Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 12 de 12
Filtrar
1.
Proc Natl Acad Sci U S A ; 121(37): e2408262121, 2024 Sep 10.
Artigo em Inglês | MEDLINE | ID: mdl-39226352

RESUMO

Truncating genetic variants of SORL1, encoding the endosome recycling receptor SORLA, have been accepted as causal of Alzheimer's disease (AD). However, most genetic variants observed in SORL1 are missense variants, for which it is complicated to determine the pathogenicity level because carriers come from pedigrees too small to be informative for penetrance estimations. Here, we describe three unrelated families in which the SORL1 coding missense variant rs772677709, that leads to a p.Y1816C substitution, segregates with Alzheimer's disease. Further, we investigate the effect of SORLA p.Y1816C on receptor maturation, cellular localization, and trafficking in cell-based assays. Under physiological circumstances, SORLA dimerizes within the endosome, allowing retromer-dependent trafficking from the endosome to the cell surface, where the luminal part is shed into the extracellular space (sSORLA). Our results showed that the p.Y1816C mutant impairs SORLA homodimerization in the endosome, leading to decreased trafficking to the cell surface and less sSORLA shedding. These trafficking defects of the mutant receptor can be rescued by the expression of the SORLA 3Fn-minireceptor. Finally, we find that iPSC-derived neurons with the engineered p.Y1816C mutation have enlarged endosomes, a defining cytopathology of AD. Our studies provide genetic as well as functional evidence that the SORL1 p.Y1816C variant is causal for AD. The partial penetrance of the mutation suggests this mutation should be considered in clinical genetic screening of multiplex early-onset AD families.


Assuntos
Doença de Alzheimer , Endossomos , Proteínas Relacionadas a Receptor de LDL , Proteínas de Membrana Transportadoras , Linhagem , Humanos , Doença de Alzheimer/genética , Doença de Alzheimer/metabolismo , Doença de Alzheimer/patologia , Endossomos/metabolismo , Proteínas Relacionadas a Receptor de LDL/genética , Proteínas Relacionadas a Receptor de LDL/metabolismo , Feminino , Masculino , Proteínas de Membrana Transportadoras/genética , Proteínas de Membrana Transportadoras/metabolismo , Mutação de Sentido Incorreto , Transporte Proteico , Multimerização Proteica , Idoso , Pessoa de Meia-Idade , Células HEK293
2.
Bioinformatics ; 36(9): 2943-2945, 2020 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-31930381

RESUMO

SUMMARY: Nuclear magnetic resonance (NMR)-based metabolomics is widely used to obtain metabolic fingerprints of biological systems. While targeted workflows require previous knowledge of metabolites, prior to statistical analysis, untargeted approaches remain a challenge. Computational tools dealing with fully untargeted NMR-based metabolomics are still scarce or not user-friendly. Therefore, we developed AlpsNMR (Automated spectraL Processing System for NMR), an R package that provides automated and efficient signal processing for untargeted NMR metabolomics. AlpsNMR includes spectra loading, metadata handling, automated outlier detection, spectra alignment and peak-picking, integration and normalization. The resulting output can be used for further statistical analysis. AlpsNMR proved effective in detecting metabolite changes in a test case. The tool allows less experienced users to easily implement this workflow from spectra to a ready-to-use dataset in their routines. AVAILABILITY AND IMPLEMENTATION: The AlpsNMR R package and tutorial is freely available to download from http://github.com/sipss/AlpsNMR under the MIT license. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.


Assuntos
Metabolômica , Software , Imageamento por Ressonância Magnética , Espectroscopia de Ressonância Magnética , Fluxo de Trabalho
3.
Int J Mol Sci ; 22(19)2021 Sep 30.
Artigo em Inglês | MEDLINE | ID: mdl-34638936

RESUMO

Nicotinamide adenine dinucleotide (NAD+) and its reduced form (NADH) are coenzymes employed in hundreds of metabolic reactions. NAD+ also serves as a substrate for enzymes such as sirtuins, poly(ADP-ribose) polymerases (PARPs) and ADP-ribosyl cyclases. Given the pivotal role of NAD(H) in health and disease, studying NAD+ metabolism has become essential to monitor genetic- and/or drug-induced perturbations related to metabolic status and diseases (such as ageing, cancer or obesity), and its possible therapies. Here, we present a strategy based on liquid chromatography-tandem mass spectrometry (LC-MS/MS), for the analysis of the NAD+ metabolome in biological samples. In this method, hydrophilic interaction chromatography (HILIC) was used to separate a total of 18 metabolites belonging to pathways leading to NAD+ biosynthesis, including precursors, intermediates and catabolites. As redox cofactors are known for their instability, a sample preparation procedure was developed to handle a variety of biological matrices: cell models, rodent tissues and biofluids, as well as human biofluids (urine, plasma, serum, whole blood). For clinical applications, quantitative LC-MS/MS for a subset of metabolites was demonstrated for the analysis of the human whole blood of nine volunteers. Using this developed workflow, our methodology allows studying NAD+ biology from mechanistic to clinical applications.


Assuntos
Metaboloma , NAD/biossíntese , Plasma/metabolismo , Soro/metabolismo , Espectrometria de Massas em Tandem/métodos , Urina/fisiologia , Animais , Doadores de Sangue , Cromatografia Líquida/métodos , Células Hep G2 , Humanos , Interações Hidrofóbicas e Hidrofílicas , Metabolômica/métodos , Camundongos , Camundongos Endogâmicos C57BL , Monitorização Fisiológica/métodos , Oxirredução , Projetos Piloto , Plasma/química , Soro/química , Urina/química
4.
J Biol Inorg Chem ; 21(3): 319-28, 2016 06.
Artigo em Inglês | MEDLINE | ID: mdl-26880094

RESUMO

Self-splicing group II introns are highly structured RNA molecules, containing a characteristic secondary and catalytically active tertiary structure, which is formed only in the presence of Mg(II). Mg(II) initiates the first folding step governed by the κζ element within domain 1 (D1κζ). We recently solved the NMR structure of D1κζ derived from the mitochondrial group II intron ribozyme Sc.ai5γ and demonstrated that Mg(II) is essential for its stabilization. Here, we performed a detailed multinuclear NMR study of metal ion interactions with D1κζ, using Cd(II) and cobalt(III)hexammine to probe inner- and outer-sphere coordination of Mg(II) and thus to better characterize its binding sites. Accordingly, we mapped (1)H, (15)N, (13)C, and (31)P spectral changes upon addition of different amounts of the metal ions. Our NMR data reveal a Cd(II)-assisted macrochelate formation at the 5'-end triphosphate, a preferential Cd(II) binding to guanines in a helical context, an electrostatic interaction in the ζ tetraloop receptor and various metal ion interactions in the GAAA tetraloop and κ element. These results together with our recently published data on Mg(II) interaction provide a much better understanding of Mg(II) binding to D1κζ, and reveal how intricate and complex metal ion interactions can be.


Assuntos
Cádmio/química , Cobalto/química , Magnésio/química , RNA/química , Sítios de Ligação , Espectroscopia de Ressonância Magnética
5.
Anal Bioanal Chem ; 408(2): 567-78, 2016 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-26577083

RESUMO

Obesity with related complications represents a widespread health problem. The etiopathogenesis of obesity is often studied using numerous rodent models. The mouse model of monosodium glutamate (MSG)-induced obesity was exploited as a model of obesity combined with insulin resistance. The aim of this work was to characterize the metabolic status of MSG mice by NMR-based metabolomics in combination with relevant biochemical and hormonal parameters. NMR analysis of urine at 2, 6, and 9 months revealed altered metabolism of nicotinamide and polyamines, attenuated excretion of major urinary proteins, increased levels of phenylacetylglycine and allantoin, and decreased concentrations of methylamine in urine of MSG-treated mice. Altered levels of creatine, citrate, succinate, and acetate were observed at 2 months of age and approached the values of control mice with aging. The development of obesity and insulin resistance in 6-month-old MSG mice was also accompanied by decreased mRNA expressions of adiponectin, lipogenetic and lipolytic enzymes and peroxisome proliferator-activated receptor-gamma in fat while mRNA expressions of lipogenetic enzymes in the liver were enhanced. At the age of 9 months, biochemical parameters of MSG mice were normalized to the values of the controls. This fact pointed to a limited predictive value of biochemical data up to age of 6 months as NMR metabolomics confirmed altered urine metabolic composition even at 9 months.


Assuntos
Metabolômica , Obesidade/urina , Glutamato de Sódio/efeitos adversos , Urina/química , Animais , Glicemia/metabolismo , Humanos , Insulina/metabolismo , Metabolismo dos Lipídeos , Fígado/metabolismo , Masculino , Camundongos , Obesidade/etiologia , Obesidade/genética , Obesidade/metabolismo , Receptores Ativados por Proliferador de Peroxissomo/genética , Receptores Ativados por Proliferador de Peroxissomo/metabolismo
6.
Chem Res Toxicol ; 28(2): 216-24, 2015 Feb 16.
Artigo em Inglês | MEDLINE | ID: mdl-25621379

RESUMO

There is mounting evidence that cyanobacterial lipopeptides can kill mammalian cells, presenting a hazard to human health. Unfortunately, their mechanism of toxicity is poorly understood. We have isolated new cyclic undecalipopeptides muscotoxin A and B containing unique lipophilicresidue 3-amino-2,5-dihydroxydecanoic acid (5-OH Ahdoa). Muscotoxin B was not used for biological studies due to its poor yield. Muscotoxin A was cytotoxic to YAC-1, Sp/2, and HeLa cancer cell lines (LC(50) ranged from 9.9 to 13.2 µM after 24 h of exposure), causing membrane damage and influx of calcium ions. Subsequently, we studied this lytic mechanism using synthetic liposomes with encapsulated fluorescent probes. Muscotoxin A permeabilized liposomes composed exclusively of phospholipids, demonstrating that no proteins or carbohydrates present in biomembranes are essential for its activity. Paradoxically, the permeabilization activity of muscotoxin A was mediated by a significant reduction in membrane surface fluidity (stiffening), the opposite of that caused by synthetic detergents and cytolytic lipopeptide puwainaphycin F. At 25 °C, muscotoxin A disrupted liposomes with and without cholesterol/sphingomyelin; however, at 37 °C, it was selective against liposomes with cholesterol/sphingomyelin. It appears that both membrane fluidity and organization can affect the lytic activity of muscotoxin A. Our findings strengthen the evidence that cyanobacterial lipopeptides specifically disrupt mammalian cell membranes and bring new insights into the mechanism of this effect.


Assuntos
Permeabilidade da Membrana Celular/efeitos dos fármacos , Membrana Celular/efeitos dos fármacos , Cianobactérias/química , Lipopeptídeos/toxicidade , Fluidez de Membrana/efeitos dos fármacos , Peptídeos Cíclicos/toxicidade , Fosfolipídeos/química , Animais , Morte Celular/efeitos dos fármacos , Membrana Celular/química , Imunofluorescência , Células HeLa , Humanos , Lipopeptídeos/química , Lipopeptídeos/isolamento & purificação , Camundongos , Conformação Molecular , Peptídeos Cíclicos/química , Peptídeos Cíclicos/isolamento & purificação , Células Tumorais Cultivadas
7.
Chembiochem ; 14(17): 2329-37, 2013 Nov 25.
Artigo em Inglês | MEDLINE | ID: mdl-24123716

RESUMO

Aeruginosin-865 (Aer-865), isolated from terrestrial cyanobacterium Nostoc sp. Lukesová 30/93, is the first aeruginosin-type peptide containing both a fatty acid and a carbohydrate moiety, and is the first aeruginosin to be found in the genus Nostoc. Mass spectrometry, chemical and spectroscopic analysis as well as one- and two-dimensional NMR and chiral HPLC analysis of Marfey derivatives were applied to determine the peptidic sequence: D-Hpla, D-Leu, 5-OH-Choi, Agma, with hexanoic and mannopyranosyl uronic acid moieties linked to Choi. We used an AlphaLISA assay to measure the levels of proinflammatory mediators IL-8 and ICAM-1 in hTNF-α-stimulated HLMVECs. Aer-865 showed significant reduction of both: with EC50 values of (3.5±1.5) µg mL(-1) ((4.0±1.7) µM) and (50.0±13.4) µg mL(-1) ((57.8±15.5) µM), respectively. Confocal laser scanning microscopy revealed that the anti-inflammatory effect of Aer-865 was directly associated with inhibition of NF-κB translocation to the nucleus. Moreover, Aer-865 did not show any cytotoxic effect.


Assuntos
Anti-Inflamatórios não Esteroides/química , Anti-Inflamatórios não Esteroides/farmacologia , Glicopeptídeos/farmacologia , Lipopeptídeos/farmacologia , Nostoc/química , Transporte Ativo do Núcleo Celular/efeitos dos fármacos , Anti-Inflamatórios não Esteroides/isolamento & purificação , Linhagem Celular , Relação Dose-Resposta a Droga , Regulação para Baixo/efeitos dos fármacos , Glicopeptídeos/química , Glicopeptídeos/isolamento & purificação , Humanos , Molécula 1 de Adesão Intercelular/biossíntese , Interleucina-8/biossíntese , Lipopeptídeos/química , Lipopeptídeos/isolamento & purificação , Estrutura Molecular , NF-kappa B/metabolismo , Nostoc/crescimento & desenvolvimento , Transporte Proteico/efeitos dos fármacos , Relação Estrutura-Atividade
8.
Nutrients ; 15(6)2023 Mar 07.
Artigo em Inglês | MEDLINE | ID: mdl-36986043

RESUMO

Faecalibacterium prausnitzii (F. prausnitzii) is a bacterial taxon in the human gut with anti-inflammatory properties, and this may contribute to the beneficial effects of healthy eating habits. However, little is known about the nutrients that enhance the growth of F. prausnitzii other than simple sugars and fibers. Here, we combined dietary and microbiome data from the American Gut Project (AGP) to identify nutrients that may be linked to the relative abundance of F. prausnitzii. Using a machine learning approach in combination with univariate analyses, we identified that sugar alcohols, carbocyclic sugar, and vitamins may contribute to F. prausnitzii growth. We next explored the effects of these nutrients on the growth of two F. prausnitzii strains in vitro and observed robust and strain-dependent growth patterns on sorbitol and inositol, respectively. In the context of a complex community using in vitro fermentation, neither inositol alone nor in combinations with vitamin B exerted a significant growth-promoting effect on F. prausnitzii, partly due to high variability among the fecal microbiota community from four healthy donors. However, the fecal communities that showed an increase in F. prausnitzii on inulin also responded with at least 60% more F. prausnitzii on any of inositol containing media than control. Future nutritional studies aiming to increase the relative abundance of F. prausnitzii should explore a personalized approach accounting for strain-level genetic variations and community-level microbiome composition.


Assuntos
Microbiota , Complexo Vitamínico B , Humanos , Faecalibacterium prausnitzii , Inositol , Inulina
9.
Cell Rep ; 42(11): 113310, 2023 11 28.
Artigo em Inglês | MEDLINE | ID: mdl-37864790

RESUMO

During the past two decades, induced pluripotent stem cells (iPSCs) have been widely used to study human neural development and disease. Especially in the field of Alzheimer's disease (AD), remarkable effort has been put into investigating molecular mechanisms behind this disease. Then, with the advent of 3D neuronal cultures and cerebral organoids (COs), several studies have demonstrated that this model can adequately mimic familial and sporadic AD. Therefore, we created an AD-CO model using iPSCs derived from patients with familial AD forms and explored early events and the progression of AD pathogenesis. Our study demonstrated that COs derived from three AD-iPSC lines with PSEN1(A246E) or PSEN2(N141I) mutations developed the AD-specific markers in vitro, yet they also uncover tissue patterning defects and altered development. These findings are complemented by single-cell sequencing data confirming this observation and uncovering that neurons in AD-COs likely differentiate prematurely.


Assuntos
Doença de Alzheimer , Presenilina-1 , Presenilina-2 , Humanos , Doença de Alzheimer/genética , Doença de Alzheimer/patologia , Células-Tronco Pluripotentes Induzidas/patologia , Mutação/genética , Neurônios , Organoides/patologia , Presenilina-1/genética , Presenilina-2/genética
10.
Front Nutr ; 9: 998044, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-36386937

RESUMO

Introduction and aims: Dietary polyphenols have long been associated with health benefits, including the prevention of obesity and related chronic diseases. Overfeeding was shown to rapidly induce weight gain and fat mass, associated with mild insulin resistance in humans, and thus represents a suitable model of the metabolic complications resulting from obesity. We studied the effects of a polyphenol-rich grape extract supplementation on the plasma metabolome during an overfeeding intervention in adults, in two randomized parallel controlled clinical trials. Methods: Blood plasma samples from 40 normal weight to overweight male adults, submitted to a 31-day overfeeding (additional 50% of energy requirement by a high calorie-high fructose diet), given either 2 g/day grape polyphenol extract or a placebo at 0, 15, 21, and 31 days were analyzed (Lyon study). Samples from a similarly designed trial on females (20 subjects) were collected in parallel (Lausanne study). Nuclear magnetic resonance (NMR)-based metabolomics was conducted to characterize metabolome changes induced by overfeeding and associated effects from polyphenol supplementation. The clinical trials are registered under the numbers NCT02145780 and NCT02225457 at ClinicalTrials.gov. Results: Changes in plasma levels of many metabolic markers, including branched chain amino acids (BCAA), ketone bodies and glucose in both placebo as well as upon polyphenol intervention were identified in the Lyon study. Polyphenol supplementation counterbalanced levels of BCAA found to be induced by overfeeding. These results were further corroborated in the Lausanne female study. Conclusion: Administration of grape polyphenol-rich extract over 1 month period was associated with a protective metabolic effect against overfeeding in adults.

11.
J Cachexia Sarcopenia Muscle ; 11(5): 1187-1199, 2020 10.
Artigo em Inglês | MEDLINE | ID: mdl-32274897

RESUMO

BACKGROUND: Constitutional thinness (CT), a non-malnourished underweight state with no eating disorders, is characterized by weight gain resistance to high fat diet. Data issued from muscle biopsies suggested blunted anabolic mechanisms in free-living state. Weight and metabolic responses to protein caloric supplementation has not been yet explored in CT. METHODS: A 2 week overfeeding (additional 600 kcal, 30 g protein, 72 g carbohydrate, and 21 g fat) was performed to compare two groups of CTs (12 women and 11 men) to normal-weight controls (12 women and 10 men). Bodyweight, food intake, energy expenditure, body composition, nitrogen balance, appetite hormones profiles, and urine metabolome were monitored before and after overfeeding. RESULTS: Before overfeeding, positive energy gap was found in both CT genders (309 ± 370 kcal in CT-F and 332 ± 709 kcal in CT-M) associated with higher relative protein intake per kilo (1.74 ± 0.32 g/kg/day in CT-F vs. 1.16 ± 0.23 in C-F, P < 0.0001; 1.56 ± 0.36 in CT-M vs. 1.22 ± 0.32 in C-M, P = 0.03), lower nitrogen (7.26 ± 2.36 g/day in CT-F vs. 11.41 ± 3.64 in C-F, P = 0.003; 9.70 ± 3.85 in CT-M vs. 14.14 ± 4.19 in C-M, P = 0.02), but higher essential amino acids urinary excretion (CT/C fold change of 1.13 for leucine and 1.14 for arginine) in free-living conditions. After overfeeding, CTs presented an accentuated positive energy gap, still higher than in controls (675 ± 540 in CTs vs. 379 ± 427 in C, P = 0.04). Increase in lean mass was induced in both controls genders but not in CTs (a trend was noticed in CT women), despite a similar nitrogen balance after overfeeding (5.06 ± 4.33 g/day in CTs vs. 4.28 ± 3.15 in controls, P = 0.49). Higher anorectic gut hormones' tone, glucagon-like peptide 1 and peptide tyrosine tyrosine, during test meal and higher snacking frequency were noticed before and after overfeeding in CTs. CONCLUSIONS: The blunted muscle energy mechanism, previously described in CTs in free-living state, is associated with basal saturated protein turn over suggested by the concordance of positive nitrogen balance and an increased urine excretion of several essential amino acids. This saturation cannot be overpassed by increasing this spontaneous high-protein intake suggesting a resistance to lean mass gain in CT phenotype.


Assuntos
Condições Sociais , Magreza , Adolescente , Composição Corporal , Metabolismo Energético , Feminino , Humanos , Masculino , Aumento de Peso , Adulto Jovem
12.
Mol Metab ; 30: 192-202, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31767171

RESUMO

OBJECTIVE: A decay in intracellular NAD+ levels is one of the hallmarks of physiological decline in normal tissue functions. Accordingly, dietary supplementation with NAD+ precursors can prevent, alleviate, or even reverse multiple metabolic complications and age-related disorders in diverse model organisms. Within the constellation of NAD+ precursors, nicotinamide riboside (NR) has gained attention due to its potent NAD+ biosynthetic effects in vivo while lacking adverse clinical effects. Nevertheless, NR is not stable in circulation, and its utilization is rate-limited by the expression of nicotinamide riboside kinases (NRKs). Therefore, there is a strong interest in identifying new effective NAD+ precursors that can overcome these limitations. METHODS: Through a combination of metabolomics and pharmacological approaches, we describe how NRH, a reduced form of NR, serves as a potent NAD+ precursor in mammalian cells and mice. RESULTS: NRH acts as a more potent and faster NAD+ precursor than NR in mammalian cells and tissues. Despite the minor structural difference, we found that NRH uses different steps and enzymes to synthesize NAD+, thus revealing a new NRK1-independent pathway for NAD+ synthesis. Finally, we provide evidence that NRH is orally bioavailable in mice and prevents cisplatin-induced acute kidney injury. CONCLUSIONS: Our data identify a new pathway for NAD+ synthesis and classify NRH as a promising new therapeutic strategy to enhance NAD+ levels.


Assuntos
NAD/biossíntese , NAD/metabolismo , Niacinamida/análogos & derivados , Animais , Linhagem Celular , Masculino , Camundongos , Niacinamida/metabolismo , Niacinamida/fisiologia , Fosfotransferases (Aceptor do Grupo Álcool) , Compostos de Piridínio , Ratos
SELEÇÃO DE REFERÊNCIAS
Detalhe da pesquisa