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1.
Free Radic Biol Med ; 145: 349-356, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31605749

RESUMO

Appropriate diet is essential for the regulation of age-related macular degeneration (AMD). In particular the type of dietary polyunsaturated fatty acids (PUFA) and poor antioxidant status including carotenoid levels concomitantly contribute to AMD risk. Build-up of oxidative stress in AMD induces PUFA oxidation, and a mix of lipid oxidation products (LOPs) are generated. However, LOPs are not comprehensively evaluated in AMD. LOPs are considered biomarkers of oxidative stress but also contributes to inflammatory response. In this cross-sectional case-control study, plasma omega-6/omega-3 PUFA ratios and antioxidant status (glutathione, superoxide dismutase and catalase), and plasma and urinary LOPs (41 types) were determined to evaluate its odds-ratio in the risk of developing exudative AMD (n = 99) compared to age-gender-matched healthy controls (n = 198) in adults with Chinese diet. The odds ratio of developing exudative AMD increased with LOPs from omega-6 PUFA and decreased from those of omega-3 PUFA. These observations were associated with a high plasma omega-6/omega-3 PUFA ratio and low carotenoid levels. In short, poor PUFA and antioxidant status increased the production of omega-6 PUFA LOPs such as dihomo-isoprostane and dihomo-isofuran, and lowered omega-3 PUFA LOPs such as neuroprostanes due to the high omega-6/omega-3 PUFA ratios; they were also correlated to the risk of AMD development. These findings indicate the generation of specific LOPs is associated with the development of exudative AMD.


Assuntos
Ácidos Graxos Ômega-3/administração & dosagem , Ácidos Graxos Ômega-6/administração & dosagem , Degeneração Macular/metabolismo , Estresse Oxidativo/efeitos dos fármacos , 3-Hidroxiacil-CoA Desidrogenases/genética , 3-Hidroxiacil-CoA Desidrogenases/metabolismo , Acetil-CoA C-Aciltransferase/genética , Acetil-CoA C-Aciltransferase/metabolismo , Idoso , Aldeídos/administração & dosagem , Antioxidantes/administração & dosagem , Biomarcadores/sangue , Isomerases de Ligação Dupla Carbono-Carbono/genética , Isomerases de Ligação Dupla Carbono-Carbono/metabolismo , Carotenoides/metabolismo , Dieta/efeitos adversos , Enoil-CoA Hidratase/genética , Enoil-CoA Hidratase/metabolismo , Ácidos Graxos Ômega-3/metabolismo , Ácidos Graxos Ômega-6/metabolismo , Feminino , Humanos , Isoprostanos/administração & dosagem , Peroxidação de Lipídeos/efeitos dos fármacos , Peroxidação de Lipídeos/genética , Degeneração Macular/etiologia , Degeneração Macular/genética , Degeneração Macular/patologia , Masculino , Pessoa de Meia-Idade , Neuroprostanos/administração & dosagem , Oxirredução/efeitos dos fármacos , Estresse Oxidativo/genética , Racemases e Epimerases/genética , Racemases e Epimerases/metabolismo , Fatores de Risco
2.
Int J Mol Sci ; 20(16)2019 Aug 20.
Artigo em Inglês | MEDLINE | ID: mdl-31434294

RESUMO

Our previous studies showed that microRNA-15a (miR-15a) was closely related to intramuscular fat (IMF) deposition in chickens; however, its regulatory mechanism remains unclear. Here, we evaluated the expression characteristics of miR-15a and its relationship with the expression of acetyl-CoA acyltransferase 1 (ACAA1), acyl-CoA oxidase 1 (ACOX1) and sterol carrier protein 2 (SCP2) by qPCR analysis in Gushi chicken breast muscle at 6, 14, 22, and 30 weeks old, where we performed transfection tests of miR-15a mimics in intramuscular preadipocytes and verified the target gene of miR-15a in chicken fibroblasts (DF1). The miR-15a expression level at 30 weeks increased 13.5, 4.5, and 2.7-fold compared with the expression levels at 6, 14, and 22 weeks, respectively. After 6 days of induction, miR-15a over-expression significantly promoted intramuscular adipogenic differentiation and increased cholesterol and triglyceride accumulation in adipocytes. Meanwhile, 48 h after transfection with miR-15a mimics, the expression levels of ACAA1, ACOX1 and SCP2 genes decreased by 56.52%, 31.18% and 37.14% at the mRNA level in intramuscular preadipocytes. In addition, the co-transfection of miR-15a mimics and ACAA1, ACOX1 and SCP2 3'UTR (untranslated region) dual-luciferase vector significantly inhibited dual-luciferase activity in DF1 cells. Taken together, our data demonstrate that miR-15a can reduce fatty acid oxidation by targeting ACAA1, ACOX1, and SCP2, which subsequently indirectly promotes the differentiation of chicken intramuscular preadipocytes.


Assuntos
Acetil-CoA C-Aciltransferase/metabolismo , Adipócitos/classificação , Adipócitos/metabolismo , Proteínas de Transporte/metabolismo , Diferenciação Celular/fisiologia , MicroRNAs/metabolismo , Acetil-CoA C-Aciltransferase/genética , Animais , Proteínas de Ligação ao Cálcio/genética , Proteínas de Ligação ao Cálcio/metabolismo , Proteínas de Transporte/genética , Diferenciação Celular/genética , Galinhas , MicroRNAs/genética
3.
Hum Mutat ; 40(10): 1641-1663, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31268215

RESUMO

Mitochondrial acetoacetyl-CoA thiolase (T2, encoded by the ACAT1 gene) deficiency is an inherited disorder of ketone body and isoleucine metabolism. It typically manifests with episodic ketoacidosis. The presence of isoleucine-derived metabolites is the key marker for biochemical diagnosis. To date, 105 ACAT1 variants have been reported in 149 T2-deficient patients. The 56 disease-associated missense ACAT1 variants have been mapped onto the crystal structure of T2. Almost all these missense variants concern residues that are completely or partially buried in the T2 structure. Such variants are expected to cause T2 deficiency by having lower in vivo T2 activity because of lower folding efficiency and/or stability. Expression and activity data of 30 disease-associated missense ACAT1 variants have been measured by expressing them in human SV40-transformed fibroblasts. Only two variants (p.Cys126Ser and p.Tyr219His) appear to have equal stability as wild-type. For these variants, which are inactive, the side chains point into the active site. In patients with T2 deficiency, the genotype does not correlate with the clinical phenotype but exerts a considerable effect on the biochemical phenotype. This could be related to variable remaining residual T2 activity in vivo and has important clinical implications concerning disease management and newborn screening.


Assuntos
Acetil-CoA C-Acetiltransferase/genética , Acetil-CoA C-Aciltransferase/deficiência , Erros Inatos do Metabolismo dos Aminoácidos/genética , Predisposição Genética para Doença , Mutação , Acetil-CoA C-Acetiltransferase/química , Acetil-CoA C-Acetiltransferase/metabolismo , Acetil-CoA C-Aciltransferase/genética , Acetil-CoA C-Aciltransferase/metabolismo , Erros Inatos do Metabolismo dos Aminoácidos/diagnóstico , Erros Inatos do Metabolismo dos Aminoácidos/metabolismo , Animais , Regulação Enzimológica da Expressão Gênica , Estudos de Associação Genética , Variação Genética , Humanos , Redes e Vias Metabólicas , Modelos Moleculares , Fenótipo , Ligação Proteica , Conformação Proteica , Domínios e Motivos de Interação entre Proteínas , Multimerização Proteica , Relação Estrutura-Atividade
4.
Int J Cardiol ; 292: 218-224, 2019 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-31023563

RESUMO

BACKGROUND: Epicardial adipose tissue (EAT) is a risk factor for cardiovascular diseases. Glucagon-like peptide 1 analogs (GLP-1A) may have beneficial cardiovascular effects and reduce EAT, possibly throughout targeting GLP-1 receptor (GLP-1R). Nevertheless, the role of EAT GLP-1R, GLP-2R and their interplay with EAT genes involved in adipogenesis and fatty acid (FA) metabolism are unknown. We analyzed whether EAT transcriptome is related to GLP-1R/GLP-2R gene expression, and GLP-1/GLP-2 plasma levels in coronary artery disease patients (CAD). METHODS: EAT was collected from 17 CAD patients undergoing CABG for microarray analysis of GLP-1R, GLP-2R and genes involved in FA metabolism and adipogenesis. EAT thickness was measured by echocardiography. GLP-1 and GLP-2 levels were quantified by ELISA in CAD and healthy subjects (CTR). RESULTS: EAT GLP-1R was directly correlated with genes promoting beta-oxidation and white-to-brown adipocyte differentiation, and inversely with pro-adipogenic genes. GLP-2R was positively correlated with genes involved in adipogenesis and lipid synthesis, and inversely with genes promoting beta-oxidation. GLP-1 and GLP-2 levels were higher in CAD than CTR and in patients with greater EAT thickness. CONCLUSIONS: GLP-1 analogs may target EAT GLP-1R and therefore reduce local adipogenesis, improve fat utilization and induce brown fat differentiation. As EAT lies in direct contiguity to myocardium and coronary arteries, the beneficial effects of GLP-1 activation may extent to the heart. The increased levels of circulating GLP-1 and GLP-2 and EAT GLP-2R may be compensatory mechanisms related to CAD and also EAT expansion, but the meaning of these observations needs to be further investigated.


Assuntos
3-Hidroxiacil-CoA Desidrogenases/metabolismo , Acetil-CoA C-Aciltransferase/metabolismo , Tecido Adiposo Marrom/metabolismo , Tecido Adiposo Branco/metabolismo , Isomerases de Ligação Dupla Carbono-Carbono/metabolismo , Doenças Cardiovasculares/sangue , Enoil-CoA Hidratase/metabolismo , Receptor do Peptídeo Semelhante ao Glucagon 1/sangue , Pericárdio/metabolismo , Racemases e Epimerases/metabolismo , 3-Hidroxiacil-CoA Desidrogenases/genética , Acetil-CoA C-Aciltransferase/genética , Tecido Adiposo Marrom/diagnóstico por imagem , Tecido Adiposo Branco/diagnóstico por imagem , Adulto , Idoso , Antropometria/métodos , Isomerases de Ligação Dupla Carbono-Carbono/genética , Doenças Cardiovasculares/diagnóstico por imagem , Doenças Cardiovasculares/genética , Enoil-CoA Hidratase/genética , Feminino , Receptor do Peptídeo Semelhante ao Glucagon 1/genética , Humanos , Masculino , Pessoa de Meia-Idade , Pericárdio/diagnóstico por imagem , Racemases e Epimerases/genética , Fatores de Risco
5.
Cell Biol Toxicol ; 35(5): 457-470, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-30721374

RESUMO

Silent information regulator 1 (SIRT1) is a nicotinamide adenine dinucleotide (NAD+)-dependent deacetylase, and the function is linked to cellular metabolism including mitochondrial biogenesis. Hepatic L-serine concentration is decreased significantly in fatty liver disease. We reported that the supplementation of the amino acid ameliorated the alcoholic fatty liver by enhancing L-serine-dependent homocysteine metabolism. In this study, we hypothesized that the metabolic production of NAD+ from L-serine and thus activation of SIRT1 contribute to the action of L-serine. To this end, we evaluated the effects of L-serine on SIRT1 activity and mitochondria biogenesis in C2C12 myotubes. L-Serine increased intracellular NAD+ content and led to the activation of SIRT1 as determined by p53 luciferase assay and western blot analysis of peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1α) acetylation. L-Serine treatment increased the expression of the genes associated with mitochondrial biogenesis and enhanced mitochondrial mass and function. In addition, L-serine reversed cellular insulin resistance determined by insulin-induced phosphorylation of Akt and GLUT4 expression and membrane translocation. L-Serine-induced mitochondrial gene expression, fatty acid oxidation, and insulin sensitization were mediated by enhanced SIRT1 activity, which was verified by selective SIRT1 inhibitor (Ex-527) and siRNA directed to SIRT1. L-Serine effect on cellular NAD+ level is dependent on the L-serine metabolism to pyruvate that is subsequently converted to lactate by lactate dehydrogenase. In summary, these data suggest that L-serine increases cellular NAD+ level and thus SIRT1 activity in C2C12 myotubes.


Assuntos
Ácidos Graxos/metabolismo , Resistência à Insulina/fisiologia , Fibras Musculares Esqueléticas/efeitos dos fármacos , Fibras Musculares Esqueléticas/metabolismo , Serina/farmacologia , Sirtuína 1/metabolismo , 3-Hidroxiacil-CoA Desidrogenases/metabolismo , Proteínas Quinases Ativadas por AMP/metabolismo , Acetil-CoA C-Aciltransferase/metabolismo , Acetilação , Animais , Isomerases de Ligação Dupla Carbono-Carbono/metabolismo , Linhagem Celular , Enoil-CoA Hidratase/metabolismo , Células Hep G2 , Humanos , Insulina/farmacologia , Metabolismo dos Lipídeos , Camundongos , Mitocôndrias/metabolismo , Fibras Musculares Esqueléticas/citologia , Músculo Esquelético/metabolismo , Oxirredução , Fosforilação , Racemases e Epimerases/metabolismo , Transdução de Sinais/efeitos dos fármacos , Transativadores/metabolismo , Fatores de Transcrição/metabolismo
6.
Biomed Pharmacother ; 112: 108668, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-30784937

RESUMO

Natural compounds are important resources for drug discovery. Using Caenorhabditis elegans (C. elegans) models, we screened active natural compounds with lipid lowering effects. Swertiamarin was found as a potent candidate to reduce lipid content in C. elegans. Using RNAi screening, we were able to demonstrate that kat-1 (ketoacyl thiolase-1) is necessary for the lipid lowering effect of swertiamarin. Furthermore, the activity of swertiamarin was verified in high fat diet induced obese mice. Consistent with the results in C. elegans, swertiamarin ameliorated high fat diet induced lipid deposition and hyperlipidemia. These results indicate that swertiamarin exerts lipid-lowering effects through kat-1 regulation and could serve as a possible therapeutic option to improve hyperlipidemia induced comorbidities.


Assuntos
Acetil-CoA C-Aciltransferase/metabolismo , Glucosídeos Iridoides/farmacologia , Produto da Acumulação Lipídica/efeitos dos fármacos , Obesidade/tratamento farmacológico , Pironas/farmacologia , Swertia , Animais , Animais Geneticamente Modificados , Caenorhabditis elegans , Dieta Hiperlipídica/efeitos adversos , Relação Dose-Resposta a Droga , Glucosídeos Iridoides/uso terapêutico , Produto da Acumulação Lipídica/fisiologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Obesidade/sangue , Obesidade/induzido quimicamente , Pironas/uso terapêutico
7.
Nat Commun ; 9(1): 79, 2018 01 08.
Artigo em Inglês | MEDLINE | ID: mdl-29311546

RESUMO

Aromatic polyesters are widely used plastics currently produced from petroleum. Here we engineer Escherichia coli strains for the production of aromatic polyesters from glucose by one-step fermentation. When the Clostridium difficile isocaprenoyl-CoA:2-hydroxyisocaproate CoA-transferase (HadA) and evolved polyhydroxyalkanoate (PHA) synthase genes are overexpressed in a D-phenyllactate-producing strain, poly(52.3 mol% 3-hydroxybutyrate (3HB)-co-47.7 mol% D-phenyllactate) can be produced from glucose and sodium 3HB. Also, various poly(3HB-co-D-phenyllactate) polymers having 11.0, 15.8, 20.0, 70.8, and 84.5 mol% of D-phenyllactate are produced from glucose as a sole carbon source by additional expression of Ralstonia eutropha ß-ketothiolase (phaA) and reductase (phaB) genes. Fed-batch culture of this engineered strain produces 13.9 g l-1 of poly(61.9 mol% 3HB-co-38.1 mol% D-phenyllactate). Furthermore, different aromatic polyesters containing D-mandelate and D-3-hydroxy-3-phenylpropionate are produced from glucose when feeding the corresponding monomers. The engineered bacterial system will be useful for one-step fermentative production of aromatic polyesters from renewable resources.


Assuntos
Escherichia coli/metabolismo , Fermentação , Glucose/metabolismo , Engenharia Metabólica/métodos , Poliésteres/metabolismo , Ácido 3-Hidroxibutírico/metabolismo , Acetil-CoA C-Aciltransferase/genética , Acetil-CoA C-Aciltransferase/metabolismo , Aciltransferases/genética , Aciltransferases/metabolismo , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , /genética , Coenzima A-Transferases/genética , Coenzima A-Transferases/metabolismo , Cupriavidus necator/enzimologia , Cupriavidus necator/genética , Escherichia coli/genética , Lactatos/metabolismo , Oxirredutases/genética , Oxirredutases/metabolismo , Polietilenotereftalatos/metabolismo , Polímeros/metabolismo
8.
Sci Rep ; 8(1): 417, 2018 01 11.
Artigo em Inglês | MEDLINE | ID: mdl-29323178

RESUMO

Mammary epithelial cells (MECs) affect milk production capacity during lactation and are critical for the maintenance of tissue homeostasis. Our previous studies have revealed that the expression of miR-152 was increased significantly in MECs of cows with high milk production. In the present study, bioinformatics analysis identified ACAA2 and HSD17B12 as the potential targets of miR-152, which were further validated by dual-luciferase repoter assay. In addition, the expressions of miR-152 was shown to be negatively correlated with levels of mRNA and protein of ACAA2, HSD17B12 genes by qPCR and western bot analysis. Furthermore, transfection with miR-152 significantly up-regulated triglyceride production, promoted proliferation and inhibited apoptosis in MECs. Furthermore, overexpression of ACAA2 and HSD17B12 could inhibit triglyceride production, cells proliferation and induce apoptosis; but sh234-ACAA2-181/sh234-HSD17B12-474 could reverse the trend. These findings suggested that miR-152 could significantly influence triglyceride production and suppress apoptosis, possibly via the expression of target genes ACAA2 and HSD17B12.


Assuntos
17-Hidroxiesteroide Desidrogenases/genética , Acetil-CoA C-Aciltransferase/genética , Glândulas Mamárias Animais/citologia , MicroRNAs/genética , Triglicerídeos/metabolismo , 17-Hidroxiesteroide Desidrogenases/metabolismo , Regiões 3' não Traduzidas , Acetil-CoA C-Aciltransferase/metabolismo , Animais , Apoptose , Bovinos , Proliferação de Células , Células Epiteliais/citologia , Células Epiteliais/metabolismo , Feminino , Regulação da Expressão Gênica , Lactação , Glândulas Mamárias Animais/metabolismo
9.
Biochemistry ; 57(22): 3155-3166, 2018 06 05.
Artigo em Inglês | MEDLINE | ID: mdl-29381332

RESUMO

Thiolases are a class of carbon-carbon bond forming enzymes with important applications in biotechnology and metabolic engineering as they provide a general method for the condensation of two acyl coenzyme A (CoA) substrates. As such, developing a greater understanding of their substrate selectivity would expand our ability to engineer the enzymatic or microbial production of a broad range of small-molecule targets. Here, we report the crystal structures and biochemical characterization of Acat2 and Acat5, two biosynthetic thiolases from Ascaris suum with varying selectivity toward branched compared to linear compounds. The structure of the Acat2-C91S mutant bound to propionyl-CoA shows that the terminal methyl group of the substrate, representing the α-branch point, is directed toward the conserved Phe 288 and Met 158 residues. In Acat5, the Phe ring is rotated to accommodate a hydroxyl-π interaction with an adjacent Thr side chain, decreasing space in the binding pocket and possibly accounting for its strong preference for linear substrates compared to Acat2. Comparison of the different Acat thiolase structures shows that Met 158 is flexible, adopting alternate conformations with the side chain rotated toward or away from a covering loop at the back of the active site. Mutagenesis of residues in the covering loop in Acat5 with the corresponding residues from Acat2 allows for highly increased accommodation of branched substrates, whereas the converse mutations do not significantly affect Acat2 substrate selectivity. Our results suggest an important contribution of second-shell residues to thiolase substrate selectivity and offer insights into engineering this enzyme class.


Assuntos
Acetil-CoA C-Aciltransferase/metabolismo , Ascaris suum/enzimologia , Acetil-CoA C-Aciltransferase/fisiologia , Sequência de Aminoácidos , Animais , Ascaris suum/fisiologia , Sítios de Ligação , Domínio Catalítico/fisiologia , Cinética , Modelos Moleculares , Conformação Proteica , Especificidade por Substrato/fisiologia
10.
Metab Brain Dis ; 32(6): 2063-2071, 2017 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-28875337

RESUMO

Hydroxysteroid (17ß) dehydrogenase 10 (HSD10) and mitochondrial acetoacetyl-CoA thiolase (ß-KT) are two adjacent enzymes for the degradation of isoleucine, thus HSD10 and ß-KT deficiencies are confusing at an early stage because of nearly the same elevation of typical metabolites in urine, such as 2-methyl-3-hydroxybutyric acid (2M3HBA) and tiglylglycine (TG). In order to better understand the differences between these two disorders, we described the clinical and molecular characteristics of two HSD10 deficiency patients and four ß-KT deficiency patients. ß-KT deficiency patients had a much more favorable outcome than that of HSD10 deficiency patients, indicating that the multifunction of HSD10, especially neurosteroid metabolic activity, other than only enzymatic degradation of isoleucine, is involved in the pathogenesis of HSD10 deficiency. Two different mutations, a novel mutation p.Ile175Met and a reported mutation p.Arg226Gln, were detected in the HSD17B10 gene of HSD10 deficiency patients. Six different mutations, including four known mutations: p.Ala333Pro, p.Thr297Lys, c.83_84delAT, c.1006-1G > C, and two novel mutations: p.Thr277Pro and c.121-3C > G were identified in the ACAT1 gene of ß-KT deficiency patients. In general, DNA diagnosis played an important role in distinguishing between these two disorders.


Assuntos
3-Hidroxiacil-CoA Desidrogenases/genética , Acetil-CoA C-Acetiltransferase/genética , Acetil-CoA C-Aciltransferase/deficiência , Erros Inatos do Metabolismo dos Aminoácidos/diagnóstico , Discinesias/diagnóstico , Epilepsia/genética , Isoleucina/metabolismo , Retardo Mental Ligado ao Cromossomo X/diagnóstico , Acetil-CoA C-Aciltransferase/genética , Acetil-CoA C-Aciltransferase/metabolismo , Erros Inatos do Metabolismo dos Aminoácidos/diagnóstico por imagem , Erros Inatos do Metabolismo dos Aminoácidos/genética , Erros Inatos do Metabolismo dos Aminoácidos/metabolismo , Encéfalo/diagnóstico por imagem , Pré-Escolar , China , Diagnóstico Diferencial , Discinesias/diagnóstico por imagem , Discinesias/genética , Discinesias/metabolismo , Epilepsia/metabolismo , Feminino , Humanos , Lactente , Masculino , Retardo Mental Ligado ao Cromossomo X/diagnóstico por imagem , Retardo Mental Ligado ao Cromossomo X/genética , Retardo Mental Ligado ao Cromossomo X/metabolismo , Modelos Moleculares , Mutação , Estudos Retrospectivos
11.
Cancer Lett ; 409: 104-115, 2017 11 28.
Artigo em Inglês | MEDLINE | ID: mdl-28923398

RESUMO

The class III deacetylase sirtuin 1 (SIRT1), a member of the sirtuin family proteins, plays a key role in many types of cancers including colorectal cancer (CRC). Here we report that SIRT1 suppressed CRC metastasis in vitro and in vivo as a negative regulator for miR-15b-5p transcription. Mechanistically, SIRT1 impaired regulatory effects of activator protein (AP-1) on miR-15b-5p trans-activation through deacetylation of AP-1. Importantly, acyl-CoA oxidase 1 (ACOX1), a key enzyme of the fatty acid oxidation (FAO) pathway, was found as a direct target for miR-15b-5p. SIRT1 expression was positively correlated with ACOX1 expression in CRC cells and in xenografts. Moreover, ACOX1 overexpression attenuated the augmentation of migration and invasion of CRC cells by miR-15b-5p overexpression. In conclusion, our study demonstrated a functional role of the SIRT1/miR-15b-5p/ACOX1 axis in CRC metastasis and suggested a potential target for metastatic CRC therapy.


Assuntos
Neoplasias Colorretais/genética , MicroRNAs/genética , Sirtuína 1/genética , 3-Hidroxiacil-CoA Desidrogenases/metabolismo , Acetil-CoA C-Aciltransferase/metabolismo , Animais , Células CACO-2 , Isomerases de Ligação Dupla Carbono-Carbono/metabolismo , Linhagem Celular Tumoral , Neoplasias Colorretais/metabolismo , Neoplasias Colorretais/patologia , Enoil-CoA Hidratase/metabolismo , Células HCT116 , Células HT29 , Xenoenxertos , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , MicroRNAs/metabolismo , Metástase Neoplásica , Racemases e Epimerases/metabolismo , Transdução de Sinais , Sirtuína 1/metabolismo , Transcrição Genética , Transfecção
12.
Appl Microbiol Biotechnol ; 101(21): 7945-7960, 2017 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-28956111

RESUMO

The actinomycete Gordonia polyisoprenivorans strain VH2 is well-known for its ability to efficiently degrade and catabolize natural rubber [poly(cis-1,4-isoprene)]. Recently, a pathway for the catabolism of rubber by strain VH2 was postulated based on genomic data and the analysis of mutants (Hiessl et al. in Appl Environ Microbiol 78:2874-2887, 2012). To further elucidate the degradation pathway of poly(cis-1,4-isoprene), 2-dimensional-polyacrylamide gel electrophoresis was performed. The analysis of the identified protein spots by matrix-assisted laser desorption/ionization-time of flight tandem mass spectrometry confirmed the postulated intracellular pathway suggesting a degradation of rubber via ß-oxidation. In addition, other valuable information on rubber catabolism of G. polyisoprenivorans strain VH2 (e.g. oxidative stress response) was provided. Identified proteins, which were more abundant in cells grown with rubber than in cells grown with propionate, implied a putative long-chain acyl-CoA-dehydrogenase, a 3-ketoacyl-CoA-thiolase, and an aldehyde dehydrogenase. The amino acid sequence of the latter showed a high similarity towards geranial dehydrogenases. The expression of the corresponding gene was upregulated > 10-fold under poly(cis-1,4-isoprene)-degrading conditions. The putative geranial dehydrogenase and a homolog were purified and used for enzyme assays. Deletion mutants for five aldehyde dehydrogenases were generated, and growth with poly(cis-1,4-isoprene) was investigated. While none of the mutants had an altered phenotype regarding growth with poly(cis-1,4-isoprene) as sole carbon and energy source, purified aldehyde dehydrogenases were able to catalyze the oxidation of oligoisoprene aldehydes indicating an involvement in rubber degradation.


Assuntos
Aldeídos/metabolismo , Gordonia (Bactéria)/enzimologia , Gordonia (Bactéria)/metabolismo , Hemiterpenos/metabolismo , Látex/metabolismo , Oxirredutases/metabolismo , Acetil-CoA C-Aciltransferase/genética , Acetil-CoA C-Aciltransferase/metabolismo , Aldeído Desidrogenase/genética , Aldeído Desidrogenase/metabolismo , Carbono/metabolismo , Eletroforese em Gel Bidimensional , Metabolismo Energético , Deleção de Genes , Perfilação da Expressão Gênica , Gordonia (Bactéria)/genética , 3-Hidroxiacil-CoA Desidrogenase de Cadeia Longa/genética , 3-Hidroxiacil-CoA Desidrogenase de Cadeia Longa/metabolismo , Redes e Vias Metabólicas/genética , Oxirredução , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz , Espectrometria de Massas em Tandem
13.
Microb Cell Fact ; 16(1): 144, 2017 Aug 17.
Artigo em Inglês | MEDLINE | ID: mdl-28818103

RESUMO

BACKGROUND: In recent years the production of biobased biodegradable plastics has been of interest of researchers partly due to the accumulation of non-biodegradable plastics in the environment and to the opportunity for new applications. Commonly investigated are the polyhydroxyalkanoates (PHAs) poly(hydroxybutyrate) and poly(hydroxybutyrate-co-hydroxyvalerate) (PHB-V). The latter has the advantage of being tougher and less brittle. The production of these polymers in bacteria is well established but production in yeast may have some advantages, e.g. the ability to use a broad spectrum of industrial by-products as a carbon sources. RESULTS: In this study we increased the synthesis of PHB-V in the non-conventional yeast Arxula adeninivorans by stabilization of polymer accumulation via genetic modification and optimization of culture conditions. An A. adeninivorans strain with overexpressed PHA pathway genes for ß-ketothiolase, acetoacetyl-CoA reductase, PHAs synthase and the phasin gene was able to accumulate an unexpectedly high level of polymer. It was found that an optimized strain cultivated in a shaking incubator is able to produce up to 52.1% of the DCW of PHB-V (10.8 g L-1) with 12.3%mol of PHV fraction. Although further optimization of cultivation conditions in a fed-batch bioreactor led to lower polymer content (15.3% of the DCW of PHB-V), the PHV fraction and total polymer level increased to 23.1%mol and 11.6 g L-1 respectively. Additionally, analysis of the product revealed that the polymer has a very low average molecular mass and unexpected melting and glass transition temperatures. CONCLUSIONS: This study indicates a potential of use for the non-conventional yeast, A. adeninivorans, as an efficient producer of polyhydroxyalkanoates.


Assuntos
Poliésteres/metabolismo , Saccharomycetales/metabolismo , Acetil-CoA C-Aciltransferase/genética , Acetil-CoA C-Aciltransferase/metabolismo , Aciltransferases/genética , Aciltransferases/metabolismo , Oxirredutases do Álcool/genética , Oxirredutases do Álcool/metabolismo , Técnicas de Cultura Celular por Lotes , Reatores Biológicos , Proteínas Fúngicas/metabolismo , Cromatografia Gasosa-Espectrometria de Massas , Microscopia Eletrônica de Transmissão , Lectinas de Plantas/genética , Lectinas de Plantas/metabolismo , Plasmídeos/genética , Plasmídeos/metabolismo , Poliésteres/análise , Poliésteres/química , Saccharomycetales/enzimologia , Saccharomycetales/crescimento & desenvolvimento
14.
Mol Nutr Food Res ; 61(10)2017 10.
Artigo em Inglês | MEDLINE | ID: mdl-28608394

RESUMO

SCOPE: The aim of this work was to study the urinary metabolomics changes of participants that consumed beer, nonalcoholic beer (na-beer), and gin. METHODS AND RESULTS: Thirty-three males at high cardiovascular risk between 55 and 75 years old participated in an open, randomized, crossover, controlled trial with three nutritional interventions consisting of beer, na-beer, and gin for 4 wk. Diet and physical activity was monitored throughout the study and compliance was assessed by measurement of urinary isoxanthohumol. Metabolomic analysis was performed in urine samples by LC coupled to an LTQ-Orbitrap mass spectrometer combined with univariate and multivariate statistical analysis. Ten metabolites were identified. Eight were exogenous metabolites related to beer, na-beer, or gin consumption, but two of them were related to endogenic changes: hydroxyadipic acid linked to fatty acid oxidation, and 4-guanidinobutanoic acid, which correlated with a decrease in urinary creatinine. Plasmatic acylcarnitines were quantified by targeted MS. A regular and moderate consumption of beer and na-beer decreased stearoylcarnitine concentrations. CONCLUSION: Humulinone and 2,3-dihydroxy-3-methylvaleric acid showed to be potential biomarkers of beer and na-beer consumption. Moreover, the results of this trial provide new evidence that the nonalcoholic fraction of beer may increase fatty oxidation.


Assuntos
Cerveja/efeitos adversos , Biomarcadores/urina , Doenças Cardiovasculares/urina , Metaboloma , Metabolômica , 3-Hidroxiacil-CoA Desidrogenases/metabolismo , Acetil-CoA C-Aciltransferase/metabolismo , Adipatos/sangue , Idoso , Consumo de Bebidas Alcoólicas , Bebidas , Isomerases de Ligação Dupla Carbono-Carbono/metabolismo , Carnitina/análogos & derivados , Carnitina/sangue , Creatinina/urina , Estudos Cross-Over , Dieta , Enoil-CoA Hidratase/metabolismo , Exercício Físico , Humanos , Masculino , Pessoa de Meia-Idade , Cooperação do Paciente , Ácidos Pentanoicos/urina , Racemases e Epimerases/metabolismo , Fatores de Risco , Xantonas/urina
15.
Arch Pediatr ; 24(8): 777-782, 2017 Aug.
Artigo em Francês | MEDLINE | ID: mdl-28647472

RESUMO

Acute fatty liver of pregnancy (AFLP) is a rare liver disease unique to pregnancy that can lead to acute liver failure. The prognosis, initially often fatal for both mother and child, has been improved by prompt delivery. The diagnosis should be highly suspected if the mother presents epigastric pain, nausea and/or vomiting, or polyuria-polydipsia in the third trimester of pregnancy. AFLP has been found associated with a genetic deficiency of fatty acid beta-oxidation, which may cause sudden death in infancy. Consequently, the mother and her newborn should undergo screening for this deficiency.


Assuntos
Parto Obstétrico , Ácidos Graxos/metabolismo , Fígado Gorduroso/diagnóstico , Fígado Gorduroso/metabolismo , Mitocôndrias/metabolismo , Complicações na Gravidez/diagnóstico , Complicações na Gravidez/metabolismo , Terceiro Trimestre da Gravidez , 3-Hidroxiacil-CoA Desidrogenases/metabolismo , Acetil-CoA C-Aciltransferase/metabolismo , Adulto , Isomerases de Ligação Dupla Carbono-Carbono/metabolismo , Enoil-CoA Hidratase/metabolismo , Medicina Baseada em Evidências , Fígado Gorduroso/genética , Fígado Gorduroso/terapia , Feminino , Humanos , Recém-Nascido , Gravidez , Complicações na Gravidez/genética , Complicações na Gravidez/terapia , Resultado da Gravidez , Prognóstico , Racemases e Epimerases/metabolismo , Fatores de Risco
16.
Mol Biosyst ; 13(8): 1504-1511, 2017 Jul 25.
Artigo em Inglês | MEDLINE | ID: mdl-28632266

RESUMO

Xuesaitong injection (XST), which mainly consists of Panax notoginseng saponins, has been widely used for treating cardio-cerebral vascular diseases. However, the underlying mechanisms of XST associated with its cardioprotective effects are still unclear. To identify the potential target proteins of XST, two-dimensional gel electrophoresis (2-DE)-based proteomics was utilized to analyze the protein profile of myocardium in rats with myocardial ischemia/reperfusion (I/R) injury. The differentially expressed proteins were identified by matrix assisted laser desorption/ionization time-of-flight mass spectrometry. It is interesting that XST can alter the expression of 7 proteins, including pyruvate dehydrogenase E1 alpha (PDHA1), hydroxyacyl-coenzyme A dehydrogenase (HADHA), peroxiredoxin 3 (PRX3), gamma-enolase, acetyl-coenzyme A acyltransferase 2 (ACAA2), etc. Functional analysis revealed that those proteins were chiefly related to cardiac energy metabolism and oxidative stress. The cardioprotective effects of XST were further validated in H9c2 cardiac muscle cells with hypoxia/reoxygenation injury. We found that XST can promote the activity of PDH, an important enzyme related to the TCA cycle, as well as increase the intracellular content of acetyl-CoA and ATP. Moreover, XST also attenuated intracellular MDA release in H2O2-injured cardiac cells. This is the first study on the proteomic expression of XST-treated myocardium with I/R injury to reveal that the cardioprotective effects of XST may be attributed to the PDH-mediated restoration of aerobic glucose oxidation.


Assuntos
Fármacos Cardiovasculares/farmacologia , Medicamentos de Ervas Chinesas/farmacologia , Metabolismo Energético/efeitos dos fármacos , Regulação da Expressão Gênica , Traumatismo por Reperfusão Miocárdica/tratamento farmacológico , Fármacos Neuroprotetores/farmacologia , Saponinas/farmacologia , Acetil-CoA C-Aciltransferase/genética , Acetil-CoA C-Aciltransferase/metabolismo , Animais , Linhagem Celular , Metabolismo Energético/genética , Perfilação da Expressão Gênica , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Masculino , Subunidade alfa da Proteína Mitocondrial Trifuncional/genética , Subunidade alfa da Proteína Mitocondrial Trifuncional/metabolismo , Traumatismo por Reperfusão Miocárdica/genética , Traumatismo por Reperfusão Miocárdica/metabolismo , Traumatismo por Reperfusão Miocárdica/patologia , Miocárdio/metabolismo , Miocárdio/patologia , Miócitos Cardíacos/efeitos dos fármacos , Miócitos Cardíacos/metabolismo , Miócitos Cardíacos/patologia , Estresse Oxidativo , Fosfopiruvato Hidratase/genética , Fosfopiruvato Hidratase/metabolismo , Piruvato Desidrogenase (Lipoamida)/genética , Piruvato Desidrogenase (Lipoamida)/metabolismo , Ratos , Ratos Sprague-Dawley , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz
17.
Metab Eng ; 42: 33-42, 2017 07.
Artigo em Inglês | MEDLINE | ID: mdl-28550000

RESUMO

ß-Oxidation is the ubiquitous metabolic strategy to break down fatty acids. In the course of this four-step process, two carbon atoms are liberated per cycle from the fatty acid chain in the form of acetyl-CoA. However, typical ß-oxidative strategies are not restricted to monocarboxylic (fatty) acid degradation only, but can also be involved in the utilization of aromatic compounds, amino acids and dicarboxylic acids. Each enzymatic step of a typical ß-oxidation cycle is reversible, offering the possibility to also take advantage of reversed metabolic pathways for applied purposes. In such cases, 3-oxoacyl-CoA thiolases, which catalyze the final chain-shortening step in the catabolic direction, mediate the condensation of an acyl-CoA starter molecule with acetyl-CoA in the anabolic direction. Subsequently, the carbonyl-group at C3 is stepwise reduced and dehydrated yielding a chain-elongated product. In the last years, several ß-oxidation pathways have been studied in detail and reversal of these pathways already proved to be a promising strategy for the production of chemicals and polymer building blocks in several industrially relevant microorganisms. This review covers recent advancements in this field and discusses constraints and bottlenecks of this metabolic strategy in comparison to alternative production pathways.


Assuntos
Acetilcoenzima A/metabolismo , Acetil-CoA C-Aciltransferase/metabolismo , Bactérias/metabolismo , Proteínas de Bactérias/metabolismo , Plásticos Biodegradáveis/metabolismo , Acetilcoenzima A/genética , Acetil-CoA C-Aciltransferase/genética , Bactérias/genética , Proteínas de Bactérias/genética , Oxirredução
18.
PLoS Comput Biol ; 13(4): e1005461, 2017 04.
Artigo em Inglês | MEDLINE | ID: mdl-28369071

RESUMO

Mitochondrial fatty-acid beta-oxidation (mFAO) plays a central role in mammalian energy metabolism. Multiple severe diseases are associated with defects in this pathway. Its kinetic structure is characterized by a complex wiring of which the functional implications have hardly been explored. Repetitive cycles of reversible reactions, each cycle shortening the fatty acid by two carbon atoms, evoke competition between intermediates of different chain lengths for a common set of 'promiscuous' enzymes (enzymes with activity towards multiple substrates). In our validated kinetic model of the pathway, substrate overload causes a steep and detrimental flux decline. Here, we unravel the underlying mechanism and the role of enzyme promiscuity in it. Comparison of alternative model versions elucidated the role of promiscuity of individual enzymes. Promiscuity of the last enzyme of the pathway, medium-chain ketoacyl-CoA thiolase (MCKAT), was both necessary and sufficient to elicit the flux decline. Subsequently, Metabolic Control Analysis revealed that MCKAT had insufficient capacity to cope with high substrate influx. Next, we quantified the internal metabolic regulation, revealing a vicious cycle around MCKAT. Upon substrate overload, MCKAT's ketoacyl-CoA substrates started to accumulate. The unfavourable equilibrium constant of the preceding enzyme, medium/short-chain hydroxyacyl-CoA dehydrogenase, worked as an amplifier, leading to accumulation of upstream CoA esters, including acyl-CoA esters. These acyl-CoA esters are at the same time products of MCKAT and inhibited its already low activity further. Finally, the accumulation of CoA esters led to a sequestration of free CoA. CoA being a cofactor for MCKAT, its sequestration limited the MCKAT activity even further, thus completing the vicious cycle. Since CoA is also a substrate for distant enzymes, it efficiently communicated the 'traffic jam' at MCKAT to the entire pathway. This novel mechanism provides a basis to explore the role of mFAO in disease and elucidate similar principles in other pathways of lipid metabolism.


Assuntos
Acetil-CoA C-Aciltransferase/metabolismo , Ácidos Graxos/metabolismo , Redes e Vias Metabólicas/fisiologia , Acetil-CoA C-Aciltransferase/fisiologia , Biologia Computacional , Simulação por Computador , Cinética , Oxirredução
19.
J Inherit Metab Dis ; 40(3): 415-422, 2017 05.
Artigo em Inglês | MEDLINE | ID: mdl-28255778

RESUMO

BACKGROUND: Mitochondrial acetoacetyl-CoA thiolase (T2) deficiency affects ketone body and isoleucine catabolism. Neurological impairment may occur secondary to ketoacidotic episodes. However, we observed neuromotor abnormalities without ketoacidotic events in two T2-deficient families. We hypothesized that the neurological signs were related to the genetic defect and may occur independently of ketoacidotic episodes. We therefore conducted a retrospective review on a French T2-deficient patient series searching for neuromotor impairment. METHODS: In total, 26 cases were retrospectively analysed for clinical, biological and neuroimaging data. RESULTS: Neurological findings were observed for 6/26 (23%) patients. Among these, two had never experienced ketoacidotic episodes, though they developed extrapyramidal signs with putamen involvement. Two of the other four patients developed neurological abnormalities before the first ketoacidotic crisis, with putamen involvement in one case. The third patient developed extrapyramidal symptoms more than 10 years after the initial decompensation with globus pallidus involvement. The last patient developed extrapyramidal signs immediately after a severe ketoacidotic crisis with putaminal lesions. CONCLUSIONS: Most T2-deficient patients achieved normal neurodevelopment. However, on account of the role of T2 in isoleucine catabolism, these patients are potentially exposed to accumulation of toxic isoleucine-derived metabolites, which may contribute to neurological impairment. Our findings confirm previous observations that neurological symptoms in T2 deficiency may occur unrelated to ketoacidosis. The role of protein restriction as a preventive measure against neurological symptoms could not be established in this study and deserves further evaluation. Long-term follow-up data on children diagnosed by newborn screening may clarify the pathogenesis of this neurometabolic association.


Assuntos
Acetil-CoA C-Acetiltransferase/deficiência , Acetil-CoA C-Aciltransferase/deficiência , Erros Inatos do Metabolismo dos Aminoácidos/metabolismo , Gânglios da Base/metabolismo , Cetose/metabolismo , Mitocôndrias/metabolismo , Acetil-CoA C-Aciltransferase/metabolismo , Adolescente , Adulto , Criança , Pré-Escolar , Feminino , Humanos , Lactente , Recém-Nascido , Isoleucina/metabolismo , Corpos Cetônicos/metabolismo , Masculino , Triagem Neonatal/métodos , Estudos Retrospectivos , Adulto Jovem
20.
Int J Cardiol ; 234: 1-6, 2017 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-28256321

RESUMO

Trimetazidine (TMZ) has traditionally been used as an anti-ischemic drug for coronary artery disease by selectively inhibiting the mitochondrial long-chain 3-ketoacyl-CoA thiolase. Recently, new applications for this therapy have been investigated. This article reviews alternative uses for TMZ in non-coronary artery diseases, such as non-ischemic cardiomyopathy, sepsis, myocardial dysfunction induced by anti-cancer drugs, diabetic cardiomyopathy and contrast-induced nephropathy.


Assuntos
Antineoplásicos/efeitos adversos , Distúrbios Induzidos Quimicamente/tratamento farmacológico , Doença da Artéria Coronariana , Cardiomiopatias Diabéticas/tratamento farmacológico , Nefropatias , Trimetazidina/farmacologia , Acetil-CoA C-Aciltransferase/metabolismo , Distúrbios Induzidos Quimicamente/etiologia , Doença da Artéria Coronariana/tratamento farmacológico , Doença da Artéria Coronariana/metabolismo , Reposicionamento de Medicamentos , Humanos , Nefropatias/induzido quimicamente , Nefropatias/tratamento farmacológico , Mitocôndrias Cardíacas/metabolismo , Vasodilatadores/farmacologia
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