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1.
Int J Mol Sci ; 22(4)2021 Feb 18.
Artigo em Inglês | MEDLINE | ID: mdl-33670449

RESUMO

With more than 25 million people affected, heart failure (HF) is a global threat. As energy production pathways are known to play a pivotal role in HF, we sought here to identify key metabolic changes in ischemic- and non-ischemic HF by using a multi-OMICS approach. Serum metabolites and mRNAseq and epigenetic DNA methylation profiles were analyzed from blood and left ventricular heart biopsy specimens of the same individuals. In total we collected serum from n = 82 patients with Dilated Cardiomyopathy (DCM) and n = 51 controls in the screening stage. We identified several metabolites involved in glycolysis and citric acid cycle to be elevated up to 5.7-fold in DCM (p = 1.7 × 10-6). Interestingly, cardiac mRNA and epigenetic changes of genes encoding rate-limiting enzymes of these pathways could also be found and validated in our second stage of metabolite assessment in n = 52 DCM, n = 39 ischemic HF and n = 57 controls. In conclusion, we identified a new set of metabolomic biomarkers for HF. We were able to identify underlying biological cascades that potentially represent suitable intervention targets.


Assuntos
Biomarcadores/metabolismo , Cardiomiopatia Dilatada/genética , Epigenômica/métodos , Perfilação da Expressão Gênica/métodos , Insuficiência Cardíaca/genética , Metabolômica/métodos , Adulto , Idoso , Biomarcadores/sangue , Cardiomiopatia Dilatada/diagnóstico , Cardiomiopatia Dilatada/metabolismo , Estudos de Coortes , Epigênese Genética , Feminino , Glicólise/genética , Insuficiência Cardíaca/diagnóstico , Insuficiência Cardíaca/metabolismo , Humanos , Masculino , Pessoa de Meia-Idade , Análise de Componente Principal
2.
Medicine (Baltimore) ; 100(3): e23836, 2021 Jan 22.
Artigo em Inglês | MEDLINE | ID: mdl-33545950

RESUMO

ABSTRACT: Bladder cancer (BC) is one of the most common malignancies worldwide. Several biomarkers related to the prognosis of patients with BC have previously been identified. However, these prognostic models use only one gene and are thus not reliable or accurate enough. The purpose of our study was to develop an innovative gene signature that has greater prognostic value in BC. So, in this study, we performed mRNA expression profiling of glycolysis-related genes in BC (n = 407) cohorts by mining data from The Cancer Genome Atlas (TCGA) database. The glycolysis-related gene sets were confirmed using the Gene Set Enrichment Analysis (GSEA). Using Cox regression analysis, a risk score staging model was built based on the genes that were determined to be significantly associated with BC outcome. Eventually, the system of risk score was structured to predict a patient's survival, and we identified four genes (CHPF, AK3, GALK1, and NUP188) that were associated with the outcomes of BC patients. According to the above-mentioned gene signature, patients were divided into two risk subgroups. The analysis showed that our constructed risk model was independent of clinical features and that the risk score was a highly powerful tool for predicting the overall survival (OS) of BC patients. Taking together, we identified a gene signature associated with glycolysis that could effectively predict the prognosis of BC patients. Our findings offer a new perspective for the clinical research and treatment of BC.


Assuntos
Predisposição Genética para Doença , Glicólise/genética , Neoplasias da Bexiga Urinária/genética , Idoso , Grupo com Ancestrais do Continente Asiático , China , Bases de Dados Factuais , Feminino , Perfilação da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Humanos , Estimativa de Kaplan-Meier , Masculino , Prognóstico , Modelos de Riscos Proporcionais , RNA Mensageiro/metabolismo , Neoplasias da Bexiga Urinária/metabolismo , Neoplasias da Bexiga Urinária/mortalidade
3.
Nan Fang Yi Ke Da Xue Xue Bao ; 41(2): 193-199, 2021 Feb 25.
Artigo em Chinês | MEDLINE | ID: mdl-33624591

RESUMO

OBJECTIVE: To investigate the effect of long non-coding RNA UPK1A-AS1 on glycolysis of hepatocellular carcinoma cells and its underlying molecular mechanisms. OBJECTIVE: A hepatocellular carcinoma (HCC) cell line with lentivirus-mediated stable UPK1A-AS1 overexpression and the cells infected with a negative control lentiviral vector were cultured under normoxic (21% O2) or hypoxic (1% O2) conditions for 24 h. The effect of UPK1A-AS1 overexpression on glycolysis of the HCC cells was examined. The expressions of glycolysis-related genes HIF1A, GLUT1, HK1, HK2 and PGK1 were detected by qRTPCR, and the effect of UPK1A-AS1 overexpression on HRE activity was detected by dual luciferase report assay. The HCC cells were treated with cycloheximide to detect the effect of UPK1A-AS1 overexpression on the stability of HIF-1α protein. Immunoprecipitation assay was used to analyze the changes in ubiquitin modification of HIF-1α protein in response to UPK1A-AS1 overexpression. The effects of UPK1A-AS1 overexpression and RNA interference of HIF-1α expression on glucose consumption, lactate production and expressions of HRE activity and glycolysis-related genes (HK1, HK2 and PGK1) were examined in the HCC cells. OBJECTIVE: Compared with the control group, overexpression of UPK1A-AS1 significantly promoted glucose consumption and lactate production in HCC cells under both normoxic and hypoxic conditions (P < 0.05). Overexpression of UPK1A-AS1 significantly increased the expression of glycolysis-related genes including HIF1A, GLUT1, HK1, HK2 and PGK1. Upregulation of UPK1A-AS1 obviously promoted the transcriptional activity of HRE (P < 0.05). Western blotting showed that UPK1A-AS1 overexpression obviously increased the stability of HIF-1α protein and significantly reduced ubiquitin modification of HIF-1α. Downregulation of HIF-1α obviously reversed the effect of UPK1A-AS1 overexpression in promoting glucose consumption, lactate production and HRE luciferase activity. Silencing HIF-1α also suppressed the upregulation of glycolysis-related gene expressions induced by UPK1A-AS1 overexpression (P < 0.05). OBJECTIVE: The long noncoding RNA UPK1A-AS1 upregulates the expression of glycolysis-related genes by stabilizing the expression of HIF-1α, thereby promoting glycolysis level in HCC cells.


Assuntos
Carcinoma Hepatocelular , Neoplasias Hepáticas , RNA Longo não Codificante , Carcinoma Hepatocelular/genética , Linhagem Celular Tumoral , Glicólise/genética , Humanos , Neoplasias Hepáticas/genética , RNA Longo não Codificante/genética , RNA Longo não Codificante/metabolismo
4.
Mol Cell ; 81(5): 922-939.e9, 2021 03 04.
Artigo em Inglês | MEDLINE | ID: mdl-33434505

RESUMO

R-2-hydroxyglutarate (R-2HG), a metabolite produced by mutant isocitrate dehydrogenases (IDHs), was recently reported to exhibit anti-tumor activity. However, its effect on cancer metabolism remains largely elusive. Here we show that R-2HG effectively attenuates aerobic glycolysis, a hallmark of cancer metabolism, in (R-2HG-sensitive) leukemia cells. Mechanistically, R-2HG abrogates fat-mass- and obesity-associated protein (FTO)/N6-methyladenosine (m6A)/YTH N6-methyladenosine RNA binding protein 2 (YTHDF2)-mediated post-transcriptional upregulation of phosphofructokinase platelet (PFKP) and lactate dehydrogenase B (LDHB) (two critical glycolytic genes) expression and thereby suppresses aerobic glycolysis. Knockdown of FTO, PFKP, or LDHB recapitulates R-2HG-induced glycolytic inhibition in (R-2HG-sensitive) leukemia cells, but not in normal CD34+ hematopoietic stem/progenitor cells, and inhibits leukemogenesis in vivo; conversely, their overexpression reverses R-2HG-induced effects. R-2HG also suppresses glycolysis and downregulates FTO/PFKP/LDHB expression in human primary IDH-wild-type acute myeloid leukemia (AML) cells, demonstrating the clinical relevance. Collectively, our study reveals previously unrecognized effects of R-2HG and RNA modification on aerobic glycolysis in leukemia, highlighting the therapeutic potential of targeting cancer epitranscriptomics and metabolism.


Assuntos
Dioxigenase FTO Dependente de alfa-Cetoglutarato/genética , Antineoplásicos/farmacologia , Glutaratos/farmacologia , Glicólise/genética , Lactato Desidrogenases/genética , Leucemia Mieloide Aguda/tratamento farmacológico , Fosfofrutoquinase-1 Tipo C/genética , Dioxigenase FTO Dependente de alfa-Cetoglutarato/antagonistas & inibidores , Dioxigenase FTO Dependente de alfa-Cetoglutarato/metabolismo , Animais , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Feminino , Fluoruracila/farmacologia , Regulação Neoplásica da Expressão Gênica , Glicólise/efeitos dos fármacos , Células HEK293 , Humanos , Células K562 , Lactato Desidrogenases/antagonistas & inibidores , Lactato Desidrogenases/metabolismo , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/mortalidade , Leucemia Mieloide Aguda/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Fosforilação Oxidativa/efeitos dos fármacos , Fosfofrutoquinase-1 Tipo C/antagonistas & inibidores , Fosfofrutoquinase-1 Tipo C/metabolismo , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Proteínas de Ligação a RNA/genética , Proteínas de Ligação a RNA/metabolismo , Transdução de Sinais , Análise de Sobrevida , Ensaios Antitumorais Modelo de Xenoenxerto
5.
Carbohydr Polym ; 255: 117532, 2021 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-33436261

RESUMO

Aldolase A (ALDOA) facilitated aerobic glycolysis in cancer cells is a potential target in the treatment of hepatocellular carcinoma (HCC). However, only few effective inhibitors of ALDOA have been reported until now. In this research, we found a polysaccharide called HDPS-4II from Holotrichia diomphalia Bates, which can specifically bind to ALDOA with a dissociation constant of 2.86 µM. HDPS-4II with a molecular weight of 19 kDa was a linear triple-helix glucan composed of ɑ-d-1,4-Glcp and ɑ-d-1,6-Glcp in a ratio of 1.0:10.0. HDPS-4II significantly inhibited aldolase enzyme activity, glycolysis, and further inhibited the expression of phosphorylated AMPKα in HCC cells. Through analyzing ALDOA-overexpressing and -knockdown cells, it was confirmed that ALDOA mediated the viability and glycolysis inhibition of HDPS-4II. Moreover, HDPS-4II administration markedly inhibited tumor growth in mice xenografted with HCCs. These findings suggest that HDPS-4II, as an ALDOA antagonist, is a promising remedy in the treatment and prevention of HCC.


Assuntos
Antineoplásicos/farmacologia , Besouros/química , Frutose-Bifosfato Aldolase/genética , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Glucanos/farmacologia , Neoplasias Hepáticas/tratamento farmacológico , Animais , Antineoplásicos/química , Antineoplásicos/isolamento & purificação , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Frutose-Bifosfato Aldolase/antagonistas & inibidores , Frutose-Bifosfato Aldolase/metabolismo , Glucanos/química , Glucanos/isolamento & purificação , Glicólise/efeitos dos fármacos , Glicólise/genética , Células Hep G2 , Hexoquinase/genética , Hexoquinase/metabolismo , Humanos , L-Lactato Desidrogenase/genética , L-Lactato Desidrogenase/metabolismo , Larva/química , Neoplasias Hepáticas/enzimologia , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/patologia , Camundongos , Piruvato Quinase/genética , Piruvato Quinase/metabolismo , Carga Tumoral/efeitos dos fármacos , Ensaios Antitumorais Modelo de Xenoenxerto
6.
Int J Mol Sci ; 22(2)2021 Jan 07.
Artigo em Inglês | MEDLINE | ID: mdl-33430235

RESUMO

The role of ketone bodies in the cerebral energy homeostasis of neurological diseases has begun to attract recent attention particularly in acute neurological diseases. In ketogenic therapies, ketosis is achieved by either a ketogenic diet or by the administration of exogenous ketone bodies. The oral ingestion of the ketone ester (KE), (R)-3-hydroxybutyl (R)-3-hydroxybutyrate, is a new method to generate rapid and significant ketosis (i.e., above 6 mmol/L) in humans. KE is hydrolyzed into ß-hydroxybutyrate (ßHB) and its precursor 1,3-butanediol. Here, we investigate the effect of oral KE administration (3 mg KE/g of body weight) on brain metabolism of non-fasted mice using liquid chromatography in tandem with mass spectrometry. Ketosis (Cmax = 6.83 ± 0.19 mmol/L) was obtained at Tmax = 30 min after oral KE-gavage. We found that ßHB uptake into the brain strongly correlated with the plasma ßHB concentration and was preferentially distributed in the neocortex. We showed for the first time that oral KE led to an increase of acetyl-CoA and citric cycle intermediates in the brain of non-fasted mice. Furthermore, we found that the increased level of acetyl-CoA inhibited glycolysis by a feedback mechanism and thus competed with glucose under physiological conditions. The brain pharmacodynamics of this oral KE strongly suggest that this agent should be considered for acute neurological diseases.


Assuntos
Acetilcoenzima A/metabolismo , Encéfalo/metabolismo , Metabolismo dos Carboidratos/genética , Cetonas/metabolismo , Animais , Dieta Cetogênica/efeitos adversos , Ingestão de Alimentos , Ésteres/metabolismo , Glucose/metabolismo , Glicólise/genética , Humanos , Corpos Cetônicos/metabolismo , Cetose/metabolismo , Cetose/patologia , Camundongos
7.
Int J Mol Sci ; 22(2)2021 Jan 07.
Artigo em Inglês | MEDLINE | ID: mdl-33430468

RESUMO

Alterations in miRNAs are associated with many metabolic disorders, such as type 2 diabetes (T2DM). The miR-23b/27b/24-1 cluster contains miR-23b, miR-27b, and miR-24-1, which are located within 881 bp on chromosome 9. Studies examining the roles of miR-23b, miR-27b, and miR-24-1 have demonstrated their multifaceted functions in variable metabolic disorders. However, their joint roles in metabolism in vivo remain elusive. To investigate this subject, we constructed miR-23b/27b/24-1 cluster knockout (KO) mice. Compared with wild-type (WT) mice, the KO mice exhibited impaired glucose tolerance, which was accompanied by a reduction in the respiratory exchange rate (RER). These alterations were more noticeable after a high-fat diet (HFD) induction. Hepatic metabolomic results showed decreased expression of reduced nicotinamide adenine dinucleotide (NADH), nicotinamide adenine dinucleotide (NAD), phosphoenolpyruvic acid (PEP), and phosphoric acid, which are involved in the glycolysis pathway. The transcriptomic results indicated that genes involved in glycolysis showed a downregulation trend. qPCR and Western blot revealed that pyruvate kinase (PKLR), the key rate-limiting enzyme in glycolysis, was significantly reduced after the deletion of the miR-23b/27b/24-1 cluster. Together, these observations suggest that the miR-23b/27b/24-1 cluster is involved in the regulation of glucose homeostasis via the glycolysis pathway.


Assuntos
Diabetes Mellitus Tipo 2/genética , Intolerância à Glucose/genética , Glucose/genética , MicroRNAs/genética , Animais , Diabetes Mellitus Tipo 2/patologia , Glucose/metabolismo , Glicólise/genética , Humanos , Camundongos , Camundongos Knockout , Família Multigênica/genética , NAD/metabolismo , Taxa Respiratória/genética , Transdução de Sinais/genética
8.
Nucleic Acids Res ; 49(3): 1769-1783, 2021 02 22.
Artigo em Inglês | MEDLINE | ID: mdl-33423048

RESUMO

The construction of microbial cell factories for sustainable production of chemicals and pharmaceuticals requires extensive genome engineering. Using Saccharomyces cerevisiae, this study proposes synthetic neochromosomes as orthogonal expression platforms for rewiring native cellular processes and implementing new functionalities. Capitalizing the powerful homologous recombination capability of S. cerevisiae, modular neochromosomes of 50 and 100 kb were fully assembled de novo from up to 44 transcriptional-unit-sized fragments in a single transformation. These assemblies were remarkably efficient and faithful to their in silico design. Neochromosomes made of non-coding DNA were stably replicated and segregated irrespective of their size without affecting the physiology of their host. These non-coding neochromosomes were successfully used as landing pad and as exclusive expression platform for the essential glycolytic pathway. This work pushes the limit of DNA assembly in S. cerevisiae and paves the way for de novo designer chromosomes as modular genome engineering platforms in S. cerevisiae.


Assuntos
Engenharia Celular , Cromossomos , Saccharomyces cerevisiae/genética , Glicólise/genética
9.
Gene ; 766: 145157, 2021 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-32949697

RESUMO

Glycolytic potential (GP) calculated based on glucose, glycogen, glucose-6-phosphate, and lactate contents is a critical factor for multiple meat quality characteristics. However, the genetic basis of glycolytic metabolism is still unclear. In this study, we constructed six RNA-Seq libraries using longissimus dorsi (LD) muscles from pigs divergent for GP phenotypic values and generated the whole genome-wide gene expression profiles. Furthermore, we identified 25,880 known and 220 novel genes from these skeletal muscle libraries, and 222 differentially expressed genes (DEGs) between the higher and lower GP groups. Notably, we found that the Lactate dehydrogenase B (LDHB) and Fructose-2, 6-biphosphatase 3 (PFKFB3) expression levels were higher in the higher GP group than the lower GP group, and positively correlated with GP and lactic acid (LA), and reversely correlated with pH value at 45 min postmortem (pH45min). Besides, LDHB and PFKFB3 expression were positively correlated with drip loss measured at 48 h postmortem (DL48h) and drip loss measured at 24 h postmortem (DL24h). Collectively, we identified a serial of DEGs as the potential key candidate genes affecting GP and found that LDHB and PFKFB3 are closely related to GP and GP-related traits. Our results lay a solid basis for in-depth studies of the regulatory mechanisms on GP and GP-related traits in pigs.


Assuntos
Glicólise/genética , Músculo Esquelético/metabolismo , Suínos/genética , Transcriptoma/genética , Animais , Perfilação da Expressão Gênica/métodos , Glucose/genética , Glicogênio/genética , Isoenzimas/genética , L-Lactato Desidrogenase/genética , Ácido Láctico/metabolismo , Carne , Fenótipo , Fosfofrutoquinase-2/genética , Suínos/metabolismo
10.
Methods Mol Biol ; 2230: 437-447, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33197031

RESUMO

Measuring cellular metabolism accurately is necessary to understand bioenergetic pathways in cells. The major ATP generating pathways in cells are oxidative phosphorylation and glycolysis. We have recently analyzed and published bioenergetic pathways active in osteoblasts undergoing differentiation in response to various substrates. Based on those studies, here we provide step-by-step procedures to isolate, culture, plate and run a seahorse assay for measuring cellular metabolism. Furthermore, we provide an example of oxygen consumption and extracellular acidification rate traces obtained from MC3T3E1-C4 cells using the XFe96 seahorse analyzer. One of the limitations of studying bioenergetics in bone cells is the current lack of techniques to analyze bioenergetics in vivo in live animals. There are currently techniques that have been developed using third harmonic generation to study osteocytes using three-photon microscopy along with metabolic changes using endogenous two-photon excited fluorescence. However, these sophisticated techniques are not widely available. The relative ease with which one can obtain data pertaining to metabolic parameters using the XF technology makes it a very attractive technique to utilize on a monolayer of adherent cells.


Assuntos
Osso e Ossos/metabolismo , Técnicas de Cultura de Células/métodos , Metabolismo Energético/genética , Osteoblastos/metabolismo , Células 3T3 , Animais , Glicólise/genética , Camundongos , Consumo de Oxigênio/genética , Fosforilação/genética
11.
Biochim Biophys Acta Mol Cell Res ; 1868(1): 118854, 2021 01.
Artigo em Inglês | MEDLINE | ID: mdl-32926942

RESUMO

Mitochondria are highly dynamic organelles. Alterations in mitochondrial dynamics are causal or are linked to numerous neurodegenerative, neuromuscular, and metabolic diseases. It is generally thought that cells with altered mitochondrial structure are prone to mitochondrial dysfunction, increased reactive oxygen species generation and widespread oxidative damage. The objective of the current study was to investigate the relationship between mitochondrial dynamics and the master cellular antioxidant, glutathione (GSH). We reveal that mouse embryonic fibroblasts (MEFs) lacking the mitochondrial fusion machinery display elevated levels of GSH, which limits oxidative damage. Moreover, targeted metabolomics and 13C isotopic labeling experiments demonstrate that cells lacking the inner membrane fusion GTPase OPA1 undergo widespread metabolic remodeling altering the balance of citric acid cycle intermediates and ultimately favoring GSH synthesis. Interestingly, the GSH precursor and antioxidant n-acetylcysteine did not increase GSH levels in OPA1 KO cells, suggesting that cysteine is not limiting for GSH production in this context. Post-mitotic neurons were unable to increase GSH production in the absence of OPA1. Finally, the ability to use glycolysis for ATP production was a requirement for GSH accumulation following OPA1 deletion. Thus, our results demonstrate a novel role for mitochondrial fusion in the regulation of GSH synthesis, and suggest that cysteine availability is not limiting for GSH synthesis in conditions of mitochondrial fragmentation. These findings provide a possible explanation for the heightened sensitivity of certain cell types to alterations in mitochondrial dynamics.


Assuntos
Antioxidantes/metabolismo , Glutationa/genética , Mitocôndrias/genética , Dinâmica Mitocondrial/genética , Trifosfato de Adenosina/metabolismo , Animais , Apoptose/genética , GTP Fosfo-Hidrolases/genética , Glutationa/biossíntese , Glicólise/genética , Humanos , Fusão de Membrana/genética , Camundongos , Mitocôndrias/metabolismo , Estresse Oxidativo/genética , Espécies Reativas de Oxigênio/metabolismo
12.
Artigo em Inglês | MEDLINE | ID: mdl-33065288

RESUMO

We previously demonstrated that astaxanthin (ASTX), a xanthophyll carotenoid, repressed ethanol-induced inflammation and oxidative stress in macrophages. We explored the role of sirtuin 1 (SIRT1) and histone deacetylase 4 (HDAC4) in the inhibitory effect of ASTX on inflammation and oxidative stress in macrophages exposed to ethanol. Ethanol decreased mRNA and protein of SIRT1 while increasing those of HDAC4, which was attenuated by ASTX in RAW 264.7 macrophages and mouse bone marrow-derived macrophages (BMDMs). Inhibition of SIRT1 expression or activity augmented ethanol-induced Hdac4 expression, but SIRT1 activation elicited the opposite effect. Consistently, Hdac4 knockdown increased Sirt1 expression with decreases in ethanol-induced inflammatory gene expression, but its overexpression resulted in the opposite effects. Furthermore, BMDMs from mice with macrophage specific-deletion of Hdac4 (Hdac4MKO) showed significant decreases in ethanol-induced inflammatory genes and ROS accumulation but an increase in Sirt1 expression. Macrophage specific deletion of Hdac4 or ASTX abolished the changes in genes for mitochondrial biogenesis and glycolysis by ethanol. Ethanol increased mitochondrial respiration, ATP production, and proton leak, but decreased maximal respiration and spare respiratory capacity, all of which were abolished by ASTX in RAW 264.7 macrophages. The ethanol-induced alterations in mitochondrial respiration were abrogated in Hdac4MKO BMDMs. In conclusion, the anti-inflammatory and antioxidant properties of ASTX in ethanol-treated macrophages may be mediated, at least partly, by its opposite effect on SIRT1 and HDAC4 to empower SIRT1 to counteract ethanol-induced activation of HDAC4.


Assuntos
Antioxidantes/farmacologia , Etanol/antagonistas & inibidores , Histona Desacetilases/genética , Macrófagos/efeitos dos fármacos , Sirtuína 1/genética , Trifosfato de Adenosina/biossíntese , Animais , Etanol/farmacologia , Regulação da Expressão Gênica , Glicólise/efeitos dos fármacos , Glicólise/genética , Histona Desacetilases/metabolismo , Inflamação , Macrófagos/citologia , Macrófagos/metabolismo , Camundongos , Camundongos Knockout , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/genética , Mitocôndrias/metabolismo , Biogênese de Organelas , Fosforilação Oxidativa/efeitos dos fármacos , Estresse Oxidativo , Cultura Primária de Células , Células RAW 264.7 , Espécies Reativas de Oxigênio/agonistas , Espécies Reativas de Oxigênio/antagonistas & inibidores , Espécies Reativas de Oxigênio/metabolismo , Sirtuína 1/metabolismo , Xantofilas/farmacologia
13.
Life Sci ; 268: 118925, 2021 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-33358903

RESUMO

AIMS: Osteosarcoma (OS) is an extremely malignant bone cancer with high incidence and rapid progression. This study aims to investigate the role and underlying mechanisms of MALAT1 and miR-485-3p in OS. MATERIALS AND METHODS: qRT-PCR and Western blotting were utilized to measure the levels of miR-485-3p, MALAT1, c-MET, AKT3, p-mTOR, mTOR, glycolysis-related proteins or migration-related proteins. Colony formation and transwell assay were used to test the roles of miR-485-3p, MALAT1, c-MET and AKT3 in cancer cell proliferation, migration and invasion. Dual luciferase assay was used to validate the interactions of miR-485-3p/c-MET, miR-485-3p/AKT3, and MALAT1/miR-485-3p. Glucose uptake assay and measurement of lactate production were employed to determine the glycolysis process. Mouse tumour xenograft model was used to determine the effect of shMALAT1 and miR-485-3p mimics on tumour growth and metastasis in vivo. KEY FINDINGS: miR-485-3p was decreased while c-MET, AKT3, and MALAT1 were increased in human OS tissues and cells. miR-485-3p bound directly to c-MET and AKT3 mRNAs and repressed OS cell glycolysis, proliferation, migration, and invasion through decreasing glycolysis-related proteins and migration-related proteins via inhibiting c-MET and AKT3/mTOR pathway. In addition, MALAT1 interacted with miR-485-3p and disinhibited c-MET and AKT3/mTOR signalling. Knockdown MALAT1 or overexpression of miR-485-3p restrained OS tumour growth and lung metastasis in vivo. SIGNIFICANCE: miR-485-3p suppresses OS glycolysis, proliferation, and metastasis via inhibiting c-MET and AKT3/mTOR signalling and MALAT1 acts as a sponge of miR-485-3p. MALAT1 and miR-485-3p may be the key regulators in OS progression, and potential molecular targets for future OS therapy.


Assuntos
Neoplasias Ósseas/patologia , MicroRNAs/genética , Osteossarcoma/patologia , Proteínas Proto-Oncogênicas c-met/genética , RNA Longo não Codificante/genética , Animais , Neoplasias Ósseas/genética , Neoplasias Ósseas/metabolismo , Linhagem Celular Tumoral , Movimento Celular/genética , Regulação Neoplásica da Expressão Gênica , Técnicas de Silenciamento de Genes , Glicólise/genética , Humanos , Masculino , Camundongos Endogâmicos BALB C , Osteossarcoma/genética , Osteossarcoma/metabolismo , Proteínas Proto-Oncogênicas c-akt/genética , Proteínas Proto-Oncogênicas c-akt/metabolismo , Proteínas Proto-Oncogênicas c-met/metabolismo , Serina-Treonina Quinases TOR/genética , Serina-Treonina Quinases TOR/metabolismo , Ensaios Antitumorais Modelo de Xenoenxerto
14.
Biochim Biophys Acta Rev Cancer ; 1875(1): 188485, 2021 01.
Artigo em Inglês | MEDLINE | ID: mdl-33309965

RESUMO

Adenine nucleotide translocases (ANTs) are a class of transporters located in the inner mitochondrial membrane that not only couple processes of cellular productivity and energy expenditure, but are also involved in the composition of the mitochondrial membrane permeability transition pore (mPTP). The function of ANTs has been found to be most closely related to their own conformational changes. Notably, as multifunctional proteins, ANTs play a key role in oncogenesis, which provides building blocks for tumor anabolism, control oxidative phosphorylation and glycolysis homeostasis, and govern cell death. Thus, ANTs constitute promising targets for the development of novel anticancer agents. Here, we review the recent findings regarding ANTs and their important mechanisms in cancer, with a focus on the therapeutic potential of targeting ANTs for cancer therapy.


Assuntos
Carcinogênese/genética , Transformação Celular Neoplásica/genética , Translocases Mitocondriais de ADP e ATP/genética , Neoplasias/genética , Carcinogênese/metabolismo , Permeabilidade da Membrana Celular/genética , Transformação Celular Neoplásica/metabolismo , Metabolismo Energético/genética , Glicólise/genética , Humanos , Mitocôndrias/genética , Mitocôndrias/metabolismo , Neoplasias/metabolismo , Neoplasias/patologia , Fosforilação Oxidativa
15.
Science ; 371(6531)2021 02 19.
Artigo em Inglês | MEDLINE | ID: mdl-33335020

RESUMO

Single-cell RNA sequencing (scRNA-seq) has become an essential tool for characterizing gene expression in eukaryotes, but current methods are incompatible with bacteria. Here, we introduce microSPLiT (microbial split-pool ligation transcriptomics), a high-throughput scRNA-seq method for Gram-negative and Gram-positive bacteria that can resolve heterogeneous transcriptional states. We applied microSPLiT to >25,000 Bacillus subtilis cells sampled at different growth stages, creating an atlas of changes in metabolism and lifestyle. We retrieved detailed gene expression profiles associated with known, but rare, states such as competence and prophage induction and also identified unexpected gene expression states, including the heterogeneous activation of a niche metabolic pathway in a subpopulation of cells. MicroSPLiT paves the way to high-throughput analysis of gene expression in bacterial communities that are otherwise not amenable to single-cell analysis, such as natural microbiota.


Assuntos
Bacillus subtilis/genética , Regulação Bacteriana da Expressão Gênica , Redes e Vias Metabólicas/genética , RNA-Seq/métodos , Análise de Célula Única/métodos , Antibacterianos/biossíntese , Fagos Bacilares/fisiologia , Bacillus subtilis/crescimento & desenvolvimento , Bacillus subtilis/metabolismo , Carbono/metabolismo , Meios de Cultura , Escherichia coli/genética , Fermentação/genética , Gluconeogênese/genética , Glicólise/genética , Resposta ao Choque Térmico/genética , Inositol/metabolismo , Transporte de Íons , Metais/metabolismo , Movimento , Óperon , RNA Bacteriano/genética , Estresse Fisiológico , Transcrição Genética , Transcriptoma , Ativação Viral
16.
Cell Prolif ; 54(2): e12913, 2021 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-33332677

RESUMO

MicroRNAs (miRNAs) are small and highly conserved non-coding RNAs that silence target mRNAs, and compelling evidence suggests that they play an essential role in the pathogenesis of human diseases, especially cancer. miR-125b, which is the mammalian orthologue of the first discovered miRNA lin-4 in Caenorhabditis elegans, is one of the most important miRNAs that regulate various physiological and pathological processes. The role of miR-125b in many types of cancer has been well established, and so here we review the current knowledge of how miR-125b is deregulated in different types of cancer; its oncogenic and/or tumour-suppressive roles in tumourigenesis and cancer progression; and its regulation with regard to treatment response, all of which are underlined in multiple studies. The emerging information that elucidates the essential functions of miR-125b might help support its potentiality as a diagnostic and prognostic biomarker as well as an effective therapeutic tool against cancer.


Assuntos
MicroRNAs/metabolismo , Neoplasias/patologia , Apoptose/genética , Resistencia a Medicamentos Antineoplásicos/genética , Glicólise/genética , Histonas/metabolismo , Humanos , Metástase Neoplásica , Neoplasias/genética , RNA Longo não Codificante/metabolismo , Transdução de Sinais/genética
17.
PLoS One ; 15(12): e0243842, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-33320884

RESUMO

Activation of the aryl hydrocarbon receptor (AHR) by the environmental toxin dioxin (2,3,7,8-tetrachlorodibenzo-p-dioxin, TCDD) causes diverse toxicities, including thymus atrophy and hepatosteatosis. The mechanisms by which AHR activation by TCDD leads to these toxicities are not fully understood. Here we studied the effects of TCDD on a major energy pathway, glycolysis, using the chick embryo close to hatching, a well-established model for studying dioxin toxicity. We showed that 24 hr of TCDD treatment causes changes in glycolysis in both thymus and liver. In thymus glands, TCDD decreased mRNAs for glycolytic genes and glucose transporters, glycolytic indices and levels of IL7 mRNA, phosphorylated AKT (pAKT) and HIF1A, stimulators of glycolysis and promoters of survival and proliferation of thymic lymphocytes. In contrast, in liver, TCDD increased mRNA levels for glycolytic genes and glucose transporters, glycolytic endpoints and pAKT levels. Similarly, increases by TCDD in mRNA levels for glycolytic genes and glucose transporters in human primary hepatocytes showed that effects in chick embryo liver pertain also to human cells. Treatment with the glycolytic inhibitor 2-deoxy-d-glucose exacerbated the effects on thymus atrophy by TCDD, supporting a role for decreased glycolysis in thymus atrophy by TCDD, but did not prevent hepatosteatosis. NAD+ precursors abolished TCDD effects on glycolytic endpoints in both thymus and liver. In summary, we report here that dioxin disrupts glycolysis mediated energy metabolism in both thymus and liver, and that it does so in opposite ways, decreasing it in the thymus and increasing it in the liver. Further, the findings support NAD+ boosting as a strategy against metabolic effects of environmental pollutants such as dioxins.


Assuntos
Dioxinas/toxicidade , Glicólise/efeitos dos fármacos , Especificidade de Órgãos/efeitos dos fármacos , Receptores de Hidrocarboneto Arílico/metabolismo , Adulto , Animais , Células Cultivadas , Embrião de Galinha , Feminino , Regulação da Expressão Gênica/efeitos dos fármacos , Glucose/metabolismo , Glicólise/genética , Humanos , Fígado/efeitos dos fármacos , Fígado/metabolismo , Proteínas de Membrana Transportadoras/genética , Proteínas de Membrana Transportadoras/metabolismo , Niacinamida/análogos & derivados , Niacinamida/metabolismo , Dibenzodioxinas Policloradas/toxicidade , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Timo/efeitos dos fármacos , Timo/embriologia , Timo/metabolismo
18.
Inflamm Res ; 69(12): 1215-1234, 2020 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-33044562

RESUMO

OBJECTIVE AND DESIGN: Macrophages exhibit strong phenotypic plasticity and can mediate renal inflammation by polarizing into an M1 phenotype. They play a pivotal role in diabetic nephropathy (DN). Here, we have investigated the regulatory role of transforming growth factor ß-activated kinase 1-binding protein 1 (TAB1) in glycolysis and activation of macrophages during DN. METHODS: TAB1 was inhibited using siRNA in high glucose (HG)-stimulated bone marrow-derived macrophages (BMMs) and lentiviral vector-mediated TAB1 knockdown was used in streptozotocin (STZ)-induced diabetic mice. Western blotting, flow cytometry, qRT-PCR, ELISA, PAS staining and immunohistochemical staining were used for assessment of TAB1/nuclear factor-κB (NF-κB)/hypoxia-inducible factor-1α (HIF-1α), iNOS, glycolysis, inflammation and the clinical and pathological manifestations of diabetic nephropathy. RESULTS: We found that TAB1/NF-κB/HIF-1α, iNOS and glycolysis were up-regulated in BMMs under HG conditions, leading to release of further inflammatory factors, Downregulation of TAB1 could inhibit glycolysis/polarization of macrophages and inflammation in vivo and in vitro. Furthermore, albuminuria, the tubulointerstitial damage index and glomerular mesangial expansion index of STZ-induced diabetic nephropathy mice were decreased by TAB1 knockdown. CONCLUSIONS: Our results suggest that the TAB1/NF-κB/HIF-1α signaling pathway regulates glycolysis and activation of macrophages in DN.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Neuropatias Diabéticas/genética , Neuropatias Diabéticas/metabolismo , Glicólise/genética , Ativação de Macrófagos/genética , Proteínas Adaptadoras de Transdução de Sinal/antagonistas & inibidores , Albuminúria/tratamento farmacológico , Albuminúria/patologia , Animais , Diabetes Mellitus Experimental/complicações , Diabetes Mellitus Experimental/metabolismo , Técnicas de Silenciamento de Genes , Subunidade alfa do Fator 1 Induzível por Hipóxia/efeitos dos fármacos , Células Mesangiais/patologia , Camundongos , Camundongos Endogâmicos C57BL , NF-kappa B/efeitos dos fármacos , Nefrite Intersticial/patologia , Óxido Nítrico Sintase Tipo II/efeitos dos fármacos , RNA Interferente Pequeno/farmacologia
19.
Nat Commun ; 11(1): 5163, 2020 10 14.
Artigo em Inglês | MEDLINE | ID: mdl-33057020

RESUMO

Parkinson's disease-associated kinase LRRK2 has been linked to IFN type II (IFN-γ) response in infections and to dopaminergic neuronal loss. However, whether and how LRRK2 synergizes with IFN-γ remains unclear. In this study, we employed dopaminergic neurons and microglia differentiated from patient-derived induced pluripotent stem cells carrying LRRK2 G2019S, the most common Parkinson's disease-associated mutation. We show that IFN-γ enhances the LRRK2 G2019S-dependent negative regulation of AKT phosphorylation and NFAT activation, thereby increasing neuronal vulnerability to immune challenge. Mechanistically, LRRK2 G2019S suppresses NFAT translocation via calcium signaling and possibly through microtubule reorganization. In microglia, LRRK2 modulates cytokine production and the glycolytic switch in response to IFN-γ in an NFAT-independent manner. Activated LRRK2 G2019S microglia cause neurite shortening, indicating that LRRK2-driven immunological changes can be neurotoxic. We propose that synergistic LRRK2/IFN-γ activation serves as a potential link between inflammation and neurodegeneration in Parkinson's disease.


Assuntos
Neurônios Dopaminérgicos/imunologia , Interferon gama/metabolismo , Serina-Treonina Proteína Quinase-2 com Repetições Ricas em Leucina/metabolismo , Microglia/imunologia , Doença de Parkinson/imunologia , Sinalização do Cálcio/genética , Diferenciação Celular , Citocinas/metabolismo , Neurônios Dopaminérgicos/metabolismo , Técnicas de Inativação de Genes , Glicólise/genética , Células HEK293 , Humanos , Células-Tronco Pluripotentes Induzidas/fisiologia , Interferon gama/imunologia , Microscopia Intravital , Serina-Treonina Proteína Quinase-2 com Repetições Ricas em Leucina/genética , Microglia/metabolismo , Microtúbulos/metabolismo , Mutação , Fatores de Transcrição NFATC/metabolismo , Doença de Parkinson/genética , Doença de Parkinson/patologia , Cultura Primária de Células , Transdução de Sinais/genética , Transdução de Sinais/imunologia , Células THP-1
20.
Life Sci ; 259: 118215, 2020 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-32768579

RESUMO

AIMS: Infantile hemangioma (IH) is one of the most common tumors in infancy, which etiology and pathogenesis has not been fully elucidated, hypoxia and abnormal glucose metabolism is regarded as critical pathogenic factors. This study investigated the expression and function of glycolysis-associated molecules (GLUT1, HK2, PFKFB3, PKM2, and LDHA) under normoxic and hypoxic conditions to further understand the pathogenesis of IH. MAIN METHODS: Hemangioma-derived endothelial cells (HemECs) were isolated from proliferating phase infantile hemangiomas and identified by immunofluorescence. HemECs and human umbilical vein endothelial cells (HUVECs) were cultured under normoxic and hypoxic conditions. RNA and protein expression of glycolysis-associated molecules were analyzed by quantitative real-time RT-PCR, western blotting, and immunohistochemistry. Glucose consumption, ATP production and lactate production were measured. Glycolysis-associated molecules were inhibited by WZB117, 3BP, 3PO, SKN, and GSK 2837808A and the resulting effects on HemECs proliferation, migration, and tube formation were quantified. KEY FINDINGS: Glycolysis-associated molecules were highly expressed at both mRNA and protein levels in HemECs compared with HUVECs (P < 0.05). Glucose consumption and ATP production were higher in HemECs than in HUVECs, while lactate production in HemECs was lower than in HUVECs (P < 0.05). Inhibition of some glycolysis-associated molecules reduced the proliferation, migration, and tube formation capacity of HemECs (P < 0.05). SIGNIFICANCE: Our study revealed that glycolysis-associated molecules were highly expressed in IH. Glucose metabolismin HemECs differed from normal endothelial cells. Altering the expression of glycolysis-associated molecules may influence the phenotype of HemECs and provide new therapeutic approaches to the successful treatment of IH.


Assuntos
Glicólise/fisiologia , Hemangioma/metabolismo , Apoptose/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , China , Feminino , Transportador de Glucose Tipo 1/genética , Transportador de Glucose Tipo 1/metabolismo , Glicólise/genética , Hemangioma/fisiopatologia , Hexoquinase/genética , Hexoquinase/metabolismo , Células Endoteliais da Veia Umbilical Humana , Humanos , Hipóxia , Lactente , Recém-Nascido , L-Lactato Desidrogenase/genética , L-Lactato Desidrogenase/metabolismo , Masculino , Fosfofrutoquinase-2/genética , Fosfofrutoquinase-2/metabolismo , Cultura Primária de Células , Proteínas Proto-Oncogênicas c-akt/metabolismo , Transdução de Sinais/efeitos dos fármacos , Transcriptoma/genética
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