Rate Of Lactate Production Research Articles

Carbamazepine induces a bioenergetics disruption to microvascular endothelial cells from the blood-brain barrier

Carbamazepine (CBZ) is a widely employed anti-seizure medication that crosses the blood-brain barrier (BBB) to exert its anti-convulsant action. The effects of CBZ on components of the BBB have yet to be completely delineated. Hence the current study evaluated the effects of CBZ upon mitochondrial functionality of BBB-derived microvascular endothelial cells isolated from Albino rats. The influence of CBZ on cell viability and barrier functions were evaluated by 3-(4,5 dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide (MTT), lactate dehydrogenase, and electrophysiological assays over a drug concentration range of 0.1–1000 μM. Bioenergetics effects were measured via ATP production, mitochondrial complexes I and III activities, lactate production, and oxygen consumption rates (OCRs), and mitochondrial membrane potential, fluidity and lipid content. CBZ was cytotoxic to microvascular endothelial cells in a concentration and duration dependent manner. CBZ significantly diminished the endothelial cell’s barrier functions, and impacted upon cellular bioenergetics: reducing mitochondrial complex activities with a parallel decrease in OCRs and increased anaerobic lactate production. CBZ significantly decreased mitochondrial membrane potential and induced an increase of membrane fluidity and decrease in levels of mitochondrial saturated and unsaturated fatty acids. In summary, CBZ disrupted functional activity of BBB endothelial cells via damage and modification of mitochondria functionality at therapeutically relevant concentrations.

Parametric optimization and kinetic study of l-lactic acid production by homologous batch fermentation of Lactobacillus pentosus cells.

Parametric optimization always plays important roles in bioengineering systems to obtain a high product yield under the proper conditions. The parametric conditions of lactic acid production by homologous batch fermentation of Lactobacillus pentosus cells was optimized by the Box-Behnken design. The highest l-lactic acid yield was obtained as 0.836±0.003 g/g glucose with the productivity of 0.906±0.003g/(L×H) under the optimum conditions of 34.7 °C, pH 6.2, 148 rpm agitation speed, and 9.3g/L nitrogen source concentration determined by quadratic response surface with high accuracy. The adequate kinetic models of cell growth rate, lactic production rate, and glucose consumption rate were also established to describe the fermentation behavior of L. pentosus cells with the correlation coefficients of 09985, 0.9990, and 0.9989, respectively.

Methods For Estimating Anaerobic Glycolytic Capacity (vLa Max ) In Trained Cyclists

PURPOSE: The maximum rate of lactate production (vLaMax) can be used to estimate the maximum anaerobic energy contribution, which can be used to optimize endurance training. The aim of this study was to examine the reliability of a 20-sec vLaMax estimate using either a variable or standardized alactic time interval (talac). METHODS: 16 trained male cyclists and triathletes (35.3 + 8.9 yo, 1.80 + 0.05 m, and 73.3 + 11.2 kg, average 30-sec Wingate power of 9.0 +1.6 W.kg-1) were recruited. Subjects completed two familiarization trials on their own before completing two 20-sec maximal sprints on their own bikes which were attached to a Wahoo Kickr trainer. Each session began with a 10-min warm-up at 100 W before each sprint and were then followed by 12-min passive rest. A lactate (HLa) sample was taken from the finger at the end of the sprint and then every 2-min after that until HLa dropped > 1 mM. vLaMax was then calculated using (HLa post - pre)/(20-sec - alactic time), where talac was either time 0-sec to time where power drops 3.5%, or was set at 5-sec. RESULTS: Individual sprint sessions data were analyzed and no significant differences (p>0.05) nor were noted between session 1 and 2 data; all data were pooled and then vLaMax calculated using methods described above. Mean 20-sec sprint power was 9.8 + 1.0 W.kg-1 (~109% of Wingate) and a total work of 14.5 + 2.7. The talac was calculated to be 4.0 + 2.3-sec. There was no significant difference (p>0.05) between the calculated vLaMax 0.51 + 0.18 mM.L-1.sec-1 and vLaMax using the a standard 5-sec value, 0.54 + 0.15 mM.L-1.sec-1. Neither vLaMax value appears to be associated (r2 < 0.10, p>0.05) with absolute sprint performance. CONCLUSIONS: These results indicate that using a standard talac of 5-sec produced similar results as calculating it using a 3.5% drop in power. While vLaMax may ultimately influence HLa testing curves (i.e., lactate threshold) and endurance performance, it does not appear to be associated with actual sprint power output.

Lactic Acidosis in the Presence of Glucose Diminishes Warburg Effect in Lung Adenocarcinoma Cells.

Lactic acidosis (3 to 40 mM, pH < 6.9) is a condition found in solid tumors because tumor cells have a high rate of glucose consumption and lactate production even in the presence of oxygen; nevertheless, the microenvironment might still provide a sufficient glucose supply. Lactic acidosis has been proposed to shift metabolism from aerobic glycolysis toward oxidative phosphorylation (OXPHOS). We tested if lung tumor cells cultured under lactic acidosis shift their metabolism from glycolysis to OXPHOS by consuming extracellular lactate, increasing growth rate. We analyzed lung adenocarcinoma (A-549, A-427) cell lines and non-transformed fibroblast cells (MRC-5), which were cultured using RPMI-1640 medium initially containing lactate (2 mM) and glucose (10 mM), at pH 7.2 or 6.2 and oxygen tension 21% O2 (normoxia) or 2% O2 (hypoxia). We obtained growth curves, as well as glucose consumption and lactate production rates (measured during exponential growth) for each cell line. HIF-1α (Hypoxia-inducible factor 1 α), CS (citrate synthase) and AMPK (AMP-activated protein kinase) transcript levels were analyzed using RT-qPCR. By flow cytometry, we determined: (a) expression of glucose transporters (GLUT)1 and 4; (b) lactate transporters (MCT)1 and 4; (c) cell cycle profile, and (d) protein levels of HIF-1α, total and phosphorylated AMPK (pAMPK). Mitochondrial functionality was evaluated by measuring O2 consumption in tumor cells using polarography and a Clark-type electrode. Tumor and non-transformed cells used both aerobic glycolysis and OXPHOS for obtaining energy. As of 48 h of culture, lactate levels ranged from (4.5–14 mM), thus forming a lactic environment. Lactic acidosis diminished GLUT1/GLUT4 expression and glucose consumption in A-549, but not in A-427 cells, and induced differential expression of HIF-1α, AMPK, and CS transcripts. A-427 cells increased pAMPK and HIF-1α levels and shifted their metabolism increasing OXPHOS; thus supporting cell growth. Conversely, A-549 cells increased HIF-1α protein levels, but did not activate AMPK and diminished OXPHOS. A-549 cells survived by arresting cells in G1-phase. Our findings show that lactic acidosis diminishes Warburg effect in tumor cells, but this change does not necessarily promote a shift to OXPHOS. Hence, lung adenocarcinomas show a differential metabolic response even when they are under the same microenvironmental conditions.

Lactate Production Precedes Inflammatory Cell Recruitment in Arthritic Ankles: an Imaging Study

PurposeInflammation is involved in many disease processes. However, accurate imaging tools permitting diagnosis and characterization of inflammation are still missing. As inflamed tissues exhibit a high rate of glycolysis, pyruvate metabolism may offer a unique approach to follow the inflammatory response and disease progression. Therefore, the aim of the study was to follow metabolic changes and recruitment of inflammatory cells after onset of inflammation in arthritic ankles using hyperpolarized 1-13C-pyruvate magnetic resonance spectroscopy (MRS) and 19F magnetic resonance imaging (MRI), respectively.ProcedureExperimental rheumatoid arthritis (RA) was induced by intraperitoneal injection of glucose-6-phosphate-isomerase-specific antibodies (GPI) containing serum. To monitor pyruvate metabolism, the transformation of hyperpolarized 1-13C-pyruvate into hyperpolarized 1-13C-lactate was followed using MRS. To track phagocytic immune cell homing, we intravenously injected a perfluorocarbon emulsion 48 h before imaging. The animals were scanned at days 1, 3, or 6 after GPI-serum injection to examine the different stages of arthritic inflammation. Finally, to confirm the pyruvate metabolic activity and the link to inflammatory cell recruitment, we conducted hematoxylin-eosin histopathology and monocarboxylase transporter (MCT-1) immune histochemistry (IHC) of inflamed ankles.ResultsHyperpolarized 1-13C-pyruvate MRS revealed a high rate of lactate production immediately at day 1 after GPI-serum transfer, which remained elevated during the progression of the disease, while 19F-MRI exhibited a gradual recruitment of phagocytic immune cells in arthritic ankles, which correlated well with the course of ankle swelling. Histopathology and IHC revealed that MCT-1 was expressed in regions with inflammatory cell recruitment, confirming the metabolic shift identified in arthritic ankles.ConclusionsOur study demonstrated the presence of a very early metabolic shift in arthritic joints independent of phagocytic immune cell recruitment. Thus, hyperpolarized 1-13C-pyruvate represents a promising tracer to monitor acute arthritic joint inflammation, even with minor ankle swelling. Furthermore, translated to the clinics, these methods add a detailed characterization of disease status and could substantially support patient stratification and therapy monitoring.

Determination of Anaerobic Capacity - Reliability and Validity of Sprint Running Tests

PURPOSE: A number of physiological diagnostics were developed. However, the timeline-related diagnostics of maximal anaerobic glycolytic capacity remain unclear. The objective of this study was to evaluate the reliability and validity of a sprint running test to assess the anaerobic capacity.METHODS: The study was divided into three parts. Sixty-one male (24±4 years, 181.0±4.3 cm; 78.5±5.9 kg) and twelve female (25±3 years, 167.0±0.6 cm, 60.4±5.7 kg) sports students participated in this study. Twenty-five subjects (13 males, 24±2 years, 181.0±0.5 cm, 78.5±5.9 kg; 12 females, 25±3 years, 167.0±0.6 cm, 60.4±5.7 kg) performed incremental step tests at running track and several linear sprints on a running track (LSRT) with different time durations (8, 10, 12, and 14 seconds)(part I) on different days. Twenty-five male subjects (24±3 years, 180.7±6.7 cm, 84.6±8.8 kg) conducted a 10 or 12 second sprint running on a non-motorized treadmill (NMT)(part II). In part III, twenty-three male subjects (24±2 years, 181.4±5.8 cm, 74.5±7.4 kg) ran a 10 second LSRT and NMT on consecutive days. Capillary blood samplings were taken before (Lac&lt;sub&gt;r&lt;/sub&gt;) and after the sprint running for ten minutes at one minute intervals to find out maximal lactate concentration after exercise and to calculate the maximum lactate production rate (LPR&lt;sub&gt;max&lt;/sub&gt;).RESULTS: For all parts reliability for LPR&lt;sub&gt;max&lt;/sub&gt; was proven (Part I: 8 seconds: ICC: &lt;i&gt;r&lt;/i&gt;=.89; 10 seconds: ICC: &lt;i&gt;r&lt;/i&gt;=.82; 12 seconds: ICC: &lt;i&gt;r&lt;/i&gt;=.92; 14 seconds: &lt;i&gt;r&lt;/i&gt;=.84, respectively; Part II: 10 seconds: ICC: &lt;i&gt;r&lt;/i&gt;=.76; 12 seconds: ICC: &lt;i&gt;r&lt;/i&gt;=.79). To analyze validity for LPR&lt;sub&gt;max&lt;/sub&gt;, Part III was conducted and proven valid (ICC: &lt;i&gt;r&lt;/i&gt;=.96, p=.074).CONCLUSIONS: We demonstrate that LSRT and NMT reliably determine anaerobic capacity and can be used as a valid tool for physiological performance diagnostics.

Determination of Anaerobic Capacity - Reliability and Validity of Sprint Running Tests

PURPOSE: A number of physiological diagnostics were developed. However, the timeline-related diagnostics of maximal anaerobic glycolytic capacity remain unclear. The objective of this study was to evaluate the reliability and validity of a sprint running test to assess the anaerobic capacity.METHODS: The study was divided into three parts. Sixty-one male (24±4 years, 181.0±4.3 cm; 78.5±5.9 kg) and twelve female (25±3 years, 167.0±0.6 cm, 60.4±5.7 kg) sports students participated in this study. Twenty-five subjects (13 males, 24±2 years, 181.0±0.5 cm, 78.5±5.9 kg; 12 females, 25±3 years, 167.0±0.6 cm, 60.4±5.7 kg) performed incremental step tests at running track and several linear sprints on a running track (LSRT) with different time durations (8, 10, 12, and 14 seconds)(part I) on different days. Twenty-five male subjects (24±3 years, 180.7±6.7 cm, 84.6±8.8 kg) conducted a 10 or 12 second sprint running on a non-motorized treadmill (NMT)(part II). In part III, twenty-three male subjects (24±2 years, 181.4±5.8 cm, 74.5±7.4 kg) ran a 10 second LSRT and NMT on consecutive days. Capillary blood samplings were taken before (Lac&lt;sub&gt;r&lt;/sub&gt;) and after the sprint running for ten minutes at one minute intervals to find out maximal lactate concentration after exercise and to calculate the maximum lactate production rate (LPR&lt;sub&gt;max&lt;/sub&gt;).RESULTS: For all parts reliability for LPR&lt;sub&gt;max&lt;/sub&gt; was proven (Part I: 8 seconds: ICC: &lt;i&gt;r&lt;/i&gt;=.89; 10 seconds: ICC: &lt;i&gt;r&lt;/i&gt;=.82; 12 seconds: ICC: &lt;i&gt;r&lt;/i&gt;=.92; 14 seconds: &lt;i&gt;r&lt;/i&gt;=.84, respectively; Part II: 10 seconds: ICC: &lt;i&gt;r&lt;/i&gt;=.76; 12 seconds: ICC: &lt;i&gt;r&lt;/i&gt;=.79). To analyze validity for LPR&lt;sub&gt;max&lt;/sub&gt;, Part III was conducted and proven valid (ICC: &lt;i&gt;r&lt;/i&gt;=.96, p=.074).CONCLUSIONS: We demonstrate that LSRT and NMT reliably determine anaerobic capacity and can be used as a valid tool for physiological performance diagnostics.

The Role of Surface Termination in Halide Perovskites for Efficient Photocatalytic Synthesis.

Halide perovskites have received attention in the field of photocatalysis owing to their excellent optoelectronic properties. However, the semiconductor properties of halide perovskite surfaces and the influence on photocatalytic performance have not been systematically clarified. Now, the conversion of triose (such as 1,3-dihydroxyacetone (DHA)) is employed as a model reaction to explore the surface termination of MAPbI3 . By rational design of the surface termination for MAPbI3 , the production rate of butyl lactate is substantially improved to 7719 μg g-1 cat. h-1 under visible-light illumination. The MAI-terminated MAPbI3 surface governs the photocatalytic performance. Specially, MAI-terminated surface is susceptible to iodide oxidation, which thus promotes the exposure of PbII as active sites for this photocatalysis process. Moreover, MAI-termination induces a p-doping effect near the surface for MAPbI3 , which facilitates carrier transport and thus photosynthesis.

The Role of Surface Termination in Halide Perovskites for Efficient Photocatalytic Synthesis

AbstractHalide perovskites have received attention in the field of photocatalysis owing to their excellent optoelectronic properties. However, the semiconductor properties of halide perovskite surfaces and the influence on photocatalytic performance have not been systematically clarified. Now, the conversion of triose (such as 1,3‐dihydroxyacetone (DHA)) is employed as a model reaction to explore the surface termination of MAPbI3. By rational design of the surface termination for MAPbI3, the production rate of butyl lactate is substantially improved to 7719 μg g−1 cat. h−1 under visible‐light illumination. The MAI‐terminated MAPbI3 surface governs the photocatalytic performance. Specially, MAI‐terminated surface is susceptible to iodide oxidation, which thus promotes the exposure of PbII as active sites for this photocatalysis process. Moreover, MAI‐termination induces a p‐doping effect near the surface for MAPbI3, which facilitates carrier transport and thus photosynthesis.

MiR-16-5p/PDK4-Mediated Metabolic Reprogramming Is Involved in Chemoresistance of Cervical Cancer

Cervical cancer is one of the most prevalent malignancies in women worldwide. Therefore, investigation about molecular pathogenesis and related therapy targets of cervical cancer is an emergency. The molecular mechanisms responsible for the chemoresistance of cervical cancer were investigated by the use of doxorubicin (Dox)-resistant HeLa/Dox and SiHa/Dox cells. Our data showed that chemoresistant cells exhibited significantly higher glucose consumption, lactate production rate, and ATP levels than that of their parental cells. Among metabolic and glycolytic related genes, the expression of PDK4 was upregulated in Dox-resistant cells. Knockdown of PDK4 can decrease glucose consumption, lactate production rate, and ATP levels and further sensitize resistant cervical cancer cells to Dox treatment. By screening microRNAs (miRNAs), which can regulate expression of PDK4, we found that miR-16-5p was downregulated in chemoresistant cells. Overexpression of miR-16-5p can decrease the expression of PDK4 and sensitize the resistant cells to Dox treatment. Xenograft models confirmed that knockdown of PDK4 can increase chemotherapy efficiency for in vivo tumor growth. Collectively, our data suggested that miR-16-5p/PDK4-mediated metabolic reprogramming is involved in chemoresistance of cervical cancer.

Metformin enhances cytotoxic action of dichloroacetate against Lewis lung carcinoma cells in vitro.

To study the capability of MTF to enhance the antitumor action of DCA against Lewis lung carcinoma cells in vitro. LLC/R9, a low metastatic variant of Lewis lung carcinoma cells, was used. Effects of 30mM DCA in combination with 2mM MTF on cell survival, cell cycle distribution, apoptosis, mitochondrial potential, intracellular ATP level, glucose consumption, and lactate production rates were determined in vitro. MTF was shown to enhance the cytotoxic/cytostatic action of DCA against LLC/R9cells in vitro. Treatment of LLC/R9cells with 30mM DCA in combination with 2mM MTF resulted in a 39% decrease in the number of viable cells (p < 0.05), a 2.8-fold increase of the number of dead cells (p < 0.05), a near 2-fold decrease in the proportion of cells at the S-phase (p < 0.05), a 4-fold increase in the apoptosis (p < 0.05) and significant reduction (p < 0.05) of the mitochondrial membrane potential of tumor cells as compared to corresponding values in control. DCA alone reduced glucose consumption and lactate production rates by more than 26% (p < 0.05) and 34% (p < 0.05), respectively, whereas MTF counteracted these effects. Nevertheless, in the cells treated with both DCA and DCA in combination with MTF, the intracellular adenosine triphosphate increased by 33-35% compared with that in the control (p < 0.05). MTF enhanced the cytotoxic/cytostatic action of DCA against LLC/R9 cells in vitro, which points on their possible synergistic antitumor action in vivo.

Kinetics of Methyl Lactate Formation from the Transesterification of Polylactic Acid Catalyzed by Zn(II) Complexes.

The kinetics of the transesterification of polylactic acid (PLA) with methanol to form methyl lactate catalyzed by Zn(II) complexes was studied experimentally and numerically. The complexes, Zn(1Et)2 and Zn(2Pr)2, were synthesized from ethylenediamine and propylenediamine Schiff bases, respectively. The temperature range covered was 313.2–383.2 K. An increase in the reaction rate with the increase in temperature was observed for the Zn(1Et)2-catalyzed reaction. The temperature relationship of the rate coefficients can be explained by a linear Arrhenius dependency with constant activation energy. The kinetics of Zn(2Pr)2, on the other hand, is only explained by non-Arrhenius kinetics with convex variable activation energy, resulting in faster methyl lactate production rates at 323.2 and 343.2 K. The formation of a new catalyst species, likely through reaction with protic reagents, appears to promote the formation of intermediate complexes, resulting in the nonlinear behavior. Stirring speed induced the stability of the intermediate complexes. Contrary to Zn(1Et)2, Zn(2Pr)2 was susceptible to the presence of air/moisture in solution. The kinetic parameters were obtained by fitting the experimental data to the mass and energy balance of a consecutive second step reversible reaction taking place in a jacketed stirred batch reactor. For the case of Zn(2Pr)2, the activation energy was fitted to a four-parameter equation. The kinetic parameters presented in this work are valuable for the design of processes involving the chemical recycling of PLA into green solvents.

Brewers' spent grain liquor as a feedstock for lactate production with Lactobacillusdelbrueckii subsp. lactis.

Brewers’ spent grain (BSG) is a low‐cost by‐product of the brewing process. BSG liquor names the liquid components of BSG, mainly glucose, maltose, and long‐chain α‐1,4‐glycosidic bond glucose oligomers. These substances should be separated in existing BSG biorefineries, as they might lead to an increased formation of microbe‐inhibiting compounds in well‐established hydrothermal/enzymatic saccharification processes. In most cases, this liquid fraction is discarded. The present study presents for the first time an optimized process with BSG liquor for the purpose of producing bulk chemicals (e.g., lactate) in relevant concentrations. The process comprises the application of yeast extract, produced from own brewing processes, as the sole supplemented complex constituent in a simultaneous fermentation and saccharification process. Kinetic parameters for the final optimized process conditions with the organism Lactobacillus delbrueckii subsp. lactis were: maximum specific growth rate µmax = 0.47 h−1, maximum lactate concentration cLac, max = 79.06 g L−1, process yield YPS = 0.89 gLac gSugar −1, lactate production rate qP = 4.18 gLac gCDW −1 h−1, and productivity P 15 h = 4.93 gLac L−1 h−1. BSG liquor, linked with yeast extract from Brewers’ yeast, can be a promising substrate for further bioprocess engineering tasks and contribute to a holistic and sustainable usage of Brewers’ spent grain.

Role of Lactic Acid in Cancer Metabolism Under the Influences on Tumor Expansion and Metamorphosis: A Review

Anaerobic glycolysis advertise the production of energy under hypoxic condition (absence of oxygen), while in aerobic glycolysis, the Warburg effect gives a momentous benefit through remodeling carbohydrates seeds from production of energy to biochemical pathways. On the other side, high degree of lactate have been cooperated with feeble outcome in human tumors as well as glycolytic switch is correlated with high rate of glucose uptake and lactate production. Although lactic acid was primarily remind as indicator of the glycolytic process in which many mechanisms initiates from the study of normal tissue physiology and transported to the tumor indicating lactic acid in essence lactate anion and protons, straightly contributes to tumor growth and development. MCT1 & MCT4 monocarboxylate transporters have been justified as well-known facilitators of lactate exchanges between various cancer cells with many metabolic behaviors. In this review, we summarize the current knowledge on the role of lactic acidosis and metastasis, lactate shuttles and lactate signaling molecules.

Resibufogenin suppresses tumor growth and inhibits glycolysis in ovarian cancer by modulating PIM1.

Ovarian cancer is a common human malignancy of the female reproductive system. However, chemotherapy has been proven to have limited effectiveness in a majority of patients. Resibufogenin (RB) is a major active ingredient in cinobufacini, which has been used in the treatment of human malignancies as adjunct agents. This study was designed to examine the anti-cancer effect of RB and the underlying mechanisms in ovarian cancer. Our results showed that RB treatment resulted in cell death, cell cycle arrest, and apoptosis in ovarian cancer cells. The anti-growth and pro-apoptotic effects of RB were also validated in xenograft mice models. Proteomics analysis indicated that RB was able to alter the expressions of several genes, which were involved in the regulation of glycolysis. The suppression effect of RB in the glycolysis pathway of ovarian cancer cells was validated by decreased glucose consumption, lactate production, and extracellular acidification rate (ECAR). We proposed that PIM1 functioned as the key target that mediated the anti-cancer effect of RB against ovarian cancer cells. Our results have revealed that RB downregulated PIM1 in ovarian cancer cells and its downstream genes involved in glycolysis. Moreover, our results indicated that the anti-growth activities and suppressing effect of RB on glycolysis were enhanced significantly by PIM1 knockdown but was attenuated by ectopic PIM1 expression. This provided evidence to support the role of PIM1 in the anti-cancer activities of RB.

MiR-489-3p/SIX1 Axis Regulates Melanoma Proliferation and Glycolytic Potential

Sine oculis homeobox 1 (SIX1), a key transcription factor for regulating aerobic glycolysis, participates in the occurrence of various cancer types. However, the role of SIX1 in melanoma and the upstream regulating mechanisms of SIX1 remain to be further investigated. MicroRNAs (miRNAs) have emerged as key regulators in tumorigenesis and progression. Here, we show that miR-489-3p suppresses SIX1 expression by directly targeting its 3′ untranslated region (3′ UTR) in melanoma cells. miR-489-3p suppressed melanoma cell proliferation, migration, and invasion through inhibition of SIX1. Mechanistically, by targeting SIX1, miR-489-3p dampens glycolysis, with decreased glucose uptake, lactate production, ATP generation, and extracellular acidification rate (ECAR), as well as an increased oxygen consumption rate (OCR). Importantly, glycolysis regulated by the miR-489-3p/SIX1 axis is critical for its regulation of melanoma growth and metastasis both in vitro and in vivo. In melanoma patients, miR-489-3p expression is negatively correlated with SIX1 expression. In addition, patients who had increased glucose uptake in tumors and with metastasis assessed by positron emission tomography (PET) scans showed decreased miR-489-3p expression and increased expression of SIX1. Collectively, our study demonstrates the importance of the miR-489-3p/SIX1 axis in melanoma, which can be a potential and a promising therapeutic target in melanoma.

Regulation of the Elevated T Cell Glycolysis May Alleviate Acute Graft-Versus-Host Disease Post-Allotransplant

Background: Acute graft-versus-host disease(aGVHD) remains a major complication following allogeneic hematopoietic stem cell transplantation(allo-HSCT). The pathogenesis of aGVHD is commonly considered to be caused by exaggerated and undesirable immune responses. Metabolism not only provide energy and substrates for T cell growth and survival, but also instruct effector functions, differentiation, and gene expression of T cells. In this regard, the metabolic profile of T cells was reported to play a critical role in the occurrence and development of many immunological disorders such as systemic lupus erythematosus and rheumatoid arthritis. Murine studies found that alloreactive T cells use aerobic glycolysis as the predominant metabolic process to meet activation and proliferation demand after allo-HSCT. However, the metabolic profile of T cells and the approach for regulating T cell metabolism in aGVHD patients remains to be elucidated. Aims: To determine the metabolic state in T cells of patients with aGVHD. Moreover, to investigate the effect of the novel approach targeting the abnormal metabolism in T cells of aGVHD patients, which may provide a potential therapeutic target for aGVHD patients after allo-HSCT. Methods: In this prospective case-control study, a total of 25 patients with aGVHD and 25 matched patients without aGVHD(non-aGVHD) after allo-HSCT were enrolled. T cell subsets were analyzed in aGVHD and non-aGVHD patients by flow cytometry. Th1, Th2, Th17, and Treg cells were identified as CD4+IFN-γ+, CD4+IL-4+, CD4+IL17A+, and CD4+CD25+Foxp3+, respectively. Tc1 and Tc2 cells were identified as CD8+IFN-γ+ and CD8+IL-4+, respectively. In order to determine the metabolic state in T cells of patients with aGVHD and non-aGVHD, the metabolic profile was determined using a Seahorse XF96 Analyzer. The glucose consumption and lactate production rates were detected by glucose assay kit and lactate assay kit. The mitochondrial mass, the mitochondrial membrane potential, the protein expressions for the lipid metabolism enzyme CTP1a and the glycolytic activator PFKFB3 were measured by flow cytometry. To further understand the metabolic state of T cells in aGVHD and non-aGVHD patients and investigate its mechanism, RNA sequencing (RNA-Seq) was performed to analyze the gene expression profiles of T cells. Subsequently, to explore the potential way of targeting the abnormal metabolism in T cells, the glycolysis inhibitor 3-PO was administrated to T cells from aGVHD patients. Results: When compared with T cells in non-aGVHD patients, T cells in aGVHD patients were polarized towards pro-inflammatory T cells, characterized by an elevated proportion of Tc1, Th1 and Th17. Furthermore, T cells isolated from aGVHD patients exhibited higher extracellular acidification rate, as well as the increased glucose consumption rate and lactate production rate compared to those in non-aGVHD patients. Moreover, elevated expression of PFKFB3 was observed in T cells, especially in naïve T cells of aGVHD patients, but oxygen consumption rate, CPT1A, mitochondrial mass or membrane potential showed no significant differences in T cells between aGVHD and non-aGVHD patients. These results implied higher glycolytic activity of T cells in aGVHD patients when compared with those in non-aGVHD patients. Consistent with the increased glycolytic activity observed in T cells from aGVHD patients, the mRNA levels of genes involved in the glycolytic pathway were substantially elevated in T cells of aGVHD patients compared to those in non-aGVHD patients. Importantly, in vitro treatment with glycolysis inhibitor 3-PO improved the activity of T cells derived from aGVHD patients through down-regulating glycolytic activity of T cells. Summary/Conclusion: The current study demonstrated that T cells in aGVHD patients preferentially depend on glycolysis to meet activation and proliferation demands. Furthermore, the activity of T cells from aGVHD patients could be ameliorated by glycolysis inhibitor 3-PO in vitro. Although further validation is required, T cell glycolysis promises to be a novel therapeutic target for aGVHD patients after allo-HSCT. Disclosures No relevant conflicts of interest to declare.

Programmed cell death ligand-1-mediated enhancement of hexokinase 2 expression is inversely related to T-cell effector gene expression in non-small-cell lung cancer

BackgroundWe investigated the role of PD-L1 in the metabolic reprogramming of non-small cell lung cancer (NSCLC).MethodsChanges in glycolysis-related molecules and glycolytic activity were evaluated in PD-L1low and PD-L1high NSCLC cells after transfection or knockdown of PD-L1, respectively. Jurkat T-cell activation was assessed after co-culture with NSCLC cells. The association between PD-L1 and immune response-related molecules or glycolysis were analyzed in patients with NSCLC and The Cancer Genome Atlas (TCGA).ResultsTransfecting PD-L1 in PD-L1low cells enhanced hexokinase-2 (HK2) expression, lactate production, and extracellular acidification rates, but minimally altered GLUT1 and PKM2 expression and oxygen consumption rates. By contrast, knocking-down PD-L1 in PD-L1high cells decreased HK2 expression and glycolysis by suppressing PI3K/Akt and Erk pathways. Interferon-γ (IFNγ) secretion and activation marker expression was decreased in stimulated Jurkat T-cells when co-cultured with HK2-overexpressing vector-transfected tumor cells rather than empty vector-transfected tumor cells. Immunohistochemistry revealed that PD-L1 expression was positively correlated with HK2 expression in NSCLC (p < 0.001). In TCGA, HK2 exhibited a positive linear association with CD274 (PD-L1) expression (p < 0.001) but an inverse correlation with the expression of CD4, CD8A, and T-cell effector function-related genes in the CD274high rather than CD274low group. Consistently, there were fewer CD8+ T-cells in PD-L1positive/HK2high tumors compared to PD-L1positive/HK2low tumors in squamous cell carcinoma.ConclusionsPD-L1 enhances glycolysis in NSCLC by upregulating HK2, which might dampen anti-tumor immunity. PD-L1 may contribute to NSCLC oncogenesis by inducing metabolic reprogramming and immune checkpoint.

TWIST1 transcriptionally regulates glycolytic genes to promote the Warburg metabolism in pancreatic cancer

Reprogrammed glucose metabolism is essential for tumor initiation and development, especially for pancreatic ductal adenocarcinoma (PDAC). Most cancer cells rely on aerobic glycolysis, a phenomenon termed “the Warburg effect”, to support uncontrolled proliferation and evade apoptosis. However, the direct regulators of the Warburg effect remain areas of active investigation. In this study, we found that the highly conserved transcription factor, TWIST1, is a crucial regulator of aerobic glycolysis in PDAC. Genetic silencing of TWIST1 significantly inhibited the glycolytic phenotypes of PDAC cells as revealed by reduced glucose uptake, lactate production, and extracellular acidification rate, which can be restored by re-expression of siRNA-resistant TWIST1. Moreover, tamoxifen-inducible expression of TWIST1 promoted the Warburg metabolism of PDAC cells. Mechanistically, by luciferase reporter assay and chromatin immunoprecipitation experiment, we showed that TWIST1 can directly increase the expression of several glycolytic genes, including SLC2A1, HK2, ENO1, and PKM2. Of note, the transcriptional regulation by TWIST1 was not dependent on HIF1α or c-Myc. In The Cancer Genome Atlas and Gene Expression Omnibus accession GSE15471, we confirmed that TWIST1 was closely associated with the glycolysis pathway. Collectively, our findings indicate that TWIST1 is likely to act as important regulator of the Warburg effect in PDAC.

Comparison of dynamic brain metabolism during antegrade cerebral perfusion versus deep hypothermic circulatory arrest using proton magnetic resonance spectroscopy

Comparison of dynamic brain metabolism during antegrade cerebral perfusion versus deep hypothermic circulatory arrest using proton magnetic resonance spectroscopy