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1.
Am J Alzheimers Dis Other Demen ; 37: 15333175221124949, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-36113018

RESUMO

The ketone bodies, especially the ß-hydroxybutyrate, had been shown to modulate the function of the central nervous system and prevent the pathological progression of Alzheimer's disease (AD). However, little is known about the role of acetoacetate in the AD brain. Thus, we intraventricularly injected acetoacetate into familial AD mice (APPSWE) for 14 days and monitored their memory and biochemical changes. During the behavior test, acetoacetate at 100 mg/kg led to significant improvement in both Y-maze and novel object recognition tests (NORTs) (both P < .05), indicating ameliorating spatial and recognition memory, respectively. Biomedical tests revealed two mechanisms were involved. Firstly, acetoacetate inhibited the GPR43-pERK pathway, which led to apparent inhibition in tumor necrosis factor-α and Interleukin-6 expression in the hippocampus in a concentration-dependent manner. Secondarily, acetoacetate stimulated the expression of hippocampal brain-derived neurotrophic factor (BDNF). We concluded that acetoacetate could ameliorate AD symptoms and exhibited promising features as a therapeutic for AD.


Assuntos
Doença de Alzheimer , Fator Neurotrófico Derivado do Encéfalo , Ácido 3-Hidroxibutírico , Acetoacetatos/uso terapêutico , Animais , Inflamação/tratamento farmacológico , Interleucina-6/uso terapêutico , Camundongos , Fator de Necrose Tumoral alfa/uso terapêutico
2.
Exp Mol Med ; 54(8): 1125-1132, 2022 08.
Artigo em Inglês | MEDLINE | ID: mdl-35931735

RESUMO

Recent investigations have revealed that the human microbiome plays an essential role in the occurrence of type 2 diabetes (T2D). However, despite the importance of understanding the involvement of the microbiota throughout the body in T2D, most studies have focused specifically on the intestinal microbiota. Extracellular vesicles (EVs) have been recently found to provide important evidence regarding the mechanisms of T2D pathogenesis, as they act as key messengers between intestinal microorganisms and the host. Herein, we explored microorganisms potentially associated with T2D by tracking changes in microbiota-derived EVs from patient urine samples collected three times over four years. Mendelian randomization analysis was conducted to evaluate the causal relationships among microbial organisms, metabolites, and clinical measurements to provide a comprehensive view of how microbiota can influence T2D. We also analyzed EV-derived metagenomic (N = 393), clinical (N = 5032), genomic (N = 8842), and metabolite (N = 574) data from a prospective longitudinal Korean community-based cohort. Our data revealed that GU174097_g, an unclassified Lachnospiraceae, was associated with T2D (ß = -189.13; p = 0.00006), and it was associated with the ketone bodies acetoacetate and 3-hydroxybutyrate (r = -0.0938 and -0.0829, respectively; p = 0.0022 and 0.0069, respectively). Furthermore, a causal relationship was identified between acetoacetate and HbA1c levels (ß = 0.0002; p = 0.0154). GU174097_g reduced ketone body levels, thus decreasing HbA1c levels and the risk of T2D. Taken together, our findings indicate that GU174097_g may lower the risk of T2D by reducing ketone body levels.


Assuntos
Diabetes Mellitus Tipo 2 , Microbiota , Acetoacetatos , Diabetes Mellitus Tipo 2/complicações , Hemoglobina A Glicada , Humanos , Estudos Longitudinais , Estudos Prospectivos
3.
J Org Chem ; 87(15): 10241-10249, 2022 08 05.
Artigo em Inglês | MEDLINE | ID: mdl-35849640

RESUMO

Two new complementary Au(I)-catalyzed methods for the preparation of ester-substituted indolizines from easily accessible 2-propargyloxypyridines and either acetoacetates or dimethyl malonate are reported. These reactions tolerate a wide range of functionality, allowing for diversification at three distinct positions of the product (R, R1, R2). For electron-poor substrates, the highest yields are observed upon reaction with acetoacetates, while neutral and electron-rich substrates give higher yields upon treatment with dimethyl malonate.


Assuntos
Indolizinas , Acetoacetatos , Catálise , Ciclização , Ésteres
4.
Respir Res ; 23(1): 172, 2022 Jun 27.
Artigo em Inglês | MEDLINE | ID: mdl-35761396

RESUMO

BACKGROUND: Acute respiratory distress syndrome (ARDS) is a challenging clinical problem. Discovering the potential metabolic alterations underlying the ARDS is important to identify novel therapeutic target and improve the prognosis. Serum and urine metabolites can reflect systemic and local changes and could help understanding metabolic characterization of community-acquired pneumonia (CAP) with ARDS. METHODS: Clinical data of patients with suspected CAP at the First Affiliated Hospital of Wenzhou Medical University were collected from May 2020 to February 2021. Consecutive patients with CAP were enrolled and divided into two groups: CAP with and without ARDS groups. 1H nuclear magnetic resonance-based metabolomics analyses of serum and urine samples were performed before and after treatment in CAP with ARDS (n = 43) and CAP without ARDS (n = 45) groups. Differences metabolites were identifed in CAP with ARDS. Furthermore, the receiver operating characteristic (ROC) curve was utilized to identify panels of significant metabolites for evaluating therapeutic effects on CAP with ARDS. The correlation heatmap was analyzed to further display the relationship between metabolites and clinical characteristics. RESULTS: A total of 20 and 42 metabolites were identified in the serum and urine samples, respectively. Serum metabolic changes were mainly involved in energy, lipid, and amino acid metabolisms, while urine metabolic changes were mainly involved in energy metabolism. Elevated levels of serum 3-hydroxybutyrate, lactate, acetone, acetoacetate, and decreased levels of serum leucine, choline, and urine creatine and creatinine were detected in CAP with ARDS relative to CAP without ARDS. Serum metabolites 3-hydroxybutyrate, acetone, acetoacetate, citrate, choline and urine metabolite 1-methylnicotinamide were identified as a potential biomarkers for assessing therapeutic effects on CAP with ARDS, and with AUCs of 0.866 and 0.795, respectively. Moreover, the ROC curve analysis revealed that combined characteristic serum and urine metabolites exhibited a better classification system for assessing therapeutic effects on CAP with ARDS, with a AUC value of 0.952. In addition, differential metabolites strongly correlated with clinical parameters in patients with CAP with ARDS. CONCLUSIONS: Serum- and urine-based metabolomics analyses identified characteristic metabolic alterations in CAP with ARDS and might provide promising circulatory markers for evaluating therapeutic effects on CAP with ARDS.


Assuntos
Infecções Comunitárias Adquiridas , Pneumonia , Síndrome do Desconforto Respiratório , Ácido 3-Hidroxibutírico , Acetoacetatos , Acetona , Biomarcadores , Infecções Comunitárias Adquiridas/diagnóstico , Humanos , Espectroscopia de Ressonância Magnética/métodos , Metabolômica/métodos , Pneumonia/diagnóstico por imagem , Curva ROC , Síndrome do Desconforto Respiratório/diagnóstico por imagem
5.
Carbohydr Polym ; 289: 119468, 2022 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-35483865

RESUMO

In the present work, we constructed a methodology to graft starch with special groups, such as alkyl, phenolic, naphthalimide derivatives (ND) and polymer, by a simple reaction under generally mild conditions without catalysts or UV irradiation, based on precursor starch acetoacetate (SAA). The completeness of these reactions has been proved to be ideal. After grafting, the starch derivatives have some common changes, for instance, their solubility is improved in certain solvents. On the other hand, the introduction of different functional groups will also bring some characteristics to the derivatives (e.g. ND brings fluorescence). In addition, part of the derivatives shows excellent machinability, and their hot-pressed samples exhibit great transparency and mechanical strength. Specially, the alkyl grafted starch displays excellent toughness, properties of deformation and self-recovery. In conclusion, this method has good universality and methodological significance, and offers insights into the larger-scale industrial application of starch.


Assuntos
Acetoacetatos , Amido , Polímeros , Solubilidade , Solventes
6.
J Biol Chem ; 298(5): 101884, 2022 05.
Artigo em Inglês | MEDLINE | ID: mdl-35367206

RESUMO

2-Ketopropyl-coenzyme M oxidoreductase/carboxylase (2-KPCC) is a member of the flavin and cysteine disulfide containing oxidoreductase family (DSOR) that catalyzes the unique reaction between atmospheric CO2 and a ketone/enolate nucleophile to generate acetoacetate. However, the mechanism of this reaction is not well understood. Here, we present evidence that 2-KPCC, in contrast to the well-characterized DSOR enzyme glutathione reductase, undergoes conformational changes during catalysis. Using a suite of biophysical techniques including limited proteolysis, differential scanning fluorimetry, and native mass spectrometry in the presence of substrates and inhibitors, we observed conformational differences between different ligand-bound 2-KPCC species within the catalytic cycle. Analysis of site-specific amino acid variants indicated that 2-KPCC-defining residues, Phe501-His506, within the active site are important for transducing these ligand induced conformational changes. We propose that these conformational changes promote substrate discrimination between H+ and CO2 to favor the metabolically preferred carboxylation product, acetoacetate.


Assuntos
Carboxiliases , Mesna , Acetoacetatos/metabolismo , Dióxido de Carbono/metabolismo , Carboxiliases/metabolismo , Catálise , Ligantes , Mesna/metabolismo , Oxirredutases/metabolismo , Xanthobacter/metabolismo
7.
Environ Sci Pollut Res Int ; 29(36): 54432-54447, 2022 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-35304716

RESUMO

A solid polymer, poly[(sodium methacrylate)-co-2-(methacryloyloxy)ethyl acetoacetate], p(MAA-co-MEAA) was synthesized and then grafted onto carbon nanotubes to prepare poly(MAA-co-MEAA)-grafted carbon nanotubes [CNT-g-p(MAA-co-MEAA)]. NMR, TGA, and FT-IR characterized the synthesized polymers and adsorbents. SEM-EDX was used to investigate the surface characteristics of the adsorbents. Pb2+ was removed from the aqueous solution using the CNT-g-p(MAA-co-MEAA). A batch adsorption experiment was performed at different Pb2+ concentrations (1, 10, 25, 50 mg/L), pH (4 and 6.75), temperature (25 and 35 °C), and contact periods (1, 5, 20, 60, and 1440 min) to study the adsorption kinetics and isotherm. The adsorbent dose of 2.5 g/L could effectively lower the initial Pb2+ concentration of 1000 to 2 ppb. The maximum adsorption capacity of the adsorbent was found to be 1178 mg/g. In addition, the adsorbents have been shown to effectively reduce the coexisting metal ion concentrations from industrial wastewater, which indicated the potential of the proposed adsorbent in removing metal ions from coexisting metals containing wastewater. To predict the adsorption efficiency of Pb2+, various linear, non-linear, and neural network models were established. An additional data set, not incorporated in model training, was used to validate the models. A number of models showed excellent performance with R2 in the range of 0.89-0.98. In model validation studies, the correlation coefficients (r) ranged from 0.94 to 0.99. The novel adsorbent and models will most likely aid in the development of a robust treatment technique for removing Pb2+ ions from water and wastewater.


Assuntos
Nanotubos de Carbono , Poluentes Químicos da Água , Purificação da Água , Acetoacetatos , Adsorção , Concentração de Íons de Hidrogênio , Íons , Cinética , Chumbo , Metacrilatos , Polímeros/química , Espectroscopia de Infravermelho com Transformada de Fourier , Águas Residuárias/química , Água/química , Poluentes Químicos da Água/análise , Purificação da Água/métodos
8.
Pediatr Nephrol ; 37(6): 1347-1353, 2022 06.
Artigo em Inglês | MEDLINE | ID: mdl-34757480

RESUMO

BACKGROUND: The presence of ketone bodies (KBs) can interfere with creatinine (Cr) measurement in both enzymatic and Jaffe methods. Since a high proportion of children hospitalized for diabetic ketoacidosis (DKA) develop acute kidney injury (AKI), here we investigate whether KB interferences affect the accuracy of pediatric Cr measurement. METHODS: Residual patient plasma samples were pooled to make three Cr levels (~ 50, 100, and 250 µM). KBs (acetone, acetoacetate, and ß-hydroxybutyrate) were used to spike the pooled samples. All samples were measured for Cr by two enzymatic methods (E1 and E2), two Jaffe methods (J1 and J2), and LC-MS/MS. LC-MS/MS was considered the gold standard, and the % difference in Cr concentration was calculated for each method. RESULTS: E1 and E2 were unaffected by the presence of all three KBs. J1 and J2 were unaffected by the presence of ß-hydroxybutyrate. The presence of acetone resulted in dose-dependent positive interference in both Jaffe methods, whereas the presence of acetoacetate resulted in dose-dependent positive and negative interference in J1 and J2, respectively. CONCLUSIONS: Compared to the enzymatic methods, the Jaffe methods were much more susceptible to interference by acetone and acetoacetate, especially at lower Cr values which are commonly seen in pediatrics. Interpretation of changes in Cr concentration between different hospitals when transferring patients can become ambiguous and true kidney function unclear if different methods are used without awareness of method-specific biases. To improve DKA patient care, we recommend standardizing all of the Cr methods to an enzymatic method. A higher resolution version of the Graphical abstract is available as Supplementary information.


Assuntos
Cetoacidose Diabética , Ácido 3-Hidroxibutírico , Acetoacetatos , Acetona , Criança , Cromatografia Líquida/métodos , Creatinina , Cetoacidose Diabética/diagnóstico , Humanos , Corpos Cetônicos , Espectrometria de Massas em Tandem
9.
Vet Immunol Immunopathol ; 244: 110370, 2022 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-34952251

RESUMO

Repeat breeding, which is non-pregnancy following three or more breeding attempts, is a serious reproductive disorder in cattle. In the present study, metabolomic profiling was used to identify metabolites in the blood plasma of repeat breeder cows (RBCs) and non-RBCs. Metabolomic analysis showed that acetoacetate (AcAc), a ketone body, was detected in RBCs, but not in non-RBCs. In contrast, ß-hydroxybutyrate (BHB) was at similar levels in both RBCs and non-RBCs. We hypothesized that an imbalance of AcAc and BHB induces abnormal inflammatory conditions, especially the NLRP3 inflammasome, which regulates sterile inflammation to control interleukin (IL)-1ß secretion, and may be associated with repeat breeding in cattle. To investigate this hypothesis, blood samples were collected from both non-RBCs and RBCs on day 7 of the estrous cycle. The mRNA expression of IL1B in peripheral blood mononuclear cells (PBMCs) was observed to be higher in RBCs than in non-RBCs. To test the effects of AcAc and BHB on inflammatory responses, blood samples were collected from healthy cows and PBMCs were isolated. PBMCs were treated with AcAc and BHB to investigate the activation of the NLRP3 inflammasome (complex of NLRP3, ASC, and caspase-1) and IL-1ß secretion. AcAc treatment resulted in higher protein and/or mRNA expression of NLRP3 and IL-1ß in PBMCs. Moreover, AcAc increased the co-localization of NLRP3 and ASC and stimulated caspase-1 activation, indicating the formation of the platform of the NLRP3 inflammasome. Addition of specific NLRP3 inhibitor, MCC950, suppressed AcAc stimulation-induced IL-1ß secretion. Contrary to the effects of AcAc, BHB treatment suppressed the activation of NLRP3 inflammasome and IL-1ß secretion in response to AcAc and typical NLRP3 inflammasome triggers. These findings demonstrate that AcAc can potentially trigger NLRP3 inflammasome activation, resulting in IL-1ß secretion, and that these inflammatory responses are suppressed by BHB in bovine PBMCs. In addition, the imbalance between AcAc and BHB with higher levels of IL-1ß may be associated with repeat breeding in cattle.


Assuntos
Acetoacetatos/farmacologia , Inflamassomos , Leucócitos Mononucleares/efeitos dos fármacos , Proteína 3 que Contém Domínio de Pirina da Família NLR , Ácido 3-Hidroxibutírico , Animais , Caspase 1 , Bovinos , Feminino , Inflamassomos/metabolismo , Interleucina-1beta , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo
10.
Nat Commun ; 12(1): 7115, 2021 12 08.
Artigo em Inglês | MEDLINE | ID: mdl-34880237

RESUMO

Lactic acidosis, the extracellular accumulation of lactate and protons, is a consequence of increased glycolysis triggered by insufficient oxygen supply to tissues. Macrophages are able to differentiate from monocytes under such acidotic conditions, and remain active in order to resolve the underlying injury. Here we show that, in lactic acidosis, human monocytes differentiating into macrophages are characterized by depolarized mitochondria, transient reduction of mitochondrial mass due to mitophagy, and a significant decrease in nutrient absorption. These metabolic changes, resembling pseudostarvation, result from the low extracellular pH rather than from the lactosis component, and render these cells dependent on autophagy for survival. Meanwhile, acetoacetate, a natural metabolite produced by the liver, is utilized by monocytes/macrophages as an alternative fuel to mitigate lactic acidosis-induced pseudostarvation, as evidenced by retained mitochondrial integrity and function, retained nutrient uptake, and survival without the need of autophagy. Our results thus show that acetoacetate may increase tissue tolerance to sustained lactic acidosis.


Assuntos
Acetoacetatos/farmacologia , Acidose Láctica/tratamento farmacológico , Macrófagos/efeitos dos fármacos , Mitocôndrias/metabolismo , Substâncias Protetoras/farmacologia , Reprogramação Celular , Metabolismo Energético , Expressão Gênica , Humanos , Concentração de Íons de Hidrogênio , Ácido Láctico/metabolismo , Macrófagos/metabolismo , Engenharia Metabólica , Mitofagia , Microambiente Tumoral
11.
ACS Sens ; 6(11): 3967-3977, 2021 11 26.
Artigo em Inglês | MEDLINE | ID: mdl-34761912

RESUMO

Cellular redox is intricately linked to energy production and normal cell function. Although the redox states of mitochondria and cytosol are connected by shuttle mechanisms, the redox state of mitochondria may differ from redox in the cytosol in response to stress. However, detecting these differences in functioning tissues is difficult. Here, we employed 13C magnetic resonance spectroscopy (MRS) and co-polarized [1-13C]pyruvate and [1,3-13C2]acetoacetate ([1,3-13C2]AcAc) to monitor production of hyperpolarized (HP) lactate and ß-hydroxybutyrate as indicators of cytosolic and mitochondrial redox, respectively. Isolated rat hearts were examined under normoxic conditions, during low-flow ischemia, and after pretreatment with either aminooxyacetate (AOA) or rotenone. All interventions were associated with an increase in [Pi]/[ATP] measured by 31P NMR. In well-oxygenated untreated hearts, rapid conversion of HP [1-13C]pyruvate to [1-13C]lactate and [1,3-13C2]AcAc to [1,3-13C2]ß-hydroxybutyrate ([1,3-13C2]ß-HB) was readily detected. A significant increase in HP [1,3-13C2]ß-HB but not [1-13C]lactate was observed in rotenone-treated and ischemic hearts, consistent with an increase in mitochondrial NADH but not cytosolic NADH. AOA treatments did not alter the productions of HP [1-13C]lactate or [1,3-13C2]ß-HB. This study demonstrates that biomarkers of mitochondrial and cytosolic redox may be detected simultaneously in functioning tissues using co-polarized [1-13C]pyruvate and [1,3-13C2]AcAc and 13C MRS and that changes in mitochondrial redox may precede changes in cytosolic redox.


Assuntos
Acetoacetatos , Ácido Pirúvico , Acetoacetatos/metabolismo , Animais , Citosol/metabolismo , Ácido Láctico , Mitocôndrias/metabolismo , Oxirredução , Ácido Pirúvico/metabolismo , Ratos
12.
Biochem Biophys Res Commun ; 585: 61-67, 2021 12 31.
Artigo em Inglês | MEDLINE | ID: mdl-34794035

RESUMO

Leucine, isoleucine and valine, known as branched chain amino acids (BCAAs), have been reported to be degraded by different cancer cells, and their biodegradation pathways have been suggested as anticancer targets. However, the mechanisms by which the degradation of BCAAs could support the growth of cancer cells remains unclear. In this work, 13C experiments have been carried out in order to elucidate the metabolic role of BCAA degradation in two breast cancer cell lines (MCF-7 and BCC). The results revealed that up to 36% of the energy production via respiration by MCF-7 cells was supported by the degradation of BCAAs. Also, 67% of the mevalonate (the precursor of cholesterol) synthesized by the cells was coming from the degradation of leucine. The results were lower for BCC cells (14 and 30%, respectively). The non-tumorigenic epythelial cell line MCF-10A was used as a control, showing that 10% of the mitochondrial acetyl-CoA comes from the degradation of BCAAs and no mevalonate production. Metabolic flux analysis around the mevalonate node, also revealed that significant amounts of acetoacetate are being produced from BCAA derived carbon, which could be at the source of lipid synthesis. From these results we can conclude that the degradation of BCAAs is an important energy and carbon source for the proliferation of some cancer cells and its therapeutic targeting could be an interesting option.


Assuntos
Aminoácidos de Cadeia Ramificada/metabolismo , Neoplasias da Mama/metabolismo , Metabolismo Energético , Análise do Fluxo Metabólico/métodos , Ácido Mevalônico/metabolismo , Acetoacetatos/metabolismo , Algoritmos , Neoplasias da Mama/patologia , Carbono/metabolismo , Linhagem Celular , Linhagem Celular Tumoral , Ciclo do Ácido Cítrico , Feminino , Humanos , Leucina/metabolismo , Células MCF-7 , Redes e Vias Metabólicas , Mitocôndrias/metabolismo , Modelos Biológicos
13.
Biotechnol Bioeng ; 118(11): 4278-4289, 2021 11.
Artigo em Inglês | MEDLINE | ID: mdl-34289076

RESUMO

Whole-cell biosensors hold potential in a variety of industrial, medical, and environmental applications. These biosensors can be constructed through the repurposing of bacterial sensing mechanisms, including the common two-component system (TCS). Here we report on the construction of a range of novel biosensors that are sensitive to acetoacetate, a molecule that plays a number of roles in human health and biology. These biosensors are based on the AtoSC TCS. An ordinary differential equation model to describe the action of the AtoSC TCS was developed and sensitivity analysis of this model used to help inform biosensor design. The final collection of biosensors constructed displayed a range of switching behaviours at physiologically relevant acetoacetate concentrations and can operate in several Escherichia coli host strains. It is envisaged that these biosensor strains will offer an alternative to currently available commercial strip tests and, in future, may be adopted for more complex in vivo or industrial monitoring applications.


Assuntos
Acetoacetatos/metabolismo , Técnicas Biossensoriais , Proteínas de Escherichia coli , Escherichia coli , Regulação Bacteriana da Expressão Gênica , Acetoacetatos/análise , Escherichia coli/genética , Escherichia coli/metabolismo , Proteínas de Escherichia coli/genética , Proteínas de Escherichia coli/metabolismo , Humanos , Óperon
14.
Appl Microbiol Biotechnol ; 105(14-15): 5821-5832, 2021 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-34324009

RESUMO

Pathway engineering is a useful technology for producing desired compounds on a large scale by modifying the biosynthetic pathways of host organisms using genetic engineering. We focused on acetoacetate esters as novel low-cost substrates and established an efficient terpenoid production system using pathway-engineered recombinant Escherichia coli. Functional analysis using recombinant E. coli proteins of 18 carboxylesterases identified from the microbial esterases and lipases database showed that the p-nitrobenzyl esterase (PnbA) from Bacillus subtilis specifically hydrolyzed two acetoacetate esters: methyl acetoacetate (MAA) and ethyl acetoacetate (EAA). We generated a plasmid (pAC-Mev/Scidi/Aacl/PnbA) co-expressing PnbA and six enzymes of the mevalonate pathway gene cluster from Streptomyces, isopentenyl diphosphate isomerase type I from Saccharomyces cerevisiae, and acetoacetyl-coenzyme A ligase from Rattus norvegicus. The plasmid pAC-Mev/Scidi/Aacl/PnbA was introduced into E. coli along with plasmid expressing carotenoid (lycopene) or sesquiterpene (ß-bisabolene) biosynthesis genes, and the terpenoid production was evaluated following the addition of acetoacetate esters as substrates. These recombinant E. coli strains used MAA and EAA as substrates for the biosynthesis of terpenoids and produced almost equivalent concentrations of target compounds compared with the previous production system that used mevalonolactone and lithium acetoacetate. The findings of this study will enable the production of useful terpenoids from low-cost substrates, which may facilitate their commercial production on an industrial scale in the future. KEY POINTS: • PnbA from Bacillus subtilis exhibits acetoacetate hydrolysis activity. • A plasmid enabling terpenoid synthesis from acetoacetate esters was constructed. • Acetoacetate esters as substrates enable a low-cost production of terpenoids.


Assuntos
Escherichia coli , Terpenos , Acetoacetatos , Animais , Hidrolases de Éster Carboxílico , Escherichia coli/genética , Ésteres , Hidrólise , Engenharia Metabólica , Ratos
15.
J Appl Physiol (1985) ; 131(2): 435-441, 2021 08 01.
Artigo em Inglês | MEDLINE | ID: mdl-34166120

RESUMO

Inert gas bubbles are widely accepted as the causative factor of decompression sickness (DCS), resulting in gas embolism and systemic inflammatory responses. The anticonvulsive ketone ester 1,3-butanediol acetoacetate diester (BD-AcAc2) was reported to have the characteristics of increasing blood oxygen partial pressure (ppO2) and anti-inflammation and was thought to have the potential to reduce bubble formation and alleviate the pathological process of DCS. This study aims to investigate the potential protection of BD-AcAc2 against DCS in a rat model. A single dose of BD-AcAc2 was administered orally to adult male rats (5 g/kg body wt), followed by pharmacokinetic analysis or simulated air dives. After decompression, signs of DCS were monitored, and blood was sampled for biochemical measurements. Blood ketosis peaked at 2 h and lasted for more than 4 h. The incidence of DCS was decreased and postponed significantly in rats treated with BD-AcAc2 compared with those treated with saline (P < 0.05). Although BD-AcAc2 failed to reduce bubble load (P > 0.05), it showed an obvious decreasing trend. BD-AcAc2 significantly increased blood ppO2 and ameliorated oxidative and inflammatory responses, represented by increased plasma malondialdehyde (MDA), IL-1, IL-6, and TNF-α and decreased glutathione thiol (P < 0.05) levels, whereas blood pH remained unchanged (P > 0.05). These results suggest that BD-AcAc2 exerted beneficial effects on DCS rats mainly related to increasing ppO2 and anti-inflammatory and antioxidant properties. Together with its capacity for delaying central nervous system (CNS) oxygen toxicity seizures, BD-AcAc2 might be an ideal drug candidate for DCS prevention and treatment.NEW & NOTEWORTHY This is the first study exploring the effects of BD-AcAc2 on DCS prevention, and it was proven to be an efficient and simple method. The role of BD-AcAc2 in increasing ppO2, anti-inflammatory and antioxidant properties was thought to be the critical mechanism in DCS prevention.


Assuntos
Doença da Descompressão , Mergulho , Acetoacetatos , Animais , Butileno Glicóis , Descompressão , Doença da Descompressão/tratamento farmacológico , Masculino , Ratos , Convulsões
16.
Acta Biochim Biophys Sin (Shanghai) ; 53(8): 1009-1016, 2021 Jul 28.
Artigo em Inglês | MEDLINE | ID: mdl-34184741

RESUMO

Acetoacetate (AA) is an important ketone body that is used as an oxidative fuel to supply energy for the cellular activities of various tissues, including the brain and skeletal muscle. We recently revealed a new signaling role for AA by showing that it promotes muscle cell proliferation in vitro, enhances muscle regeneration in vivo, and ameliorates the dystrophic muscle phenotype of Mdx mice. In this study, we provide new molecular insight into this function of AA. We show that AA promotes C2C12 cell proliferation by transcriptionally upregulating the expression of muscle-specific miR-133b, which in turn stimulates muscle cell proliferation by targeting serum response factor. Furthermore, we show that the AA-induced upregulation of miR-133b is transcriptionally mediated by MEF2 via the Mek-Erk1/2 signaling pathway. Mechanistically, our findings provide further convincing evidence that AA acts as signaling metabolite to actively regulate various cellular activities in mammalian cells.


Assuntos
Acetoacetatos/farmacologia , Proliferação de Células/efeitos dos fármacos , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , MicroRNAs/metabolismo , Mioblastos/metabolismo , Fator de Resposta Sérica/metabolismo , Animais , Linhagem Celular , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , MAP Quinase Quinase Quinases/metabolismo , Fatores de Transcrição MEF2/metabolismo , Camundongos
17.
Int J Mol Sci ; 22(9)2021 Apr 30.
Artigo em Inglês | MEDLINE | ID: mdl-33946157

RESUMO

The metabolic ratios lactate/pyruvate and ß-hydroxybutyrate/acetoacetate are considered valuable tools to evaluate the in vivo redox cellular state by estimating the free NAD+/NADH in cytoplasm and mitochondria, respectively. The aim of the current study was to validate a gas-chromatography mass spectrometry method for simultaneous determination of the four metabolites in plasma and liver tissue. The procedure included an o-phenylenediamine microwave-assisted derivatization, followed by liquid-liquid extraction with ethyl acetate and silylation with bis(trimethylsilyl)trifluoroacetamide:trimethylchlorosilane 99:1. The calibration curves presented acceptable linearity, with a limit of quantification of 0.001 mM for pyruvate, ß-hydroxybutyrate and acetoacetate and of 0.01 mM for lactate. The intra-day and inter-day accuracy and precision were within the European Medicines Agency's Guideline specifications. No significant differences were observed in the slope coefficient of three-point standard metabolite-spiked curves in plasma or liver and water, and acceptable recoveries were obtained in the metabolite-spiked samples. Applicability of the method was tested in precision-cut liver rat slices and also in HepG2 cells incubated under different experimental conditions challenging the redox state. In conclusion, the validated method presented good sensitivity, specificity and reproducibility in the quantification of lactate/pyruvate and ß-hydroxybutyrate/acetate metabolites and may be useful in the evaluation of in vivo redox states.


Assuntos
Ácido 3-Hidroxibutírico/metabolismo , Acetoacetatos/metabolismo , Cromatografia Gasosa-Espectrometria de Massas , Lactatos/metabolismo , Piruvatos/metabolismo , Ácido 3-Hidroxibutírico/análise , Ácido 3-Hidroxibutírico/sangue , Acetoacetatos/análise , Acetoacetatos/sangue , Animais , Feminino , Cromatografia Gasosa-Espectrometria de Massas/métodos , Células Hep G2 , Humanos , Lactatos/análise , Lactatos/sangue , Limite de Detecção , Fígado/química , Fígado/metabolismo , Oxirredução , Piruvatos/análise , Piruvatos/sangue , Ratos Wistar
18.
Int J Mol Sci ; 22(6)2021 Mar 13.
Artigo em Inglês | MEDLINE | ID: mdl-33805788

RESUMO

Persistent chronic liver diseases increase the scar formation and extracellular matrix accumulation that further progress to liver fibrosis and cirrhosis. Nevertheless, there is no antifibrotic therapy to date. The ketogenic diet is composed of high fat, moderate to low-protein, and very low carbohydrate content. It is mainly used in epilepsy and Alzheimer's disease. However, the effects of the ketogenic diet on liver fibrosis remains unknown. Through ketogenic diet consumption, ß-hydroxybutyrate (bHB) and acetoacetate (AcAc) are two ketone bodies that are mainly produced in the liver. It is reported that bHB and AcAc treatment decreases cancer cell proliferation and promotes apoptosis. However, the influence of bHB and AcAc in hepatic stellate cell (HSC) activation and liver fibrosis are still unclear. Therefore, this study aimed to investigate the effect of the ketogenic diet and ketone bodies in affecting liver fibrosis progression. Our study revealed that feeding a high-fat ketogenic diet increased cholesterol accumulation in the liver, which further enhanced the carbon tetrachloride (CCl4)- and thioacetamide (TAA)-induced liver fibrosis. In addition, more severe liver inflammation and the loss of hepatic antioxidant and detoxification ability were also found in ketogenic diet-fed fibrotic mouse groups. However, the treatment with ketone bodies (bHB and AcAc) did not suppress transforming growth factor-ß (TGF-ß)-induced HSC activation, platelet-derived growth factor (PDGF)-BB-triggered proliferation, and the severity of CCl4-induced liver fibrosis in mice. In conclusion, our study demonstrated that feeding a high-fat ketogenic diet may trigger severe steatohepatitis and thereby promote liver fibrosis progression. Since a different ketogenic diet composition may exert different metabolic effects, more evidence is necessary to clarify the effects of a ketogenic diet on disease treatment.


Assuntos
Ácido 3-Hidroxibutírico/farmacologia , Acetoacetatos/farmacologia , Colesterol/biossíntese , Dieta Cetogênica/efeitos adversos , Cirrose Hepática/metabolismo , Fígado/efeitos dos fármacos , Ácido 3-Hidroxibutírico/biossíntese , Acetoacetatos/metabolismo , Actinas/genética , Actinas/metabolismo , Animais , Becaplermina/farmacologia , Tetracloreto de Carbono/administração & dosagem , Catalase/genética , Catalase/metabolismo , Proliferação de Células/efeitos dos fármacos , Colesterol/sangue , Colágeno Tipo I/genética , Colágeno Tipo I/metabolismo , Citocromo P-450 CYP1A2/genética , Citocromo P-450 CYP1A2/metabolismo , Desmina/genética , Desmina/metabolismo , Progressão da Doença , Regulação da Expressão Gênica/efeitos dos fármacos , Células Estreladas do Fígado/citologia , Células Estreladas do Fígado/efeitos dos fármacos , Células Estreladas do Fígado/metabolismo , Fígado/metabolismo , Fígado/patologia , Cirrose Hepática/induzido quimicamente , Cirrose Hepática/genética , Cirrose Hepática/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Cultura Primária de Células , Índice de Gravidade de Doença , Superóxido Dismutase/genética , Superóxido Dismutase/metabolismo , Superóxido Dismutase-1/genética , Superóxido Dismutase-1/metabolismo , Tioacetamida/administração & dosagem , Fator de Crescimento Transformador beta/biossíntese , Fator de Crescimento Transformador beta/farmacologia
19.
Zhonghua Wei Zhong Bing Ji Jiu Yi Xue ; 33(2): 180-185, 2021 Feb.
Artigo em Chinês | MEDLINE | ID: mdl-33729137

RESUMO

OBJECTIVE: To investigate the diagnostic value of mitochondrial associated protein fumarylacetoacetate domain containing protein 1 (FAHD1) and growth differentiation factor-15 (GDF-15) in sepsis. METHODS: Based on the database of the whole process of sepsis early warning, diagnosis and treatment management, which was established on the study of sepsis early warning and standardized diagnosis and treatment system, adult patients with suspected infection admitted to the department of critical care medicine of Zhejiang Hospital, Second Affiliated Hospital of Zhejiang University, the First Affiliated Hospital of Sun Yat-Sen University, West China Hospital of Sichuan University, Ningbo First Hospital from May 2014 to October 2015 were enrolled. The basic vital signs, and the main blood routine results, liver and kidney function, blood gas, acute physiology and chronic health evaluation II (APACHE II) and sequential organ failure assessment (SOFA) score at the time of diagnosis of patients with or without sepsis were analyzed. The preserved serum samples were taken, the levels of procalcitonin (PCT), C-reactive protein (CRP) were detected by electrochemiluminescence method, immunoturbidimetry respectively, and FAHD1 and GDF-15 were detected by enzyme linked immunosorbent assay (ELISA). Univariate and multivariate Logistic regression were used to analyze the risk factors for sepsis diagnose. The indexes' diagnostic efficacy in sepsis were analyzed by receiver operating characteristics curve (ROC curve). RESULTS: A total of 132 patients were enrolled, including 76 cases of sepsis and 56 cases of non-sepsis. Compared with the non-sepsis group, the heart rate in the sepsis group was increased (bpm: 116.4±17.8 vs. 97.4±19.1), while the mean arterial pressure (MAP), platelet count (PLT), arterial partial pressure of oxygen (PaO2) were significantly decreased [MAP (mmHg, 1 mmHg = 0.133 kPa): 65.8±9.7 vs. 74.7±10.3, PLT (×109/L): 120 (69, 204) vs. 163 (117, 239), PaO2 (mmHg): 83.0 (66.6, 108.0) vs. 108.0 (84.4, 130.0), all P < 0.05], direct bilirubin (DBil), serum creatinine (SCr), lactic acid (Lac), APACHE II score and SOFA score were significantly increased [DBil (µmol/L): 13.00 (5.55, 55.31) vs. 6.20 (2.20, 21.90), SCr (µmol/L): 118.00 (70.00, 191.73) vs. 77.20 (59.65, 110.86), Lac (mmol/L): 2.90 (1.50, 4.10) vs. 1.90 (1.20, 2.80), APACHE II score: 20.0 (16.0, 25.0) vs. 16.0 (10.0, 21.0), SOFA score: 12.0 (8.0, 16.0) vs. 8.0 (5.0, 13.0), all P < 0.05]. In addition, the serum levels of FAHD1, GDF-15, PCT and CRP in sepsis group were significantly higher than those in non-sepsis group [FAHD1 (µg/L): 3.96 (2.25, 5.92) vs. 2.47 (1.03, 3.54), GDF-15 (µg/L): 8.46 (4.37, 19.68) vs. 4.32 (1.74, 10.39), PCT (µg/L): 3.79 (1.37, 11.32) vs. 0.42 (0.12, 2.14), CRP (mg/L): 154.43 (61.33, 283.20) vs. 65.95 (28.15, 144.69), all P < 0.01]. Multivariate Logistic regression showed that serum FAHD1 [odds ratio (OR) = 1.135, 95% confidence interval (95%CI) was 1.045-1.234], GDF-15 (OR = 1.090, 95%CI was 1.029-1.155) and CRP (OR = 1.007, 95%CI was 1.002-1.011) were risk factors for sepsis (all P < 0.05). ROC curve analysis of sepsis showed that the areas under ROC curve (AUC) of serum mitochondrial associated proteins FAHD1 and GDF-15 were 0.727 (95%CI was 0.641-0.802) and 0.677 (95%CI was 0.588-0.757), respectively; and the AUC of classical infection indexes PCT and CRP were 0.767 (95%CI was 0.683-0.837) and 0.680 (95%CI was 0.59-0.760), respectively. There was no significant difference between the AUC of mitochondrial associated proteins and classical infection indexes. The combination of FAHD1, GDF-15, PCT and CRP had the largest AUC, which was 0.809 (95%CI was 0.730-0.874), and the sensitivity was 75.00%, and the specificity was 80.00%. CONCLUSIONS: Mitochondrial associated protein FAHD1 and GDF-15 are associated with sepsis, and the diagnostic efficiency is improved when combined with PCT and CRP, which might provide experimental basis for screening diagnostic markers of sepsis.


Assuntos
Fator 15 de Diferenciação de Crescimento , Sepse , Acetoacetatos , Adulto , China , Humanos , Hidrolases , Proteínas Mitocondriais , Prognóstico , Curva ROC , Estudos Retrospectivos , Sepse/diagnóstico
20.
ACS Appl Mater Interfaces ; 13(11): 12928-12940, 2021 Mar 24.
Artigo em Inglês | MEDLINE | ID: mdl-33709691

RESUMO

The fabrication of covalently cross-linked high-surface-area biopolymeric nanogel fibers by nanopore extrusion is reported for the first time. The biopolymer pullulan was functionalized with tert-butyl acetoacetate via a transesterification reaction to synthesize the water-soluble ketone-rich precursor pullulan acetoacetate (PUAA). PUAA and carbonic dihydrazide (CDH) as cross-linker were extruded through anodic aluminum oxide (AAO) nanoporous membranes, which possessed an average pore diameter of 61 ± 2 nm. By changing the concentration of PUAA, the flow rate, and extrusion time, the step polymerization cross-linking reaction was controlled so that the polymer can be extruded gradually during cross-linking through the membrane, avoiding the formation of macroscopic bulk hydrogels and rupture of the AAO membrane. Fibers with diameters on the order of 250 nm were obtained. This approach was also expanded to functionalized PUAA derivatives together with the fluorogenic substrate 4-methylumbelliferyl-ß-d-glucuronide MUGlcU in (PUAA-MUGlcU), which exhibited a mean equilibrium swelling ratio of 5.7 and 9.0 in Milli-Q water and in phosphate-buffered saline, respectively. ß-Glucuronidase was sensitively detected via fluorescence of 4-methylumbelliferone, which was liberated in the enzymatic hydrolysis reaction of PUAA-MUGlcU. Compared to hydrogel slabs, the rate of the hydrolysis was >20% higher in the nanogel fibers, facilitating the rapid detection of ß-glucuronidase-producing Escherichia coli (E. coli Mach1-T1). Nanopore extruded nanogel fibers are therefore considered a viable approach to enhance the functionality of hydrogels in surface-dominated processes.


Assuntos
Escherichia coli/enzimologia , Corantes Fluorescentes/química , Glucanos/química , Glucuronidase/análise , Nanogéis/química , Acetoacetatos/química , Ensaios Enzimáticos/métodos
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