactate in mice exposed to LPS. Moreover, LPS administration up-regulated the expression of LDHA and MCT4 on BM neutrophils. Taken together, these results reveal that during acute LPS induced inflammation BM neutrophils produce and release high levels of lactate. Unexpectedly, we found that in vivo lactate treatment specifically induced rapid ROShigh neutrophil activation and mobilization from the BM to PB and liver in an NADPH oxidase/ROS dependent manner. Accordingly, reduced activated neutrophil levels in PB and liver were observed following inhibition of ROS production. Furthermore, in a genetic mouse model of Chronic Granulomatous Disease (CGD) due to NADPH oxidase inactivation, with reduced ROS generation, treatment with lactate partially reduced neutrophil mobilization from the BM. Our results demonstrate that NADPH oxidase/ROS signaling play a role in lactate-induced neutrophil mobilization. HIF-1α is a major regulator of energy metabolism and glycolysis pathway in immune cells (Reviewed by Weidemann and Johnson, Cell Death and Diff. 2008, and by O'Neill and Pearce, J. Exp. Med. 2015). Notably, our results show in myeloid specific HIF-1α-deficient mice exposed to LPS a reduction in the expression of LDHA and MCT4 on BM neutrophils compared to wild type mice, suggesting that HIF-1α is upstream of lactate. Accordingly, exogenous lactate administration restored neutrophil mobilization in myeloid HIF-1α-deficient mice implicating HIF-1α in lactate production and release by BM neutrophils and in their mobilization from the BM following acute inflammation.Further deciphering the mechanisms of lactate-induced neutrophil mobilization, we identified that BM endothelial cells (BMEC) highly express the lactate receptor GPR81. This GPCR was downregulated and internalized by lactate, as well as GPR81 agonist or LPS treatment. Interestingly, we found that lactate or GPR81 agonist reduced BMEC VE-Cadherin expression, leading to increased blood-BM vascular permeability as a possible cause for the enhanced neutrophil mobilization from the BM into the circulation. Lactate effects on BM neutrophils required functional GPR81 signaling since lactate injection together with a GPR81 antagonist, or to GPR81-deficient mice interfered with BM vascular permeability and reduced neutrophil mobilization from the BM. Moreover, in chimeras of GPR81-deficient BM reconstituted in wild type mice recipients we found that lactate induced-neutrophil mobilization is dependent on host BMEC GPR81 signaling rather than hematopoietic cell GPR81. This finding eludes to a role of endothelial GPR81 rather than to neutrophil GPR81 in lactate induced-neutrophil mobilization. In summary, our study links, for the first time, the metabolic by-product lactate secreted by BM neutrophils exposed to inflammatory conditions, to a novel GPR81 signaling axis on the BM endothelial cells which controls vascular permeability and thereby neutrophil mobilization. Our study highlights lactate as a potential target for attenuating blood neutrophilia and neutrophil injury associated with sepsis and other infectious conditions. DisclosuresNo relevant conflicts of interest to declare.

Neuroprotective Function of High Glycolytic Activity in Astrocytes: Common Roles in Stroke and Neurodegenerative Diseases.

Astrocytes (also, astroglia) consume huge amounts of glucose and produce lactate regardless of sufficient oxygen availability, indicating a high capacity for aerobic glycolysis. Glycolysis in astrocytes is activated in accordance with neuronal excitation and leads to increases in the release of lactate from astrocytes. Although the fate of this lactate remains somewhat controversial, it is believed to fuel neurons as an energy substrate. Besides providing lactate, astrocytic glycolysis plays an important role in neuroprotection. Among the minor pathways of glucose metabolism, glucose flux to the pentose-phosphate pathway (PPP), a major shunt pathway of glycolysis, is attracting research interest. In fact, PPP activity in astrocytes is five to seven times higher than that in neurons. The astrocytic PPP plays a key role in protecting neurons against oxidative stress by providing neurons with a reduced form of glutathione, which is necessary to eliminate reactive oxygen species. Therefore, enhancing astrocytic glycolysis might promote neuronal protection during acute ischemic stroke. Contrariwise, the dysfunction of astrocytic glycolysis and the PPP have been implicated in the pathogenesis of various neurodegenerative diseases such as Parkinson’s disease, Alzheimer’s disease, and amyotrophic lateral sclerosis, since mitochondrial dysfunction and oxidative stress trigger and accelerate disease progression.

The role of lactate in sepsis and COVID-19: Perspective from contracting skeletal muscle metabolism.

New Findings What is the topic of this review? Lactate is considered an important substrate for mitochondria in the muscles, heart and brain during exercise and is the main gluconeogenetic precursor in the liver and kidneys. In this light, we review the (patho)physiology of lactate metabolism in sepsis and coronavirus disease 2019 (COVID‐19). What advances does it highlight? Elevated blood lactate is strongly associated with mortality in septic patients. Lactate seems unrelated to tissue hypoxia but is likely to reflect mitochondrial dysfunction and high adrenergic stimulation. Patients with severe COVID‐19 exhibit near‐normal blood lactate, indicating preserved mitochondrial function, despite a systemic hyperinflammatory state similar to sepsis. In critically ill patients, elevated plasma lactate is often interpreted as a sign of organ hypoperfusion and/or tissue hypoxia. This view on lactate is likely to have been influenced by the pioneering exercise physiologists around 1920. August Krogh identified an oxygen deficit at the onset of exercise that was later related to an oxygen ‘debt’ and lactate accumulation by A. V. Hill. Lactate is considered to be the main gluconeogenetic precursor in the liver and kidneys during submaximal exercise, but hepatic elimination is attenuated by splanchnic vasoconstriction during high‐intensity exercise, causing an exponential increase in blood lactate. With the development of stable isotope tracers, lactate has become established as an important energy source for muscle, brain and heart tissue, where it is used for mitochondrial respiration. Plasma lactate > 4 mM is strongly associated with mortality in septic shock, with no direct link between lactate release and tissue hypoxia. Herein, we provide evidence for mitochondrial dysfunction and adrenergic stimulation as explanations for the sepsis‐induced hyperlactataemia. Despite profound hypoxaemia and intense work of breathing, patients with severe coronavirus disease 2019 (COVID‐19) rarely exhibit hyperlactataemia (> 2.5 mM), while presenting a systemic hyperinflammatory state much like sepsis. However, lactate dehydrogenase, which controls the formation of lactate, is markedly elevated in plasma and strongly associated with mortality in severe COVID‐19. We briefly review the potential mechanisms of the lactate dehydrogenase elevation in COVID‐19 and its relationship to lactate metabolism based on mechanisms established in contracting skeletal muscle and the acute respiratory distress syndrome.