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1.
J Transl Med ; 18(1): 257, 2020 06 25.
Artigo em Inglês | MEDLINE | ID: mdl-32586380

RESUMO

BACKGROUND: The recent global pandemic has placed a high priority on identifying drugs to prevent or lessen clinical infection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), caused by Coronavirus disease-2019 (COVID-19). METHODS: We applied two computational approaches to identify potential therapeutics. First, we sought to identify existing FDA approved drugs that could block coronaviruses from entering cells by binding to ACE2 or TMPRSS2 using a high-throughput AI-based binding affinity prediction platform. Second, we sought to identify FDA approved drugs that could attenuate the gene expression patterns induced by coronaviruses, using our Disease Cancelling Technology (DCT) platform. RESULTS: Top results for ACE2 binding iincluded several ACE inhibitors, a beta-lactam antibiotic, two antiviral agents (Fosamprenavir and Emricasan) and glutathione. The platform also assessed specificity for ACE2 over ACE1, important for avoiding counterregulatory effects. Further studies are needed to weigh the benefit of blocking virus entry against potential counterregulatory effects and possible protective effects of ACE2. However, the data herein suggest readily available drugs that warrant experimental evaluation to assess potential benefit. DCT was run on an animal model of SARS-CoV, and ranked compounds by their ability to induce gene expression signals that counteract disease-associated signals. Top hits included Vitamin E, ruxolitinib, and glutamine. Glutathione and its precursor glutamine were highly ranked by two independent methods, suggesting both warrant further investigation for potential benefit against SARS-CoV-2. CONCLUSIONS: While these findings are not yet ready for clinical translation, this report highlights the potential use of two bioinformatics technologies to rapidly discover existing therapeutic agents that warrant further investigation for established and emerging disease processes.


Assuntos
Betacoronavirus/fisiologia , Biologia Computacional , Infecções por Coronavirus/genética , Infecções por Coronavirus/terapia , Pneumonia Viral/genética , Pneumonia Viral/terapia , Animais , Betacoronavirus/genética , Regulação da Expressão Gênica , Glutamina/metabolismo , Humanos , Camundongos , Pandemias , Peptidil Dipeptidase A/metabolismo , Serina Endopeptidases/metabolismo
2.
Nature ; 583(7814): 109-114, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32528181

RESUMO

Hibernating mammals actively lower their body temperature to reduce energy expenditure when facing food scarcity1. This ability to induce a hypometabolic state has evoked great interest owing to its potential medical benefits2,3. Here we show that a hypothalamic neuronal circuit in rodents induces a long-lasting hypothermic and hypometabolic state similar to hibernation. In this state, although body temperature and levels of oxygen consumption are kept very low, the ability to regulate metabolism still remains functional, as in hibernation4. There was no obvious damage to tissues and organs or abnormalities in behaviour after recovery from this state. Our findings could enable the development of a method to induce a hibernation-like state, which would have potential applications in non-hibernating mammalian species including humans.


Assuntos
Metabolismo Energético/fisiologia , Hibernação/fisiologia , Hipotálamo/citologia , Hipotálamo/fisiologia , Vias Neurais/citologia , Vias Neurais/fisiologia , Animais , Metabolismo Basal/fisiologia , Núcleo Hipotalâmico Dorsomedial/citologia , Núcleo Hipotalâmico Dorsomedial/fisiologia , Feminino , Neurônios GABAérgicos/metabolismo , Glutamina/metabolismo , Masculino , Camundongos , Consumo de Oxigênio/fisiologia
3.
Nature ; 583(7814): 115-121, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32528180

RESUMO

The advent of endothermy, which is achieved through the continuous homeostatic regulation of body temperature and metabolism1,2, is a defining feature of mammalian and avian evolution. However, when challenged by food deprivation or harsh environmental conditions, many mammalian species initiate adaptive energy-conserving survival strategies-including torpor and hibernation-during which their body temperature decreases far below its homeostatic set-point3-5. How homeothermic mammals initiate and regulate these hypothermic states remains largely unknown. Here we show that entry into mouse torpor, a fasting-induced state with a greatly decreased metabolic rate and a body temperature as low as 20 °C6, is regulated by neurons in the medial and lateral preoptic area of the hypothalamus. We show that restimulation of neurons that were activated during a previous bout of torpor is sufficient to initiate the key features of torpor, even in mice that are not calorically restricted. Among these neurons we identify a population of glutamatergic Adcyap1-positive cells, the activity of which accurately determines when mice naturally initiate and exit torpor, and the inhibition of which disrupts the natural process of torpor entry, maintenance and arousal. Taken together, our results reveal a specific neuronal population in the mouse hypothalamus that serves as a core regulator of torpor. This work forms a basis for the future exploration of mechanisms and circuitry that regulate extreme hypothermic and hypometabolic states, and enables genetic access to monitor, initiate, manipulate and study these ancient adaptations of homeotherm biology.


Assuntos
Metabolismo Energético/fisiologia , Hipotálamo/citologia , Vias Neurais/fisiologia , Neurônios/fisiologia , Torpor/fisiologia , Animais , Jejum , Feminino , Privação de Alimentos , Glutamina/metabolismo , Hipotálamo/fisiologia , Masculino , Camundongos , Polipeptídeo Hipofisário Ativador de Adenilato Ciclase/metabolismo
4.
J Anim Sci ; 98(6)2020 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-32415842

RESUMO

This study determined whether extracellular citrulline is degraded by ruminal bacteria of sheep. In the first experiment, whole rumen fluid (3 mL) from six adult Suffolk sheep was incubated at 37 °C with 5 mM l-glutamine (Gln), l-glutamate (Glu), l-arginine (Arg), or l-citrulline (Cit) for 0, 0.5, 1, and 2 h or with 0, 0.5, 2, or 5 mM Gln, Glu, Arg, or Cit for 2 h. An aliquot (50 µL) of the incubation solution was collected at the predetermined time points for amino acids (AA) analyses. Results showed extensive hydrolysis of Gln into Glu and ammonia, of Arg into l-ornithine and l-proline, but little or no degradation of extracellular Cit or Glu by ruminal microbes. In the second experiment, six adult Suffolk sheep were individually fed each of three separate supplements (8 g Gln , Cit, or urea) on three separate days along with regular feed (800 g/animal). Blood (2 mL) was sampled from the jugular vein prior to feeding (time 0) and at 0.5, 1, 2, and 4 h after consuming the supplement. Plasma was analyzed for AA, glucose, ammonia, and urea. The concentrations of Cit in the plasma of sheep consuming this AA increased (P < 0.001) by 117% at 4 h and those of Arg increased by 23% at 4 h, compared with the baseline values. Urea or Gln feeding did not affect (P > 0.05) the concentrations of Cit or Arg in plasma. These results indicate that Cit is not metabolized by ruminal microbes of sheep and is, therefore, absorbed as such by the small intestine and used for the synthesis of Arg by extrahepatic tissues.


Assuntos
Bactérias/metabolismo , Citrulina/metabolismo , Rúmen/microbiologia , Ovinos/microbiologia , Amônia/metabolismo , Ração Animal/análise , Animais , Arginina/metabolismo , Dieta/veterinária , Suplementos Nutricionais , Feminino , Ácido Glutâmico/metabolismo , Glutamina/metabolismo , Ornitina/sangue , Prolina/metabolismo , Ureia
5.
PLoS One ; 15(4): e0231770, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32298377

RESUMO

The Warburg effect, a hallmark of cancer, has recently been identified as a metabolic limitation of Chinese Hamster Ovary (CHO) cells, the primary platform for the production of monoclonal antibodies (mAb). Metabolic engineering approaches, including genetic modifications and feeding strategies, have been attempted to impose the metabolic prevalence of respiration over aerobic glycolysis. Their main objective lies in decreasing lactate production while improving energy efficiency. Although yielding promising increases in productivity, such strategies require long development phases and alter entangled metabolic pathways which singular roles remain unclear. We propose to apply drugs used for the metabolic therapy of cancer to target the Warburg effect at different levels, on CHO cells. The use of α-lipoic acid, a pyruvate dehydrogenase activator, replenished the Krebs cycle through increased anaplerosis but resulted in mitochondrial saturation. The electron shuttle function of a second drug, methylene blue, enhanced the mitochondrial capacity. It pulled on anaplerotic pathways while reducing stress signals and resulted in a 24% increase of the maximum mAb production. Finally, the combination of both drugs proved to be promising for stimulating Krebs cycle activity and mitochondrial respiration. Therefore, drugs used in metabolic therapy are valuable candidates to understand and improve the metabolic limitations of CHO-based bioproduction.


Assuntos
Anticorpos Monoclonais/biossíntese , Ciclo do Ácido Cítrico/fisiologia , Glicólise/efeitos dos fármacos , Engenharia Metabólica/métodos , Azul de Metileno/farmacologia , Ácido Tióctico/farmacologia , Animais , Células CHO , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Cricetulus , Glucose/metabolismo , Glutamina/metabolismo , Glicólise/fisiologia , Ácido Láctico/metabolismo , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Azul de Metileno/metabolismo , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Fosforilação Oxidativa/efeitos dos fármacos , Estresse Oxidativo/efeitos dos fármacos , Respiração , Ácido Tióctico/metabolismo
6.
PLoS Comput Biol ; 16(4): e1007780, 2020 04.
Artigo em Inglês | MEDLINE | ID: mdl-32298259

RESUMO

Metabolism plays an essential role in cell fate decisions. However, the methods used for metabolic characterization and for finding potential metabolic regulators are still based on characterizing cellular metabolic steady-state which is dependent on the extracellular environment. In this work, we hypothesized that the response dynamics of intracellular metabolic pools to extracellular stimuli is controlled in a cell type-specific manner. We applied principles of process dynamics and control to human induced pluripotent stem cells (hiPSC) and human neural stem cells (hNSC) subjected to a sudden extracellular glutamine step. The fold-changes of steady-states and the transient profiles of metabolic pools revealed that dynamic responses were reproducible and cell type-specific. Importantly, many amino acids had conserved dynamics and readjusted their steady state concentration in response to the increased glutamine influx. Overall, we propose a novel methodology for systematic metabolic characterization and identification of potential metabolic regulators.


Assuntos
Células-Tronco Pluripotentes Induzidas , Redes e Vias Metabólicas/fisiologia , Células-Tronco Neurais , Reatores Biológicos , Células Cultivadas , Biologia Computacional , Espaço Extracelular/química , Espaço Extracelular/metabolismo , Glutamina/metabolismo , Glutamina/farmacologia , Humanos , Células-Tronco Pluripotentes Induzidas/citologia , Células-Tronco Pluripotentes Induzidas/efeitos dos fármacos , Células-Tronco Pluripotentes Induzidas/metabolismo , Células-Tronco Neurais/citologia , Células-Tronco Neurais/efeitos dos fármacos , Células-Tronco Neurais/metabolismo
7.
Proc Natl Acad Sci U S A ; 117(18): 9932-9941, 2020 05 05.
Artigo em Inglês | MEDLINE | ID: mdl-32312819

RESUMO

Cellular starvation is typically a consequence of tissue injury that disrupts the local blood supply but can also occur where cell populations outgrow the local vasculature, as observed in solid tumors. Cells react to nutrient deprivation by adapting their metabolism, or, if starvation is prolonged, it can result in cell death. Cell starvation also triggers adaptive responses, like angiogenesis, that promote tissue reorganization and repair, but other adaptive responses and their mediators are still poorly characterized. To explore this issue, we analyzed secretomes from glucose-deprived cells, which revealed up-regulation of multiple cytokines and chemokines, including IL-6 and IL-8, in response to starvation stress. Starvation-induced cytokines were cell type-dependent, and they were also released from primary epithelial cells. Most cytokines were up-regulated in a manner dependent on NF-κB and the transcription factor of the integrated stress response ATF4, which bound directly to the IL-8 promoter. Furthermore, glutamine deprivation, as well as the antimetabolic drugs 2-deoxyglucose and metformin, also promoted the release of IL-6 and IL-8. Finally, some of the factors released from starved cells induced chemotaxis of B cells, macrophages, and neutrophils, suggesting that nutrient deprivation in the tumor environment can serve as an initiator of tumor inflammation.


Assuntos
Inflamação/genética , Interleucina-6/genética , Interleucina-8/genética , Neoplasias/metabolismo , Estresse Fisiológico/genética , Fator 4 Ativador da Transcrição/genética , Fator 4 Ativador da Transcrição/metabolismo , Antimetabólitos/farmacologia , Morte Celular/efeitos dos fármacos , Desoxiglucose/farmacologia , Células Epiteliais/metabolismo , Células Epiteliais/patologia , Regulação da Expressão Gênica/efeitos dos fármacos , Regulação da Expressão Gênica/imunologia , Glucose/metabolismo , Glutamina/metabolismo , Células HeLa , Humanos , Inflamação/imunologia , Inflamação/metabolismo , Macrófagos/imunologia , Macrófagos/metabolismo , Metformina/farmacologia , NF-kappa B/genética , Neoplasias/genética , Regiões Promotoras Genéticas/genética , Inanição/genética , Inanição/metabolismo , Estresse Fisiológico/imunologia
8.
Proc Natl Acad Sci U S A ; 117(19): 10294-10304, 2020 05 12.
Artigo em Inglês | MEDLINE | ID: mdl-32341162

RESUMO

Many cancer cells consume glutamine at high rates; counterintuitively, they simultaneously excrete glutamate, the first intermediate in glutamine metabolism. Glutamine consumption has been linked to replenishment of tricarboxylic acid cycle (TCA) intermediates and synthesis of adenosine triphosphate (ATP), but the reason for glutamate excretion is unclear. Here, we dynamically profile the uptake and excretion fluxes of a liver cancer cell line (HepG2) and use genome-scale metabolic modeling for in-depth analysis. We find that up to 30% of the glutamine is metabolized in the cytosol, primarily for nucleotide synthesis, producing cytosolic glutamate. We hypothesize that excreting glutamate helps the cell to increase the nucleotide synthesis rate to sustain growth. Indeed, we show experimentally that partial inhibition of glutamate excretion reduces cell growth. Our integrative approach thus links glutamine addiction to glutamate excretion in cancer and points toward potential drug targets.


Assuntos
Trifosfato de Adenosina/metabolismo , Carcinoma Hepatocelular/patologia , Citosol/metabolismo , Ácido Glutâmico/metabolismo , Glutamina/metabolismo , Neoplasias Hepáticas/patologia , Mitocôndrias/metabolismo , Carcinoma Hepatocelular/metabolismo , Células Cultivadas , Ciclo do Ácido Cítrico , Metabolismo Energético , Células Hep G2 , Humanos , Neoplasias Hepáticas/metabolismo
9.
Science ; 368(6487): 197-201, 2020 04 10.
Artigo em Inglês | MEDLINE | ID: mdl-32273471

RESUMO

Vulnerability to relapse during periods of attempted abstinence from cocaine use is hypothesized to result from the rewiring of brain reward circuitries, particularly ventral tegmental area (VTA) dopamine neurons. How cocaine exposures act on midbrain dopamine neurons to precipitate addiction-relevant changes in gene expression is unclear. We found that histone H3 glutamine 5 dopaminylation (H3Q5dop) plays a critical role in cocaine-induced transcriptional plasticity in the midbrain. Rats undergoing withdrawal from cocaine showed an accumulation of H3Q5dop in the VTA. By reducing H3Q5dop in the VTA during withdrawal, we reversed cocaine-mediated gene expression changes, attenuated dopamine release in the nucleus accumbens, and reduced cocaine-seeking behavior. These findings establish a neurotransmission-independent role for nuclear dopamine in relapse-related transcriptional plasticity in the VTA.


Assuntos
Transtornos Relacionados ao Uso de Cocaína/metabolismo , Transtornos Relacionados ao Uso de Cocaína/psicologia , Cocaína/efeitos adversos , Dopamina/metabolismo , Neurônios Dopaminérgicos/metabolismo , Comportamento de Procura de Droga , Histonas/metabolismo , Área Tegmentar Ventral/metabolismo , Animais , Transtornos Relacionados ao Uso de Cocaína/genética , Regulação da Expressão Gênica , Glutamina/metabolismo , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Plasticidade Neuronal , Núcleo Accumbens/metabolismo , Ratos , Ratos Sprague-Dawley , Transmissão Sináptica
10.
Nat Chem Biol ; 16(7): 756-765, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32284601

RESUMO

Soluble prion proteins contingently encounter foreign prion aggregates, leading to cross-species prion transmission. However, how its efficiency is regulated by structural fluctuation of the host soluble prion protein remains unsolved. In the present study, through the use of two distantly related yeast prion Sup35 proteins, we found that a specific conformation of a short disordered segment governs interspecies prion transmissibility. Using a multidisciplinary approach including high-resolution NMR and molecular dynamics simulation, we identified critical residues within this segment that allow interspecies prion transmission in vitro and in vivo, by locally altering dynamics and conformation of soluble prion proteins. Remarkably, subtle conformational differences caused by a methylene group between asparagine and glutamine sufficed to change the short segment structure and substantially modulate the cross-seeding activity. Thus, our findings uncover how conformational dynamics of the short segment in the host prion protein impacts cross-species prion transmission. More broadly, our study provides mechanistic insights into cross-seeding between heterologous proteins.


Assuntos
Asparagina/química , Glutamina/química , Proteínas Intrinsicamente Desordenadas/química , Fatores de Terminação de Peptídeos/química , Príons/química , Proteínas de Saccharomyces cerevisiae/química , Saccharomyces cerevisiae/genética , Sequência de Aminoácidos , Asparagina/metabolismo , Clonagem Molecular , Escherichia coli/genética , Escherichia coli/metabolismo , Expressão Gênica , Vetores Genéticos/química , Vetores Genéticos/metabolismo , Glutamina/metabolismo , Proteínas Intrinsicamente Desordenadas/genética , Proteínas Intrinsicamente Desordenadas/metabolismo , Simulação de Dinâmica Molecular , Fatores de Terminação de Peptídeos/genética , Fatores de Terminação de Peptídeos/metabolismo , Príons/genética , Príons/metabolismo , Domínios e Motivos de Interação entre Proteínas , Isoformas de Proteínas/química , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Estrutura Secundária de Proteína , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/metabolismo , Alinhamento de Sequência , Homologia de Sequência de Aminoácidos , Termodinâmica
11.
Nat Commun ; 11(1): 1277, 2020 03 09.
Artigo em Inglês | MEDLINE | ID: mdl-32152301

RESUMO

Although supplemental oxygen is required to promote survival of severely premature infants, hyperoxia is simultaneously harmful to premature developing tissues such as in the retina. Here we report the effect of hyperoxia on central carbon metabolism in primary mouse Müller glial cells and a human Müller glia cell line (M10-M1 cells). We found decreased flux from glycolysis entering the tricarboxylic acid cycle in Müller cells accompanied by increased glutamine consumption in response to hyperoxia. In hyperoxia, anaplerotic catabolism of glutamine by Müller cells increased ammonium release two-fold. Hyperoxia induces glutamine-fueled anaplerosis that reverses basal Müller cell metabolism from production to consumption of glutamine.


Assuntos
Células Ependimogliais/metabolismo , Glutamina/metabolismo , Hiperóxia/metabolismo , Animais , Astrócitos/metabolismo , Isótopos de Carbono , Células Cultivadas , Células Endoteliais/metabolismo , Glucose/metabolismo , Glutaminase/metabolismo , Glicólise , Humanos , Metaboloma , Camundongos , Mitocôndrias/metabolismo , Modelos Biológicos , Oxirredução , Fosforilação , Complexo Piruvato Desidrogenase/metabolismo
12.
Nat Commun ; 11(1): 1559, 2020 03 25.
Artigo em Inglês | MEDLINE | ID: mdl-32214088

RESUMO

Microglia are highly motile cells that continuously monitor the brain environment and respond to damage-associated cues. While glucose is the main energy substrate used by neurons in the brain, the nutrients metabolized by microglia to support surveillance of the parenchyma remain unexplored. Here, we use fluorescence lifetime imaging of intracellular NAD(P)H and time-lapse two-photon imaging of microglial dynamics in vivo and in situ, to show unique aspects of the microglial metabolic signature in the brain. Microglia are metabolically flexible and can rapidly adapt to consume glutamine as an alternative metabolic fuel in the absence of glucose. During insulin-induced hypoglycemia in vivo or in aglycemia in acute brain slices, glutaminolysis supports the maintenance of microglial process motility and damage-sensing functions. This metabolic shift sustains mitochondrial metabolism and requires mTOR-dependent signaling. This remarkable plasticity allows microglia to maintain their critical surveillance and phagocytic roles, even after brain neuroenergetic homeostasis is compromised.


Assuntos
Encéfalo/imunologia , Metabolismo Energético/fisiologia , Microglia/metabolismo , Animais , Encéfalo/patologia , Receptor 1 de Quimiocina CX3C/genética , Movimento Celular , Ácidos Graxos/metabolismo , Glucose/deficiência , Glucose/metabolismo , Glutamina/metabolismo , Vigilância Imunológica , Camundongos , Camundongos Transgênicos , Microglia/citologia , Microglia/imunologia , NAD/metabolismo , Serina-Treonina Quinases TOR/antagonistas & inibidores , Serina-Treonina Quinases TOR/metabolismo
13.
Nat Commun ; 11(1): 1320, 2020 03 17.
Artigo em Inglês | MEDLINE | ID: mdl-32184390

RESUMO

Glucose metabolism is remodeled in cancer, but the global pattern of cancer-specific metabolic changes remains unclear. Here we show, using the comprehensive measurement of metabolic enzymes by large-scale targeted proteomics, that the metabolism both carbon and nitrogen is altered during the malignant progression of cancer. The fate of glutamine nitrogen is shifted from the anaplerotic pathway into the TCA cycle to nucleotide biosynthesis, with this shift being controlled by glutaminase (GLS1) and phosphoribosyl pyrophosphate amidotransferase (PPAT). Interventions to reduce the PPAT/GLS1 ratio suppresses tumor growth of many types of cancer. A meta-analysis reveals that PPAT shows the strongest correlation with malignancy among all metabolic enzymes, in particular in neuroendocrine cancer including small cell lung cancer (SCLC). PPAT depletion suppresses the growth of SCLC lines. A shift in glutamine fate may thus be required for malignant progression of cancer, with modulation of nitrogen metabolism being a potential approach to SCLC treatment.


Assuntos
Progressão da Doença , Glutamina/metabolismo , Neoplasias/metabolismo , Neoplasias/patologia , Nitrogênio/metabolismo , Amidofosforribosiltransferase/metabolismo , Animais , Vias Biossintéticas , Linhagem Celular Tumoral , Regulação Neoplásica da Expressão Gênica , Glutaminase/metabolismo , Humanos , Metabolômica , Camundongos Nus , Modelos Biológicos , Terapia de Alvo Molecular , Neoplasias/genética , Prognóstico
14.
Poult Sci ; 99(3): 1454-1461, 2020 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-32115031

RESUMO

The aim of the present study was to evaluate the effect of glutamine (Gln) on modulating heat stress-induced oxidative damage in the broiler thigh muscle through nuclear factor erythroid 2-related 2/Kelch-like ECH-associated protein 1 (Nrf2-Keap1) pathway. Three-hundred 22-day-old Arbor Acres broilers were reallocated into 5 groups: a control group (24 °C) fed with basal diet and 4 heat stress (HS) groups (34 °C for 8 h/D) fed with basal diet containing 0, 0.5, 1.0, and 1.5% Gln. This experiment lasted 21 D. Heat stress decreased (P < 0.05) pH, redness, and Gln levels, and increased (P < 0.05) luminance, water loss rate, and cooking loss (CL) values of the thigh meat. Compared with the HS group, supplementation with 1.5% Gln increased (P < 0.05) pH, redness, and Gln levels, but decreased (P < 0.05) luminance and CL values in the thigh meat. There were significant decreases (P < 0.05) in glutathione (GSH), superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GSH-Px), total antioxidant capacity (T-AOC), and Nrf2 levels, but significant increases (P < 0.05) in the malondialdehyde (MDA) and Keap1 levels of the thigh muscle after HS treatment. Compared with the HS group, supplementation with 1.0, and 1.5% Gln decreased (P < 0.05) MDA and Keap1 levels; supplementation with 1.5% Gln increased (P < 0.05) GSH, GSH-Px, T-AOC, CAT, SOD, and Nrf2 levels in the thigh muscle of heat-stressed broilers. Furthermore, HS decreased (P < 0.05) Nrf2, SOD, CAT, and GSH-Px mRNA expression levels, but increased (P < 0.05) Keap1 mRNA level in the thigh muscle of broiler. Dietary supplementation with 1.5% Gln increased (P < 0.05) Nrf2, GSH-Px, CAT, and SOD mRNA expression levels, but decreased (P < 0.05) Keap1 mRNA level in the thigh muscle of heat-stressed broilers. In conclusion, dietary Gln improved the resistance of heat-stressed broiler muscles to oxidative damage possibly through reversing the muscle Gln level and inducing the expression of the Nrf2-Keap1 pathway.


Assuntos
Galinhas/fisiologia , Glutamina/metabolismo , Transtornos de Estresse por Calor/veterinária , Músculo Esquelético/efeitos dos fármacos , Estresse Oxidativo/efeitos dos fármacos , Transdução de Sinais , Ração Animal/análise , Animais , Proteínas Aviárias/metabolismo , Dieta/veterinária , Suplementos Nutricionais/análise , Relação Dose-Resposta a Droga , Feminino , Glutamina/administração & dosagem , Transtornos de Estresse por Calor/tratamento farmacológico , Proteína 1 Associada a ECH Semelhante a Kelch/metabolismo , Masculino , Carne/análise , Músculo Esquelético/metabolismo , Fator 2 Relacionado a NF-E2/metabolismo , Distribuição Aleatória , Coxa da Perna
16.
Mol Carcinog ; 59(4): 399-411, 2020 04.
Artigo em Inglês | MEDLINE | ID: mdl-32027051

RESUMO

Exploiting metabolic vulnerabilities of cancer cells with nontoxic, plant derived compounds constitutes a novel strategy for both chemoprevention and treatment. A high-throughput screening approach was used to evaluate a library of natural products to determine the most synergistic combination in precursor-B cell acute lymphoblast leukemia. Dimethylaminoparthenolide and shikonin effectively inhibited proliferation resulting in cell death in primary and immortalized leukemia cells, while having negligible effects on normal cells. Dimethylaminoparthenolide and shikonin have been shown separately to inhibit cell survival and proliferative signaling and activate tumor suppressors and proapoptotic pathways. Untargeted metabolomics and metabolic flux analysis with stable isotopically labeled glucose and glutamine exhibited a global shift in metabolism following treatment. Pathway analysis indicated significant differences in amino acid, antioxidant, tricarboxylic acid cycle, and nucleotide metabolism. Together, dimethylaminoparthenolide and shikonin reduced the shunting of glycolytic intermediates into the pentose phosphate pathway for biosynthetic purposes. Similarly, the incorporation of glutamine and glutamine-derived metabolites into purine and pyrimidine synthesis was inhibited by the combination of dimethylaminoparthenolide and shikonin, effectively impeding biosynthetic pathways critical for leukemia cell survival. This approach demonstrates that a synergistic pair of compounds with malignant cell specificity can effectively target metabolic pathways crucial to leukemia cell proliferation and induce apoptosis.


Assuntos
Proliferação de Células/efeitos dos fármacos , Naftoquinonas/farmacologia , Leucemia-Linfoma Linfoblástico de Células Precursoras B/metabolismo , Sesquiterpenos/farmacologia , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Criança , Ciclo do Ácido Cítrico/efeitos dos fármacos , Ensaios de Seleção de Medicamentos Antitumorais/métodos , Sinergismo Farmacológico , Glucose/metabolismo , Glutamina/metabolismo , Glicólise/efeitos dos fármacos , Humanos , Redes e Vias Metabólicas/efeitos dos fármacos , Leucemia-Linfoma Linfoblástico de Células Precursoras B/patologia
17.
Clin Interv Aging ; 15: 185-193, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32103921

RESUMO

Background: Alzheimer's disease is a devastating neurodegenerative disorder. Its worldwide prevalence is over 24 million and is expected to double by 2040. Finding ways to prevent its cognitive decline is urgent. Methods: A two-sample Mendelian randomization study was performed instrumenting glutamine, which is abundant in blood, capable of crossing the blood-brain barrier, and involved in a metabolic cycle with glutamate in the brain. Results: The results reveal a protective effect of circulating glutamine against Alzheimer's disease (inverse-variance weighted method, odds ratio per 1-standard deviation increase in circulating glutamine = 0.83; 95% CI 0.71, 0.97; P = 0.02). Conclusion: These findings lend credence to the emerging story supporting the modifiability of glutamine/glutamate metabolism for the prevention of cognitive decline. More circulating glutamine might mean that more substrate is available during times of stress, acting as a neuroprotectant. Modifications to exogenous glutamine may be worth exploring in future efforts to prevent and/or treat Alzheimer's disease.


Assuntos
Doença de Alzheimer , Barreira Hematoencefálica/fisiologia , Encéfalo/metabolismo , Disfunção Cognitiva/prevenção & controle , Glutamina , Idoso , Doença de Alzheimer/sangue , Doença de Alzheimer/metabolismo , Cognição/fisiologia , Feminino , Estudo de Associação Genômica Ampla/estatística & dados numéricos , Glutamina/sangue , Glutamina/genética , Glutamina/metabolismo , Humanos , Masculino , Análise da Randomização Mendeliana/métodos , Fármacos Neuroprotetores/metabolismo , Receptores de Glutamato/metabolismo
18.
J Biosci Bioeng ; 129(6): 672-678, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-32088137

RESUMO

l-Asparaginases have the potential to inhibit the formation of acrylamide, a harmful toxin formed during high temperature processing of food. A novel bacterium which produces l-asparaginase was screened. Type I l-asparaginase gene from Acinetobacter soli was cloned and expressed in Escherichia coli. The recombinant l-asparaginase had an activity of 42.0 IU mL-1 and showed no activity toward l-glutamine and d-asparagine. The recombinant l-asparaginase exhibited maximum catalytic activity at pH 8.0 and 40°C. The enzyme was stable in the pH ranging from 6.0 to 9.0. The activity of the recombinant enzyme was substantially enhanced by Ba2+, dithiothreitol, and ß-mercaptoethanol. The Km and Vmax values of the l-asparaginase for the l-asparagine were 3.22 mmol L-1 and 1.55 IU µg-1, respectively. Moreover, the recombinant l-asparaginase had the ability to mitigate acrylamide formation in potato chips. Compared with the untreated group, the content of acrylamide in samples treated with the enzyme was effectively decreased by 55.9%. These results indicate that the novel type I l-asparaginase has the potential for application in the food processing industry.


Assuntos
Acinetobacter/enzimologia , Acrilamida/metabolismo , Asparaginase/metabolismo , Solanum tuberosum/metabolismo , Acinetobacter/genética , Asparaginase/genética , Asparagina/metabolismo , Estabilidade Enzimática , Escherichia coli/genética , Escherichia coli/metabolismo , Glutamina/metabolismo , Lanches
19.
Genes Dev ; 34(7-8): 544-559, 2020 04 01.
Artigo em Inglês | MEDLINE | ID: mdl-32079653

RESUMO

Excessive reactive oxygen species (ROS) can cause oxidative stress and consequently cell injury contributing to a wide range of diseases. Addressing the critical gaps in our understanding of the adaptive molecular events downstream ROS provocation holds promise for the identification of druggable metabolic vulnerabilities. Here, we unveil a direct molecular link between the activity of two estrogen-related receptor (ERR) isoforms and the control of glutamine utilization and glutathione antioxidant production. ERRα down-regulation restricts glutamine entry into the TCA cycle, while ERRγ up-regulation promotes glutamine-driven glutathione production. Notably, we identify increased ERRγ expression/activation as a hallmark of oxidative stress triggered by mitochondrial disruption or chemotherapy. Enhanced tumor antioxidant capacity is an underlying feature of human breast cancer (BCa) patients that respond poorly to treatment. We demonstrate that pharmacological inhibition of ERRγ with the selective inverse agonist GSK5182 increases antitumor efficacy of the chemotherapeutic paclitaxel on poor outcome BCa tumor organoids. Our findings thus underscore the ERRs as novel redox sensors and effectors of a ROS defense program and highlight the potential therapeutic advantage of exploiting ERRγ inhibitors for the treatment of BCa and other diseases where oxidative stress plays a central role.


Assuntos
Neoplasias da Mama/fisiopatologia , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Estresse Oxidativo , Espécies Reativas de Oxigênio/metabolismo , Receptores Estrogênicos/metabolismo , Transdução de Sinais/fisiologia , Animais , Antineoplásicos/farmacologia , Técnicas Biossensoriais , Neoplasias da Mama/tratamento farmacológico , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Glutamina/metabolismo , Glutationa/metabolismo , Humanos , Camundongos , Estresse Oxidativo/efeitos dos fármacos , Estresse Oxidativo/fisiologia , Paclitaxel/farmacologia , Receptores Estrogênicos/genética , Rotenona/farmacologia , Tamoxifeno/análogos & derivados , Tamoxifeno/farmacologia
20.
Nat Commun ; 11(1): 16, 2020 01 07.
Artigo em Inglês | MEDLINE | ID: mdl-31911602

RESUMO

NAD+ synthetase is an essential enzyme of de novo and recycling pathways of NAD+ biosynthesis in Mycobacterium tuberculosis but not in humans. This bifunctional enzyme couples the NAD+ synthetase and glutaminase activities through an ammonia tunnel but free ammonia is also a substrate. Here we show that the Homo sapiens NAD+ synthetase (hsNadE) lacks substrate specificity for glutamine over ammonia and displays a modest activation of the glutaminase domain compared to tbNadE. We report the crystal structures of hsNadE and NAD+ synthetase from M. tuberculosis (tbNadE) with synthetase intermediate analogues. Based on the observed exclusive arrangements of the domains and of the intra- or inter-subunit tunnels we propose a model for the inter-domain communication mechanism for the regulation of glutamine-dependent activity and NH3 transport. The structural and mechanistic comparison herein reported between hsNadE and tbNadE provides also a starting point for future efforts in the development of anti-TB drugs.


Assuntos
Amida Sintases/metabolismo , Amônia/metabolismo , Proteínas de Bactérias/metabolismo , Carbono-Nitrogênio Ligases com Glutamina como Doadora de N-Amida/metabolismo , Mycobacterium tuberculosis/enzimologia , Amida Sintases/química , Amida Sintases/genética , Proteínas de Bactérias/química , Proteínas de Bactérias/genética , Carbono-Nitrogênio Ligases com Glutamina como Doadora de N-Amida/química , Carbono-Nitrogênio Ligases com Glutamina como Doadora de N-Amida/genética , Domínio Catalítico , Glutaminase/química , Glutaminase/genética , Glutaminase/metabolismo , Glutamina/metabolismo , Humanos , Mycobacterium tuberculosis/química , Mycobacterium tuberculosis/genética , Mycobacterium tuberculosis/metabolismo , NAD/metabolismo , Especificidade por Substrato
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