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1.
Molecules ; 26(19)2021 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-34641501

RESUMO

Diabetes mellitus is a global threat affecting millions of people of different age groups. In recent years, the development of naturally derived anti-diabetic agents has gained popularity. Okra is a common vegetable containing important bioactive components such as abscisic acid (ABA). ABA, a phytohormone, has been shown to elicit potent anti-diabetic effects in mouse models. Keeping its anti-diabetic potential in mind, in silico study was performed to explore its role in inhibiting proteins relevant to diabetes mellitus- 11ß-hydroxysteroid dehydrogenase (11ß-HSD1), aldose reductase, glucokinase, glutamine-fructose-6-phosphate amidotransferase (GFAT), peroxisome proliferator-activated receptor-gamma (PPAR-gamma), and Sirtuin family of NAD(+)-dependent protein deacetylases 6 (SIRT6). A comparative study of the ABA-protein docked complex with already known inhibitors of these proteins relevant to diabetes was compared to explore the inhibitory potential. Calculation of molecular binding energy (ΔG), inhibition constant (pKi), and prediction of pharmacokinetics and pharmacodynamics properties were performed. The molecular docking investigation of ABA with 11-HSD1, GFAT, PPAR-gamma, and SIRT6 revealed considerably low binding energy (ΔG from -8.1 to -7.3 Kcal/mol) and predicted inhibition constant (pKi from 6.01 to 5.21 µM). The ADMET study revealed that ABA is a promising drug candidate without any hazardous effect following all current drug-likeness guidelines such as Lipinski, Ghose, Veber, Egan, and Muegge.


Assuntos
Abelmoschus/química , Ácido Abscísico/farmacologia , Diabetes Mellitus/metabolismo , Hipoglicemiantes/farmacologia , Proteínas/metabolismo , 11-beta-Hidroxiesteroide Desidrogenase Tipo 1/antagonistas & inibidores , 11-beta-Hidroxiesteroide Desidrogenase Tipo 1/química , 11-beta-Hidroxiesteroide Desidrogenase Tipo 1/metabolismo , Ácido Abscísico/química , Ácido Abscísico/metabolismo , Ácido Abscísico/farmacocinética , Aldeído Redutase/química , Aldeído Redutase/metabolismo , Simulação por Computador , Inibidores Enzimáticos/química , Inibidores Enzimáticos/farmacologia , Glucoquinase/química , Glucoquinase/metabolismo , Glutamina/metabolismo , Glutamina-Frutose-6-Fosfato Transaminase (Isomerizante)/metabolismo , Quinase 3 da Glicogênio Sintase/química , Quinase 3 da Glicogênio Sintase/metabolismo , Humanos , Hipoglicemiantes/química , Simulação de Acoplamento Molecular , PPAR gama/química , PPAR gama/metabolismo , Proteínas/química , Sirtuínas/química , Sirtuínas/metabolismo
2.
Free Radic Biol Med ; 176: 335-344, 2021 11 20.
Artigo em Inglês | MEDLINE | ID: mdl-34634441

RESUMO

Whether from known or unknown causes, loss of epithelial repair plays a central role in the pathogenesis of pulmonary fibrosis. Recently, diminished mitochondrial function has been implicated as a factor contributing to the loss of epithelial repair but the mechanisms mediating these changes have not been defined. Here, we investigated the factors contributing to mitochondrial respiratory dysfunction after bleomycin, a widely accepted agent for modeling pulmonary fibrosis in mice and in vitro systems. In agreement with previous reports, we found that mitochondrial respiration was decreased in lung epithelial cells exposed to bleomycin, but also observed that responses differed depending on the type of metabolic fuel available to cells. For example, we found that mitochondrial respiration was dramatically reduced when glucose served as the primary fuel. Moreover, this associated with a marked decrease in glucose uptake, expression of glucose uptake transport 1 and capacity to augment glycolysis to either glucose or oligomycin. Conversely, mitochondrial respiration was largely preserved if glutamine was present in culture medium. The addition of glutamine also led to increased intracellular metabolite levels, including multiple TCA cycle intermediates and the glycolytic intermediate lactate, as well as reduced DNA damage and cell death to bleomycin. Taken together, these findings indicate that glutamine, rather than glucose, supports mitochondrial respiration and metabolite production in injured lung epithelial cells, and suggest that this shift away from glucose utilization serves to protect the lung epithelium from injury.


Assuntos
Bleomicina , Glutamina , Animais , Bleomicina/toxicidade , Células Epiteliais/metabolismo , Glucose/metabolismo , Glutamina/metabolismo , Glicólise , Camundongos , Mitocôndrias/metabolismo , Respiração
3.
Mol Cell ; 81(18): 3878-3878.e1, 2021 09 16.
Artigo em Inglês | MEDLINE | ID: mdl-34547243

RESUMO

Metabolic networks support cancer cell survival, proliferation, and malignant progression. Cancer cells take up large amounts of nutrients such as glucose and glutamine whose metabolism provides the energy, reducing equivalents, and biosynthetic precursors required to meet the biosynthetic demands of proliferation. Intermediates of glycolysis and the tricarboxylic acid (TCA) cycle provide critical building blocks for synthesis of non-essential amino acids, nucleotides, and fatty acids. To view this SnapShot, open or download the PDF.


Assuntos
Redes e Vias Metabólicas/fisiologia , Neoplasias/metabolismo , Aminoácidos/metabolismo , Ciclo do Ácido Cítrico/fisiologia , Metabolismo Energético , Glucose/metabolismo , Glutamina/metabolismo , Glicólise/fisiologia , Humanos , Nucleotídeos/metabolismo
4.
Cells ; 10(9)2021 09 08.
Artigo em Inglês | MEDLINE | ID: mdl-34571997

RESUMO

Acetylation is a post-translational modification that regulates the activity of enzymes fundamentally involved in cellular and mitochondrial bioenergetic metabolism. NAD+ dependent deacetylase sirtuin 3 (SIRT3) is localized to mitochondria where it plays a key role in regulating acetylation of TCA cycle enzymes and the mitochondrial respiratory complexes. Although the SIRT3 target proteins in mitochondria have been identified, the effect of SIRT3 activity on mitochondrial glucose metabolism in the brain remains elusive. The impact of abolished SIRT3 activity on glucose metabolism was determined in SIRT3 knockout (KO) and wild type (WT) mice injected with [1,6-13C]glucose using ex vivo 13C-NMR spectroscopy. The 1H-NMR spectra and amino acid analysis showed no differences in the concentration of lactate, glutamate, alanine, succinate, or aspartate between SIRT3 KO and WT mice. However, glutamine, total creatine (Cr), and GABA were lower in SIRT3 KO brain. Incorporation of label from [1,6-13C]glucose metabolism into lactate or alanine was not affected in SIRT3 KO brain. However, the incorporation of the label into all isotopomers of glutamate, glutamine, GABA and aspartate was lower in SIRT3 KO brain, reflecting decreased activity of mitochondrial and TCA cycle metabolism in both neurons and astrocytes. This is most likely due to hyperacetylation of mitochondrial enzymes due to suppressed SIRT3 activity in the brain of SIRT3 KO mice. Thus, the absence of Sirt3 results in impaired mitochondrial oxidative energy metabolism and neurotransmitter synthesis in the brain. Since the SIRT3 activity is NAD+ dependent, these results might parallel changes in glucose metabolism under pathologic reduction in mitochondrial NAD+ pools.


Assuntos
Encéfalo/metabolismo , Metabolismo dos Carboidratos/fisiologia , Glucose/metabolismo , Sirtuína 3/metabolismo , Acetilação , Animais , Astrócitos/metabolismo , Metabolismo Energético/fisiologia , Feminino , Ácido Glutâmico/metabolismo , Glutamina/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Mitocôndrias/metabolismo , Neurônios/metabolismo , Neurotransmissores/metabolismo , Processamento de Proteína Pós-Traducional/fisiologia
5.
Int J Mol Sci ; 22(18)2021 Sep 10.
Artigo em Inglês | MEDLINE | ID: mdl-34575972

RESUMO

Glutamine and lipids are two important components of proliferating cancer cells. Studies have demonstrated that glutamine synthetase (GS) boosts glutamine-dependent anabolic processes for nucleotide and protein synthesis, but the role of GS in regulating lipogenesis remains unclear. This study identified that insulin and glutamine deprivation activated the lipogenic transcription factor sterol regulatory element-binding protein 1 (SREBP1) that bound to the GS promoter and increased its transcription. Notably, GS enhanced the O-linked N-acetylglucosaminylation (O-GlcNAcylation) of the specificity protein 1 (Sp1) that induced SREBP1/acetyl-CoA carboxylase 1 (ACC1) expression resulting in lipid droplet (LD) accumulation upon insulin treatment. Moreover, glutamine deprivation induced LD formation through GS-mediated O-GlcNAc-Sp1/SREBP1/ACC1 signaling and supported cell survival. These findings demonstrate that insulin and glutamine deprivation induces SREBP1 that transcriptionally activates GS, resulting in Sp1 O-GlcNAcylation. Subsequently, O-GlcNAc-Sp1 transcriptionally upregulates the expression of SREBP1, resulting in a feedforward loop that increases lipogenesis and LD formation in liver and breast cancer cells.


Assuntos
Acetil-CoA Carboxilase/genética , Glutamato-Amônia Ligase/genética , Neoplasias Hepáticas/genética , Fator de Transcrição Sp1/genética , Proteína de Ligação a Elemento Regulador de Esterol 1/genética , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Proliferação de Células/genética , Regulação Neoplásica da Expressão Gênica/genética , Glutamina/metabolismo , Humanos , Insulina/metabolismo , Lipídeos/genética , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/patologia , Metabolismo/genética , Regiões Promotoras Genéticas/genética , Biossíntese de Proteínas/genética , Transdução de Sinais , beta-N-Acetil-Hexosaminidases/genética
6.
Theranostics ; 11(16): 7844-7868, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34335968

RESUMO

Radiotherapy is one of the curative treatment options for localized prostate cancer (PCa). The curative potential of radiotherapy is mediated by irradiation-induced oxidative stress and DNA damage in tumor cells. However, PCa radiocurability can be impeded by tumor resistance mechanisms and normal tissue toxicity. Metabolic reprogramming is one of the major hallmarks of tumor progression and therapy resistance. Specific metabolic features of PCa might serve as therapeutic targets for tumor radiosensitization and as biomarkers for identifying the patients most likely to respond to radiotherapy. The study aimed to characterize a potential role of glutaminase (GLS)-driven glutamine catabolism as a prognostic biomarker and a therapeutic target for PCa radiosensitization. Methods: We analyzed primary cell cultures and radioresistant (RR) derivatives of the conventional PCa cell lines by gene expression and metabolic assays to identify the molecular traits associated with radiation resistance. Relative radiosensitivity of the cell lines and primary cell cultures were analyzed by 2-D and 3-D clonogenic analyses. Targeting of glutamine (Gln) metabolism was achieved by Gln starvation, gene knockdown, and chemical inhibition. Activation of the DNA damage response (DDR) and autophagy was assessed by gene expression, western blotting, and fluorescence microscopy. Reactive oxygen species (ROS) and the ratio of reduced glutathione (GSH) to oxidized glutathione (GSSG) were analyzed by fluorescence and luminescence probes, respectively. Cancer stem cell (CSC) properties were investigated by sphere-forming assay, CSC marker analysis, and in vivo limiting dilution assays. Single circulating tumor cells (CTCs) isolated from the blood of PCa patients were analyzed by array comparative genome hybridization. Expression levels of the GLS1 and MYC gene in tumor tissues and amino acid concentrations in blood plasma were correlated to a progression-free survival in PCa patients. Results: Here, we found that radioresistant PCa cells and prostate CSCs have a high glutamine demand. GLS-driven catabolism of glutamine serves not only for energy production but also for the maintenance of the redox state. Consequently, glutamine depletion or inhibition of critical regulators of glutamine utilization, such as GLS and the transcription factor MYC results in PCa radiosensitization. On the contrary, we found that a combination of glutamine metabolism inhibitors with irradiation does not cause toxic effects on nonmalignant prostate cells. Glutamine catabolism contributes to the maintenance of CSCs through regulation of the alpha-ketoglutarate (α-KG)-dependent chromatin-modifying dioxygenase. The lack of glutamine results in the inhibition of CSCs with a high aldehyde dehydrogenase (ALDH) activity, decreases the frequency of the CSC populations in vivo and reduces tumor formation in xenograft mouse models. Moreover, this study shows that activation of the ATG5-mediated autophagy in response to a lack of glutamine is a tumor survival strategy to withstand radiation-mediated cell damage. In combination with autophagy inhibition, the blockade of glutamine metabolism might be a promising strategy for PCa radiosensitization. High blood levels of glutamine in PCa patients significantly correlate with a shorter prostate-specific antigen (PSA) doubling time. Furthermore, high expression of critical regulators of glutamine metabolism, GLS1 and MYC, is significantly associated with a decreased progression-free survival in PCa patients treated with radiotherapy. Conclusions: Our findings demonstrate that GLS-driven glutaminolysis is a prognostic biomarker and therapeutic target for PCa radiosensitization.


Assuntos
Glutamina/metabolismo , Neoplasias da Próstata/metabolismo , Tolerância a Radiação/genética , Animais , Autofagia , Proteína 5 Relacionada à Autofagia/metabolismo , Biomarcadores Farmacológicos , Linhagem Celular Tumoral , Glutaminase/antagonistas & inibidores , Glutaminase/genética , Glutaminase/metabolismo , Humanos , Masculino , Camundongos Nus , Células-Tronco Neoplásicas/metabolismo , Oxirredução , Proteínas Proto-Oncogênicas c-myc/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Ensaios Antitumorais Modelo de Xenoenxerto
7.
Molecules ; 26(16)2021 Aug 10.
Artigo em Inglês | MEDLINE | ID: mdl-34443413

RESUMO

Breast cancer is one of the most prevalent cancers worldwide usually treated with Tamoxifen. Tamoxifen resistance development is the most challenging issue in an initially responsive breast tumor, and mechanisms of resistance are still under investigation. The objective of this study is to develop and validate a selective, sensitive, and simultaneous high performance liquid chromatography-tandem mass spectrometry method to explore the changes in substrates and metabolites in supernatant media of developed Tamoxifen resistance MCF-7 cells. We focus on the determination of lactate, pyruvate, and L-glutamine which enables the tracking of changes in metabolic pathways as a result of the resistance process. Chromatographic separation was achieved within 3.5 min. using a HILIC column (4.6 × 100 mm, 3.5 µm particle size) and mobile phase of 0.05 M acetic acid-ammonium acetate buffer solution pH 3.0: Acetonitrile (40:60 v/v). The linear range was 0.11-2.25, 0.012-0.227, and 0.02-0.20 mM for lactate, pyruvate, and L-glutamine, respectively. Within- and between-run accuracy was in the range 98.94-105.50% with precision (CV, %) of ≤0.86%. The results revealed a significant increase in both lactate and pyruvate production after acquiring the resistant. An increase in L-glutamine levels was also observed and could be attributed to its over production or decline in its consumption. Therefore, further tracking of genes responsible of lactate, pyruvate, and glutamine metabolic pathways should be performed in parallel to provide in-depth explanation of resistance mechanism.


Assuntos
Resistencia a Medicamentos Antineoplásicos , Glutamina/metabolismo , Ácido Láctico/metabolismo , Tamoxifeno/farmacologia , Espectrometria de Massas em Tandem , Calibragem , Contagem de Células , Forma Celular/efeitos dos fármacos , Cromatografia Líquida de Alta Pressão , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Humanos , Células MCF-7 , Ácido Pirúvico/metabolismo , Reprodutibilidade dos Testes
8.
Int J Mol Sci ; 22(16)2021 Aug 09.
Artigo em Inglês | MEDLINE | ID: mdl-34445264

RESUMO

The multidrug efflux transporter ABCB1 is clinically important for drug absorption and distribution and can be a determinant of chemotherapy failure. Recent structure data shows that three glutamines donate hydrogen bonds to coordinate taxol in the drug binding pocket. This is consistent with earlier drug structure-activity relationships that implicated the importance of hydrogen bonds in drug recognition by ABCB1. By replacing the glutamines with alanines we have tested whether any, or all, of Gln347, Gln725, and Gln990 are important for the transport of three different drug classes. Flow cytometric transport assays show that Q347A and Q990A act synergistically to reduce transport of Calcein-AM, BODIPY-verapamil, and OREGON GREEN-taxol bisacetate but the magnitude of the effect was dependent on the test drug and no combination of mutations completely abrogated function. Surprisingly, Q725A mutants generally improved transport of Calcein-AM and BODIPY-verapamil, suggesting that engagement of the wild-type Gln725 in a hydrogen bond is inhibitory for the transport mechanism. To test transport of unmodified taxol, stable expression of Q347/725A and the triple mutant was engineered and shown to confer equivalent resistance to the drug as the wild-type transporter, further indicating that none of these potential hydrogen bonds between transporter and transport substrate are critical for the function of ABCB1. The implications of the data for plasticity of the drug binding pocket are discussed.


Assuntos
Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Paclitaxel/farmacologia , Subfamília B de Transportador de Cassetes de Ligação de ATP/genética , Subfamília B de Transportador de Cassetes de Ligação de ATP/metabolismo , Substituição de Aminoácidos , Resistencia a Medicamentos Antineoplásicos/genética , Glutamina/genética , Glutamina/metabolismo , Células HEK293 , Humanos , Mutação de Sentido Incorreto
9.
Nat Commun ; 12(1): 4814, 2021 08 10.
Artigo em Inglês | MEDLINE | ID: mdl-34376668

RESUMO

Glutamoptosis is the induction of apoptotic cell death as a consequence of the aberrant activation of glutaminolysis and mTORC1 signaling during nutritional imbalance in proliferating cells. The role of the bioenergetic sensor AMPK during glutamoptosis is not defined yet. Here, we show that AMPK reactivation blocks both the glutamine-dependent activation of mTORC1 and glutamoptosis in vitro and in vivo. We also show that glutamine is used for asparagine synthesis and the GABA shunt to produce ATP and to inhibit AMPK, independently of glutaminolysis. Overall, our results indicate that glutamine metabolism is connected with mTORC1 activation through two parallel pathways: an acute alpha-ketoglutarate-dependent pathway; and a secondary ATP/AMPK-dependent pathway. This dual metabolic connection between glutamine and mTORC1 must be considered for the future design of therapeutic strategies to prevent cell growth in diseases such as cancer.


Assuntos
Apoptose/fisiologia , Proliferação de Células/fisiologia , Glutamina/metabolismo , Alvo Mecanístico do Complexo 1 de Rapamicina/metabolismo , Transdução de Sinais/fisiologia , Proteínas Quinases Ativadas por AMP/metabolismo , Trifosfato de Adenosina/metabolismo , Animais , Antineoplásicos/farmacologia , Linhagem Celular Tumoral , Células HCT116 , Células HEK293 , Humanos , Masculino , Camundongos Endogâmicos NOD , Camundongos Knockout , Camundongos SCID , Sirolimo/análogos & derivados , Sirolimo/farmacologia , Ensaios Antitumorais Modelo de Xenoenxerto/métodos
10.
Curr Top Med Chem ; 21(11): 985-994, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34355684

RESUMO

BACKGROUND: Although Autism Spectrum Disorder (ASD) is considered a heterogeneous neurological disease in childhood, a growing body of evidence associates it with mitochondrial dysfunction explaining the observed comorbidities. INTRODUCTION: The aim of this study is to identify variations in cellular bioenergetics and metabolism dependent on mitochondrial function in ASD patients and healthy controls using Peripheral Blood Mononuclear Cells (PBMCs). We hypothesized that PBMCs may reveal the cellular pathology and provide evidence of bioenergetic and metabolic changes accompanying the disease. METHODS: PBMC from children with ASD and a control group of the same age and gender were isolated. All patients underwent an in-depth clinical evaluation. A well-characterized cohort of Bulgarian children is selected. Bioenergetic and metabolic studies of isolated PBMCs are performed with a Seahorse XFp analyzer. RESULTS: Our data show that PBMCs from patients with ASD have increased respiratory reserve capacity (by 27.5%), increased maximal respiration (by 67%) and altered adaptive response to oxidative stress induced by DMNQ. In addition, we demonstrate а strong dependence on fatty acids and impaired ability to reprogram cell metabolism. The listed characteristics are not observed in the control group. These results can contribute to a better understanding of the underlying causes of ASD, which is crucial for selecting a successful treatment. CONCLUSION: The current study, for the first time, provides a functional analysis of cell bioenergetics and metabolic changes in a group of Bulgarian patients with ASD. It reveals physiological abnormalities that do not allow mitochondria to adapt and meet the increased energetic requirements of the cell. The link between mitochondria and ASD is not yet fully understood, but this may lead to the discovery of new approaches for nutrition and therapy.


Assuntos
Transtorno do Espectro Autista/fisiopatologia , Metabolismo Energético/fisiologia , Leucócitos Mononucleares/metabolismo , Metaboloma/fisiologia , Mitocôndrias/metabolismo , Criança , Pré-Escolar , Ácidos Graxos/metabolismo , Glucose/metabolismo , Glutamina/metabolismo , Humanos , Mitocôndrias/ultraestrutura , Estresse Oxidativo , Respiração
11.
Curr Top Med Chem ; 21(11): 949-963, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34355686

RESUMO

Major Depressive Disorder (MDD) and Bipolar Disorder (BD) have a high prevalence and detrimental socio-economic consequences for the patients and the community. Furthermore, the depressive symptomatology of both disorders is essentially identical, thus rendering the clinical differential diagnosis between the two significantly more difficult considering the concomitant lack of objective biomarkers. Mood disorders are multifactorial disorders the pathophysiology of which includes genetic, epigenetic, neurobiological, neuroimmunological, structural and functional brain alterations, etc. Aberrant genetic variants as well as changed differential expression of microRNAs (miRNAs) and long non-coding RNAs (lncRNAs) have been implicated in the pathophysiology of MDD and BD. MiRNAs as well as lncRNAs have regulatory and modulating functions on protein-- coding gene expression thus influencing the remodeling of the architecture, neurotransmission, immunomodulation, etc. in the Central Nervous System (CNS) which are essential in the development of psychiatric disorders including MDD and BD. Moreover, both shared and distinct structural, connectivity, task-related and metabolic features have been observed via functional magnetic resonance imaging and magnetic resonance spectroscopy, suggesting the possibility of a dimensional continuum between the two disorders instead of a categorical differentiation. Aberrant connectivity within and between the Default Mode Network, the Salience Network, Executive Network, etc. as well as dysfunctional emotion, cognitive and executive processing have been associated with mood disorders. Therefore, the aim of this review is to explore a more multidimensional framework in the scientific research of mood disorders, including epigenetic and neuroimaging data in order to shape an outline for their translational capacity in clinical practice.


Assuntos
Biomarcadores/análise , Imageamento por Ressonância Magnética/métodos , Transtornos do Humor/diagnóstico , RNA não Traduzido/análise , Biomarcadores/metabolismo , Transtorno Bipolar/diagnóstico , Transtorno Bipolar/genética , Transtorno Bipolar/fisiopatologia , Encéfalo/fisiopatologia , Transtorno Depressivo Maior/diagnóstico , Transtorno Depressivo Maior/fisiopatologia , Regulação da Expressão Gênica , Ácido Glutâmico/metabolismo , Glutamina/metabolismo , Humanos , Espectroscopia de Ressonância Magnética , MicroRNAs/análise , MicroRNAs/metabolismo , Transtornos do Humor/genética , Transtornos do Humor/fisiopatologia , RNA não Traduzido/metabolismo
12.
Dev Cell ; 56(16): 2329-2347.e6, 2021 08 23.
Artigo em Inglês | MEDLINE | ID: mdl-34428399

RESUMO

Mammalian preimplantation embryos follow a stereotypic pattern of development from zygotes to blastocysts. Here, we use labeled nutrient isotopologue analysis of small numbers of embryos to track downstream metabolites. Combined with transcriptomic analysis, we assess the capacity of the embryo to reprogram its metabolism through development. Early embryonic metabolism is rigid in its nutrient requirements, sensitive to reductive stress and has a marked disequilibrium between two halves of the TCA cycle. Later, loss of maternal LDHB and transcription of zygotic products favors increased activity of bioenergetic shuttles, fatty-acid oxidation and equilibration of the TCA cycle. As metabolic plasticity peaks, blastocysts can develop without external nutrients. Normal developmental metabolism of the early embryo is distinct from cancer metabolism. However, similarities emerge upon reductive stress. Increased metabolic plasticity with maturation is due to changes in redox control mechanisms and to transcriptional reprogramming of later-stage embryos during homeostasis or upon adaptation to environmental changes.


Assuntos
Adaptação Fisiológica , Blastocisto/metabolismo , Metaboloma , Animais , Células Cultivadas , Ciclo do Ácido Cítrico , Glucose/metabolismo , Glutamina/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , NAD/metabolismo , Oxirredução , Transcriptoma
13.
Cells ; 10(7)2021 07 13.
Artigo em Inglês | MEDLINE | ID: mdl-34359941

RESUMO

Cancer stem cells (CSCs) are heterogeneous cells with stem cell-like properties that are responsible for therapeutic resistance, recurrence, and metastasis, and are the major cause for cancer treatment failure. Since CSCs have distinct metabolic characteristics that plays an important role in cancer development and progression, targeting metabolic pathways of CSCs appears to be a promising therapeutic approach for cancer treatment. Here we classify and discuss the unique metabolisms that CSCs rely on for energy production and survival, including mitochondrial respiration, glycolysis, glutaminolysis, and fatty acid metabolism. Because of metabolic plasticity, CSCs can switch between these metabolisms to acquire energy for tumor progression in different microenvironments compare to the rest of tumor bulk. Thus, we highlight the specific conditions and factors that promote or suppress CSCs properties to portray distinct metabolic phenotypes that attribute to CSCs in common cancers. Identification and characterization of the features in these metabolisms can offer new anticancer opportunities and improve the prognosis of cancer. However, the therapeutic window of metabolic inhibitors used alone or in combination may be rather narrow due to cytotoxicity to normal cells. In this review, we present current findings of potential targets in these four metabolic pathways for the development of more effective and alternative strategies to eradicate CSCs and treat cancer more effectively in the future.


Assuntos
Redes e Vias Metabólicas , Terapia de Alvo Molecular , Células-Tronco Neoplásicas/metabolismo , Animais , Glutamina/metabolismo , Humanos , Mitocôndrias/metabolismo , Fosforilação Oxidativa
14.
Plant Sci ; 310: 110963, 2021 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-34315588

RESUMO

Ammonium (NH4+) toxicity has become a serious ecological and agricultural issue owing to increasing soil nitrogen inputs and atmospheric nitrogen deposition. There is accumulating evidence for the mechanisms underlying NH4+-tolerance in rice and Arabidopsis, but similar knowledge for dryland crops is currently limited. We investigated the responses of a natural population of allotetraploid rapeseed to NH4+ and nitrate (NO3-) and screened one NH4+-tolerant genotype (T5) and one NH4+-sensitive genotype (S211). Determination of the shoot and root NH4+ concentrations showed that levels were higher in S211 than in T5. 15NH4+ uptake assays, glutamine synthetase (GS) activity quantification, and relative gene transcriptional analysis indicated that the significantly higher GS activity observed in T5 roots than that in S211 was the main reason for its NH4+-tolerance. In-depth metabolomic analysis verified that Gln metabolism plays an important role in rapeseed NH4+-tolerance. Furthermore, adaptive changes in carbon metabolism were much more active in T5 shoots than in S211. Interestingly, we found that N-glycosylation pathway was significantly induced by NH4+, especially the mannose metabolism, which concentration was 2.75-fold higher in T5 shoots than in S211 with NH4+ treatment, indicating that mannose may be a metabolomic marker which also confers physiological adaptations for NH4+ tolerance in rapeseed. The corresponding amino acid and soluble sugar concentrations and gene expression in T5 and S211 were consistent with these results. Genomic sequencing identified variations in the GLN (encoding GS) and GMP1 (encoding the enzyme that provides GDP-mannose) gene families between the T5 and S211 lines. These genes will be utilized as candidate genes for future investigations of the molecular mechanisms underlying NH4+ tolerance in rapeseed.


Assuntos
Compostos de Amônio/toxicidade , Glutamato-Amônia Ligase/metabolismo , Glutamina/metabolismo , Manose/metabolismo , Brassica napus/metabolismo , Regulação da Expressão Gênica de Plantas , Genótipo , Glutamato-Amônia Ligase/genética , Raízes de Plantas/genética , Raízes de Plantas/metabolismo
15.
Exp Eye Res ; 210: 108703, 2021 09.
Artigo em Inglês | MEDLINE | ID: mdl-34280391

RESUMO

Diabetic retinopathy (DR) is a vision-loss complication caused by diabetes with high prevalence. During DR, the retinal microvascular injury and neurodegeneration derived from chronic hyperglycemia have attracted global attention to retinal Müller cells (RMCs), the major macroglia in the retina contributes to neuroprotection. Protein Phosphatase 1 Catalytic Subunit Alpha (PPP1CA) dephosphorylates the transcriptional coactivator Yes-associated protein (YAP) to promote the transcription of glutamine synthetase (GS). GS catalyzes the transformation of neurotoxic glutamate (Glu) into nontoxic glutamine (Gln) to activate the mammalian target of rapamycin complex 1 (mTORC1), which promotes the activation of RMCs. In this study, in vitro MIO-M1 cell and in vivo mouse high-fat diet and streptozotocin (STZ)-induced diabetic model to explore the role of the PPP1CA/YAP/GS/Gln/mTORC1 pathway on the activation of MRCs during DR. Results showed that PPP1CA promoted the dephosphorylation and nuclear translocation of YAP in high glucose (HG)-exposed MIO-M1 cells. YAP transcribed GS in HG-exposed MIO-M1 cells in a TEAD1-dependent and PPP1CA-dependent way. GS promoted the biosynthesis of Gln in HG-exposed MIO-M1 cells. Gln activated mTORC1 instead of mTORC2 in HG-exposed MIO-M1 cells. The proliferation and activation of HG-exposed MIO-M1 cells were PPP1CA/YAP/GS/Gln/mTORC1-dependent. Finally, RMC proliferation and activation during DR were inhibited by the PPP1CA/YAP/GS/Gln/mTORC1 blockade. The findings supplied a potential idea to protect RMCs and alleviate the development of DR.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Retinopatia Diabética/metabolismo , Células Ependimogliais/metabolismo , Glutamato-Amônia Ligase/metabolismo , Glutamina/metabolismo , Alvo Mecanístico do Complexo 1 de Rapamicina/metabolismo , Proteína Fosfatase 1/metabolismo , Animais , Western Blotting , Proliferação de Células , Células Cultivadas , Diabetes Mellitus Experimental/metabolismo , Ensaio de Imunoadsorção Enzimática , Glucose/farmacologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Microscopia de Fluorescência , Reação em Cadeia da Polimerase em Tempo Real , Transdução de Sinais/fisiologia , Estreptozocina
16.
Sci Rep ; 11(1): 15558, 2021 07 30.
Artigo em Inglês | MEDLINE | ID: mdl-34330957

RESUMO

N-Acetylcysteine (NAC) is an antioxidant, anti-adhesive, and antimicrobial compound. Even though there is much information regarding the role of NAC as an antioxidant and anti-adhesive agent, little is known about its antimicrobial activity. In order to assess its mode of action in bacterial cells, we investigated the metabolic responses triggered by NAC at neutral pH. As a model organism, we chose the Gram-negative plant pathogen Xanthomonas citri subsp. citri (X. citri), the causal agent of citrus canker disease, due to the potential use of NAC as a sustainable molecule against phytopathogens dissemination in citrus cultivated areas. In presence of NAC, cell proliferation was affected after 4 h, but damages to the cell membrane were observed only after 24 h. Targeted metabolite profiling analysis using GC-MS/TOF unravelled that NAC seems to be metabolized by the cells affecting cysteine metabolism. Intriguingly, glutamine, a marker for nitrogen status, was not detected among the cells treated with NAC. The absence of glutamine was followed by a decrease in the levels of the majority of the proteinogenic amino acids, suggesting that the reduced availability of amino acids affect protein synthesis and consequently cell proliferation.


Assuntos
Acetilcisteína/metabolismo , Cromatografia Gasosa-Espectrometria de Massas/métodos , Metabolômica/métodos , Xanthomonas/metabolismo , Aminoácidos/metabolismo , Membrana Celular/metabolismo , Citrus/metabolismo , Glutamina/metabolismo
17.
J Cancer Res Clin Oncol ; 147(11): 3169-3181, 2021 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-34235580

RESUMO

PURPOSE: Glutamine plays an important role in cell viability and growth of various tumors. For the fetal subtype of hepatoblastoma, growth inhibition through glutamine depletion was shown. We studied glutamine depletion in embryonal cell lines of hepatoblastoma carrying different mutations. Since asparagine synthetase was identified as a prognostic factor and potential therapeutic target in adult hepatocellular carcinoma, we investigated the expression of its gene ASNS and of the gene GLUL, encoding for glutamine synthetase, in hepatoblastoma specimens and cell lines and investigated the correlation with overall survival. METHODS: We correlated GLUL and ASNS expression with overall survival using publicly available microarray and clinical data. We examined GLUL and ASNS expression by RT-qPCR and by Western blot analysis in the embryonal cell lines Huh-6 and HepT1, and in five hepatoblastoma specimens. In the same cell lines, we investigated the effects of glutamine depletion. Hepatoblastoma biopsies were examined for histology and CTNNB1 mutations. RESULTS: High GLUL expression was associated with a higher median survival time. Independent of mutations and histology, hepatoblastoma samples showed strong GLUL expression and glutamine synthesis. Glutamine depletion resulted in the inhibition of proliferation and of cell viability in both embryonal hepatoblastoma cell lines. ASNS expression did not correlate with overall survival. CONCLUSION: Growth inhibition resulting from glutamine depletion, as described for the hepatoblastoma fetal subtype, is also detected in established embryonal hepatoblastoma cell lines carrying different mutations. At variance with adult hepatocellular carcinoma, in hepatoblastoma asparagine synthetase has no prognostic significance.


Assuntos
Glutamato-Amônia Ligase/biossíntese , Glutamina/metabolismo , Hepatoblastoma/metabolismo , Neoplasias Hepáticas/metabolismo , Carbono-Nitrogênio Ligases com Glutamina como Doadora de N-Amida/biossíntese , Carbono-Nitrogênio Ligases com Glutamina como Doadora de N-Amida/genética , Linhagem Celular Tumoral , Sobrevivência Celular/fisiologia , Éxons , Expressão Gênica , Glutamato-Amônia Ligase/genética , Glutamina/deficiência , Hepatoblastoma/genética , Hepatoblastoma/patologia , Humanos , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/patologia , Mutação , beta Catenina/genética
18.
Sci Rep ; 11(1): 14161, 2021 07 08.
Artigo em Inglês | MEDLINE | ID: mdl-34239013

RESUMO

The integrated stress response (ISR) is a central cellular adaptive program that is activated by diverse stressors including ER stress, hypoxia and nutrient deprivation to orchestrate responses via activating transcription factor 4 (ATF4). We hypothesized that ATF4 is essential for the adaptation of human glioblastoma (GB) cells to the conditions of the tumor microenvironment and is contributing to therapy resistance against chemotherapy. ATF4 induction in GB cells was modulated pharmacologically and genetically and investigated in the context of temozolomide treatment as well as glucose and oxygen deprivation. The relevance of the ISR was analyzed by cell death and metabolic measurements under conditions to approximate aspects of the GB microenvironment. ATF4 protein levels were induced by temozolomide treatment. In line, ATF4 gene suppressed GB cells (ATF4sh) displayed increased cell death and decreased survival after temozolomide treatment. Similar results were observed after treatment with the ISR inhibitor ISRIB. ATF4sh and ISRIB treated GB cells were sensitized to hypoxia-induced cell death. Our experimental study provides evidence for an important role of ATF4 for the adaptation of human GB cells to conditions of the tumor microenvironment characterized by low oxygen and nutrient availability and for the development of temozolomide resistance. Inhibiting the ISR in GB cells could therefore be a promising therapeutic approach.


Assuntos
Fator 4 Ativador da Transcrição/metabolismo , Adaptação Fisiológica , Glioblastoma/tratamento farmacológico , Glioblastoma/metabolismo , Temozolomida/uso terapêutico , Hipóxia Tumoral , Acetamidas/farmacologia , Fator 4 Ativador da Transcrição/genética , Adaptação Fisiológica/efeitos dos fármacos , Morte Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Cicloexilaminas/farmacologia , Estresse do Retículo Endoplasmático/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Glioblastoma/genética , Glutamina/metabolismo , Humanos , Consumo de Oxigênio/efeitos dos fármacos , Estresse Fisiológico/efeitos dos fármacos , Estresse Fisiológico/genética , Temozolomida/farmacologia , Hipóxia Tumoral/efeitos dos fármacos
19.
J Biol Chem ; 297(3): 100954, 2021 09.
Artigo em Inglês | MEDLINE | ID: mdl-34270958

RESUMO

Peroxisome proliferator-activated receptor δ (PPARδ) is a nuclear receptor transcription factor that plays an important role in the regulation of metabolism, inflammation, and cancer. In addition, the nutrient-sensing kinase 5'AMP-activated protein kinase (AMPK) is a critical regulator of cellular energy in coordination with PPARδ. However, the molecular mechanism of the AMPK/PPARδ pathway on cancer progression is still unclear. Here, we found that activated AMPK induced PPARδ-S50 phosphorylation in cancer cells, whereas the PPARδ/S50A (nonphosphorylation mimic) mutant reversed this event. Further analysis showed that the PPARδ/S50E (phosphorylation mimic) but not the PPARδ/S50A mutant increased PPARδ protein stability, which led to reduced p62/SQSTM1-mediated degradation of misfolded PPARδ. Furthermore, PPARδ-S50 phosphorylation decreased PPARδ transcription activity and alleviated PPARδ-mediated uptake of glucose and glutamine in cancer cells. Soft agar and xenograft tumor model analysis showed that the PPARδ/S50E mutant but not the PPARδ/S50A mutant inhibited colon cancer cell proliferation and tumor growth, which was associated with inhibition of Glut1 and SLC1A5 transporter protein expression. These findings reveal a new mechanism of AMPK-induced PPARδ-S50 phosphorylation, accumulation of misfolded PPARδ protein, and inhibition of PPARδ transcription activity contributing to the suppression of colon tumor formation.


Assuntos
Adenilato Quinase/metabolismo , Proliferação de Células , Neoplasias do Colo/metabolismo , Neoplasias do Colo/patologia , Glucose/metabolismo , Glutamina/metabolismo , PPAR gama/metabolismo , Animais , Linhagem Celular Tumoral , Feminino , Xenoenxertos , Humanos , Camundongos Nus , Fosforilação
20.
Int J Mol Sci ; 22(11)2021 May 25.
Artigo em Inglês | MEDLINE | ID: mdl-34070527

RESUMO

Excessive proliferation and migration of vascular smooth muscle cells (VSMCs) contribute to the development of atherosclerosis and restenosis. Glycolysis and glutaminolysis are increased in rapidly proliferating VSMCs to support their increased energy requirements and biomass production. Thus, it is essential to develop new pharmacological tools that regulate metabolic reprogramming in VSMCs for treatment of atherosclerosis. The effects of 6-diazo-5-oxo-L-norleucine (DON), a glutamine antagonist, have been broadly investigated in highly proliferative cells; however, it is unclear whether DON inhibits proliferation of VSMCs and neointima formation. Here, we investigated the effects of DON on neointima formation in vivo as well as proliferation and migration of VSMCs in vitro. DON simultaneously inhibited FBS- or PDGF-stimulated glycolysis and glutaminolysis as well as mammalian target of rapamycin complex I activity in growth factor-stimulated VSMCs, and thereby suppressed their proliferation and migration. Furthermore, a DON-derived prodrug, named JHU-083, significantly attenuated carotid artery ligation-induced neointima formation in mice. Our results suggest that treatment with a glutamine antagonist is a promising approach to prevent progression of atherosclerosis and restenosis.


Assuntos
Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Diazo-Oxo-Norleucina/farmacologia , Glutamina/antagonistas & inibidores , Glicólise/efeitos dos fármacos , Músculo Liso Vascular/efeitos dos fármacos , Neointima/tratamento farmacológico , Fosforilação Oxidativa/efeitos dos fármacos , Animais , Antimetabólitos Antineoplásicos/farmacologia , Ciclo Celular/efeitos dos fármacos , Células Cultivadas , Diazo-Oxo-Norleucina/análogos & derivados , Glutamina/metabolismo , Imuno-Histoquímica , Masculino , Alvo Mecanístico do Complexo 1 de Rapamicina/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Músculo Liso Vascular/metabolismo , Neointima/metabolismo , Fator de Crescimento Derivado de Plaquetas/farmacologia , Ratos , Ratos Sprague-Dawley , Soroalbumina Bovina/farmacologia
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