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1.
BMC Bioinformatics ; 22(1): 67, 2021 Feb 12.
Artículo en Inglés | MEDLINE | ID: mdl-33579202

RESUMEN

BACKGROUND: The search for statistically significant relationships between molecular markers and outcomes is challenging when dealing with high-dimensional, noisy and collinear multivariate omics data, such as metabolomic profiles. Permutation procedures allow for the estimation of adjusted significance levels without assuming independence among metabolomic variables. Nevertheless, the complex non-normal structure of metabolic profiles and outcomes may bias the permutation results leading to overly conservative threshold estimates i.e. lower than those from a Bonferroni or Sidak correction. METHODS: Within a univariate permutation procedure we employ parametric simulation methods based on the multivariate (log-)Normal distribution to obtain adjusted significance levels which are consistent across different outcomes while effectively controlling the type I error rate. Next, we derive an alternative closed-form expression for the estimation of the number of non-redundant metabolic variates based on the spectral decomposition of their correlation matrix. The performance of the method is tested for different model parametrizations and across a wide range of correlation levels of the variates using synthetic and real data sets. RESULTS: Both the permutation-based formulation and the more practical closed form expression are found to give an effective indication of the number of independent metabolic effects exhibited by the system, while guaranteeing that the derived adjusted threshold is stable across outcome measures with diverse properties.


Asunto(s)
Metaboloma , Metabolómica , Modelos Biológicos , Marcadores Genéticos/genética , Metabolómica/métodos , Distribuciones Estadísticas
2.
Ecotoxicol Environ Saf ; 208: 111543, 2021 Jan 15.
Artículo en Inglés | MEDLINE | ID: mdl-33396091

RESUMEN

Acrylamide (ACR) is generated during thermal processing of carbohydrate-rich foods at high temperature and can directly enter the body through ingestion, inhalation and skin contact. The toxicity of ACR has been widely studied. The main results of these studies show that exposure to ACR can cause neurotoxicity in both animals and humans, and show reproductive toxicity and carcinogenicity in rodent animal models. However, the mechanism of toxicity of ACR has not been studied by metabolomics approaches, and the effect of ACR on autophagy remains unknown. Here, U2OS cell were treated with ACR 6 and 24 h and collected for further study. We have demonstrated that ACR inhibited autophagic flux, and increased ROS content. Accumulation of ROS resulted in increase of apoptosis rates and secretion of inflammatory factors. In addition, significant differences in metabolic profiles were observed between ACR treated and control cells according to multiple analysis models. A total of 73 key differential metabolites were identified. They were involved in multiple metabolic pathways. Among them, exposure to ACR caused glycolysis/gluconeogenesis attenuation by decreasing levels of glycolytic intermediates, reduced the rate of the TCA cycle, while elevating levels of several amino acid metabolites and lipid metabolites. In summary, our study provides useful evidence of cytotoxicity caused by ACR via metabolomics and multiple bioanalytic methods.


Asunto(s)
Acrilamida/toxicidad , Sustancias Peligrosas/toxicidad , Metaboloma , Metabolómica , Animales , Apoptosis/efectos de los fármacos , Autofagia/efectos de los fármacos
3.
Nat Commun ; 12(1): 101, 2021 01 04.
Artículo en Inglés | MEDLINE | ID: mdl-33397942

RESUMEN

Western diet (WD) is one of the major culprits of metabolic disease including type 2 diabetes (T2D) with gut microbiota playing an important role in modulating effects of the diet. Herein, we use a data-driven approach (Transkingdom Network analysis) to model host-microbiome interactions under WD to infer which members of microbiota contribute to the altered host metabolism. Interrogation of this network pointed to taxa with potential beneficial or harmful effects on host's metabolism. We then validate the functional role of the predicted bacteria in regulating metabolism and show that they act via different host pathways. Our gene expression and electron microscopy studies show that two species from Lactobacillus genus act upon mitochondria in the liver leading to the improvement of lipid metabolism. Metabolomics analyses revealed that reduced glutathione may mediate these effects. Our study identifies potential probiotic strains for T2D and provides important insights into mechanisms of their action.


Asunto(s)
Diabetes Mellitus Tipo 2/etiología , Diabetes Mellitus Tipo 2/microbiología , Dieta Occidental , Lactobacillus/metabolismo , Mitocondrias Hepáticas/metabolismo , Animales , Bilirrubina/sangre , Diabetes Mellitus Tipo 2/genética , Microbioma Gastrointestinal , Regulación de la Expresión Génica , Glucosa/metabolismo , Glutatión/sangre , Glutatión/metabolismo , Humanos , Metabolismo de los Lípidos , Masculino , Metabolómica , Ratones Endogámicos C57BL , Mitocondrias Hepáticas/ultraestructura , Reproducibilidad de los Resultados , Transcriptoma/genética
4.
Anticancer Res ; 41(1): 327-334, 2021 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-33419827

RESUMEN

BACKGROUND/AIM: Pancreaticobiliary maljunction (PBM), a disease with reflux of pancreatic and bile juice in the pancreaticobiliary tract, is a high-risk factor for biliary tract cancer. The aim of this study was to investigate the mechanism of carcinogenesis in PBM using a metabolomics analysis of bile sampled during surgery. PATIENTS AND METHODS: Three patients with PBM without biliary tract cancer, four patients with extrahepatic bile duct cancer (EHBC), and three controls with benign disease were enrolled. Metabolomics analysis of bile samples was performed using capillary electrophoresis-mass spectrometry and liquid chromatography-mass spectrometry to discriminate the amino acid and lipidomic profiles. RESULTS: The principal component analysis in the capillary electrophoresis-mass spectrometry and liquid chromatography-mass spectrometry revealed similar metabolites in patients with PBM and those with EHBC; furthermore, there was a clear difference between patients with PBM or EHBC compared to controls. The amino acid profiles revealed the following 20 potential carcinogenic candidates for PBM: isoleucine, phenylalanine, tyrosine, leucine, tryptophan, arginine, lysine, valine, asparagine, methionine, aspartic acid, serine, threonine, histidine, glutamine, alanine, proline, glutamic acid, and pyruvic acid. The lipidomic profiles revealed the following 11 carcinogenic candidates: lysophosphatidylcholine, lysophosphatidylethanolamine, phosphatidyl glycerol, lysophosphatidyl glycerol, triacylglycerol, diacylglycerol, ceramide, sphyngomyeline, fatty acid, hyperforin, and vitamin D. Among these characteristic metabolites, the branched-chain amino acids, methionine and lysophosphatidylcholine are known to be related to carcinogenesis. CONCLUSION: The bile metabolites were extremely similar in patients with PBM and those with EHBC. Furthermore, amino acid and lipid metabolism was markedly different in patients with PBM or EHBC compared to healthy controls.


Asunto(s)
Neoplasias de los Conductos Biliares/etiología , Bilis/metabolismo , Transformación Celular Neoplásica/metabolismo , Susceptibilidad a Enfermedades , Mal funcionamiento Pancreaticobiliar/complicaciones , Mal funcionamiento Pancreaticobiliar/metabolismo , Neoplasias de los Conductos Biliares/diagnóstico , Neoplasias de los Conductos Biliares/terapia , Cromatografía Liquida , Electroforesis Capilar , Femenino , Humanos , Masculino , Espectrometría de Masas , Metabolómica/métodos , Proyectos Piloto , Medición de Riesgo , Factores de Riesgo
5.
Nat Commun ; 12(1): 161, 2021 01 08.
Artículo en Inglés | MEDLINE | ID: mdl-33420064

RESUMEN

Calf diarrhea is associated with enteric infections, and also provokes the overuse of antibiotics. Therefore, proper treatment of diarrhea represents a therapeutic challenge in livestock production and public health concerns. Here, we describe the ability of a fecal microbiota transplantation (FMT), to ameliorate diarrhea and restore gut microbial composition in 57 growing calves. We conduct multi-omics analysis of 450 longitudinally collected fecal samples and find that FMT-induced alterations in the gut microbiota (an increase in the family Porphyromonadaceae) and metabolomic profile (a reduction in fecal amino acid concentration) strongly correlate with the remission of diarrhea. During the continuous follow-up study over 24 months, we find that FMT improves the growth performance of the cattle. This first FMT trial in ruminants suggest that FMT is capable of ameliorating diarrhea in pre-weaning calves with alterations in their gut microbiota, and that FMT may have a potential role in the improvement of growth performance.


Asunto(s)
Enfermedades de los Bovinos/terapia , Bovinos/crecimiento & desarrollo , Diarrea/terapia , Trasplante de Microbiota Fecal/veterinaria , Microbioma Gastrointestinal/genética , Animales , Bacteroidaceae/genética , Bacteroidaceae/aislamiento & purificación , Bovinos/microbiología , Enfermedades de los Bovinos/sangre , Enfermedades de los Bovinos/metabolismo , Enfermedades de los Bovinos/microbiología , ADN Bacteriano/aislamiento & purificación , Diarrea/sangre , Diarrea/metabolismo , Diarrea/microbiología , Heces/microbiología , Femenino , Estudios de Seguimiento , Genómica , Masculino , Metabolómica , ARN Ribosómico 16S/genética , Resultado del Tratamiento
6.
Hypertension ; 77(2): 308-318, 2021 02.
Artículo en Inglés | MEDLINE | ID: mdl-33390043

RESUMEN

The intent of this review is to critically consider the data that support the concept of programming and its implications. Birth weight and growth trajectories during childhood are associated with cardiometabolic disease in adult life. Both extremes, low and high birth weight coupled with postnatal growth increase the early presence of cardiometabolic risk factors and vascular imprinting, crucial elements of this framework. Data coming from epigenetics, proteomics, metabolomics, and microbiota added relevant information and contribute to better understanding of mechanisms as well as development of biomarkers helping to move forward to take actions. Research has reached a stage in which sufficiently robust data calls for new initiatives focused on early life. Prevention starting early in life is likely to have a very large impact on reducing disease incidence and its associated effects at the personal, economic, and social levels.


Asunto(s)
Enfermedades Cardiovasculares/etiología , Epigénesis Genética , Enfermedades Metabólicas/etiología , Adolescente , Adulto , Experiencias Adversas de la Infancia , Envejecimiento , Peso al Nacer , Enfermedades Cardiovasculares/embriología , Sistema Cardiovascular/crecimiento & desarrollo , Niño , Preescolar , Epigenómica/métodos , Femenino , Predicción , Edad Gestacional , Humanos , Lactante , Recién Nacido de Bajo Peso , Recién Nacido , Recien Nacido Prematuro , Masculino , Enfermedades Metabólicas/embriología , Metabolómica , Microbiota , Persona de Mediana Edad , Medicina de Precisión , Embarazo , Efectos Tardíos de la Exposición Prenatal , Proteómica , Factores de Riesgo
7.
Nat Commun ; 12(1): 377, 2021 01 15.
Artículo en Inglés | MEDLINE | ID: mdl-33452240

RESUMEN

Circadian clocks coordinate mammalian behavior and physiology enabling organisms to anticipate 24-hour cycles. Transcription-translation feedback loops are thought to drive these clocks in most of mammalian cells. However, red blood cells (RBCs), which do not contain a nucleus, and cannot perform transcription or translation, nonetheless exhibit circadian redox rhythms. Here we show human RBCs display circadian regulation of glucose metabolism, which is required to sustain daily redox oscillations. We found daily rhythms of metabolite levels and flux through glycolysis and the pentose phosphate pathway (PPP). We show that inhibition of critical enzymes in either pathway abolished 24-hour rhythms in metabolic flux and redox oscillations, and determined that metabolic oscillations are necessary for redox rhythmicity. Furthermore, metabolic flux rhythms also occur in nucleated cells, and persist when the core transcriptional circadian clockwork is absent in Bmal1 knockouts. Thus, we propose that rhythmic glucose metabolism is an integral process in circadian rhythms.


Asunto(s)
Relojes Circadianos/fisiología , Ritmo Circadiano/fisiología , Eritrocitos/metabolismo , Glucólisis/fisiología , Vía de Pentosa Fosfato/fisiología , Factores de Transcripción ARNTL/genética , Factores de Transcripción ARNTL/metabolismo , Animales , Células Cultivadas , Fibroblastos , Técnicas de Inactivación de Genes , Voluntarios Sanos , Humanos , Masculino , Metabolómica , Ratones , Oxidación-Reducción , Cultivo Primario de Células
8.
Anal Chem ; 93(4): 1924-1933, 2021 02 02.
Artículo en Inglés | MEDLINE | ID: mdl-33448796

RESUMEN

Liquid chromatography-mass spectrometry (LC-MS) is a powerful and widely used technique for measuring the abundance of chemical species in living systems. Its sensitivity, analytical specificity, and direct applicability to biofluids and tissue extracts impart great promise for the discovery and mechanistic characterization of biomarker panels for disease detection, health monitoring, patient stratification, and treatment personalization. Global metabolic profiling applications yield complex data sets consisting of multiple feature measurements for each chemical species observed. While this multiplicity can be useful in deriving enhanced analytical specificity and chemical identities from LC-MS data, data set inflation and quantitative imprecision among related features is problematic for statistical analyses and interpretation. This Perspective provides a critical evaluation of global profiling data fidelity with respect to measurement linearity and the quantitative response variation observed among components of the spectra. These elements of data quality are widely overlooked in untargeted metabolomics yet essential for the generation of data that accurately reflect the metabolome. Advanced feature filtering informed by linear range estimation and analyte response factor assessment is advocated as an attainable means of controlling LC-MS data quality in global profiling studies and exemplified herein at both the feature and data set level.


Asunto(s)
Cromatografía Liquida/métodos , Espectrometría de Masas/métodos , Metabolómica/métodos , Metabolómica/normas , Control de Calidad , Metaboloma , Transcriptoma
9.
Anal Chem ; 93(4): 1912-1923, 2021 02 02.
Artículo en Inglés | MEDLINE | ID: mdl-33467846

RESUMEN

A growing number of software tools have been developed for metabolomics data processing and analysis. Many new tools are contributed by metabolomics practitioners who have limited prior experience with software development, and the tools are subsequently implemented by users with expertise that ranges from basic point-and-click data analysis to advanced coding. This Perspective is intended to introduce metabolomics software users and developers to important considerations that determine the overall impact of a publicly available tool within the scientific community. The recommendations reflect the collective experience of an NIH-sponsored Metabolomics Consortium working group that was formed with the goal of researching guidelines and best practices for metabolomics tool development. The recommendations are aimed at metabolomics researchers with little formal background in programming and are organized into three stages: (i) preparation, (ii) tool development, and (iii) distribution and maintenance.


Asunto(s)
Nube Computacional , Metabolómica/métodos , Programas Informáticos
10.
Mol Genet Genomics ; 296(2): 379-390, 2021 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-33449160

RESUMEN

Common bean (Phaseolus vulgaris L.) is a short-day plant and its flowering time, and consequently, pod yield and quality is influenced by photoperiod. In this study, the photoperiodic-sensitive variety 'Hong jin gou', which flowers 31 days (d) earlier in short-day than in long-day, was used as the experimental material. Samples were collected to determine the growth and photosynthetic parameters in each daylength treatment, and transcriptome and metabolome data were conducted. We identified eight genes related to flowering by further screening for differentially expressed genes. These genes function to regulate the biological clock. The combination of differentially expressed genes and metabolites, together with the known regulation network of flowering time and the day-night expression pattern of related genes allow us to speculate on the regulation of flowering time in the common bean and conclude that TIMING OF CAB EXPRESSION1 (TOC1) plays a pivotal role in the network and its upregulation or downregulation causes corresponding changes in the expression of downstream genes. The regulatory network is also influenced by gibberellic acid (GA) and jasmonic acid (JA). These regulatory pathways jointly comprise the flowering regulatory network in common bean.


Asunto(s)
Perfilación de la Expresión Génica/métodos , Metabolómica/métodos , Phaseolus/fisiología , Factores de Transcripción/genética , Relojes Biológicos , Flores/fisiología , Regulación de la Expresión Génica de las Plantas/efectos de los fármacos , Redes Reguladoras de Genes/efectos de los fármacos , Giberelinas/farmacología , Ácidos Linoleicos/farmacología , Phaseolus/genética , Phaseolus/metabolismo , Fotoperiodo , Proteínas de Plantas/genética , Análisis de Secuencia de ARN
11.
J Proteome Res ; 20(2): 1107-1132, 2021 02 05.
Artículo en Inglés | MEDLINE | ID: mdl-33426872

RESUMEN

Human infectious diseases are contributed equally by the host immune system's efficiency and any pathogens' infectivity. Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the coronavirus strain causing the respiratory pandemic coronavirus disease 2019 (COVID-19). To understand the pathobiology of SARS-CoV-2, one needs to unravel the intricacies of host immune response to the virus, the viral pathogen's mode of transmission, and alterations in specific biological pathways in the host allowing viral survival. This review critically analyzes recent research using high-throughput "omics" technologies (including proteomics and metabolomics) on various biospecimens that allow an increased understanding of the pathobiology of SARS-CoV-2 in humans. The altered biomolecule profile facilitates an understanding of altered biological pathways. Further, we have performed a meta-analysis of significantly altered biomolecular profiles in COVID-19 patients using bioinformatics tools. Our analysis deciphered alterations in the immune response, fatty acid, and amino acid metabolism and other pathways that cumulatively result in COVID-19 disease, including symptoms such as hyperglycemic and hypoxic sequelae.


Asunto(s)
/prevención & control , Metabolómica/métodos , Proteómica/métodos , /metabolismo , /epidemiología , Interacciones Huésped-Patógeno , Humanos , Pandemias , /fisiología
12.
J Proteome Res ; 20(2): 1382-1396, 2021 02 05.
Artículo en Inglés | MEDLINE | ID: mdl-33426894

RESUMEN

To investigate the systemic metabolic effects of SARS-CoV-2 infection, we analyzed 1H NMR spectroscopic data on human blood plasma and co-modeled with multiple plasma cytokines and chemokines (measured in parallel). Thus, 600 MHz 1H solvent-suppressed single-pulse, spin-echo, and 2D J-resolved spectra were collected on plasma recorded from SARS-CoV-2 rRT-PCR-positive patients (n = 15, with multiple sampling timepoints) and age-matched healthy controls (n = 34, confirmed rRT-PCR negative), together with patients with COVID-19/influenza-like clinical symptoms who tested SARS-CoV-2 negative (n = 35). We compared the single-pulse NMR spectral data with in vitro diagnostic research (IVDr) information on quantitative lipoprotein profiles (112 parameters) extracted from the raw 1D NMR data. All NMR methods gave highly significant discrimination of SARS-CoV-2 positive patients from controls and SARS-CoV-2 negative patients with individual NMR methods, giving different diagnostic information windows on disease-induced phenoconversion. Longitudinal trajectory analysis in selected patients indicated that metabolic recovery was incomplete in individuals without detectable virus in the recovery phase. We observed four plasma cytokine clusters that expressed complex differential statistical relationships with multiple lipoproteins and metabolites. These included the following: cluster 1, comprising MIP-1ß, SDF-1α, IL-22, and IL-1α, which correlated with multiple increased LDL and VLDL subfractions; cluster 2, including IL-10 and IL-17A, which was only weakly linked to the lipoprotein profile; cluster 3, which included IL-8 and MCP-1 and were inversely correlated with multiple lipoproteins. IL-18, IL-6, and IFN-γ together with IP-10 and RANTES exhibited strong positive correlations with LDL1-4 subfractions and negative correlations with multiple HDL subfractions. Collectively, these data show a distinct pattern indicative of a multilevel cellular immune response to SARS CoV-2 infection interacting with the plasma lipoproteome giving a strong and characteristic immunometabolic phenotype of the disease. We observed that some patients in the respiratory recovery phase and testing virus-free were still metabolically highly abnormal, which indicates a new role for these technologies in assessing full systemic recovery.


Asunto(s)
/diagnóstico , Quimiocinas/metabolismo , Citocinas/metabolismo , Lipoproteínas/metabolismo , Espectroscopía de Resonancia Magnética/métodos , /metabolismo , Adulto , Anciano , /virología , Quimiocinas/sangre , Citocinas/sangre , Femenino , Interacciones Huésped-Patógeno , Humanos , Lipoproteínas/sangre , Masculino , Metabolómica/métodos , Persona de Mediana Edad , Proteómica/métodos , /fisiología
13.
Nat Metab ; 3(1): 33-42, 2021 01.
Artículo en Inglés | MEDLINE | ID: mdl-33462515

RESUMEN

Mitochondrial diseases (MDs) are a heterogeneous group of disorders resulting from mutations in nuclear or mitochondrial DNA genes encoding mitochondrial proteins1,2. MDs cause pathologies with severe tissue damage and ultimately death3,4. There are no cures for MDs and current treatments are only palliative5-7. Here we show that tetracyclines improve fitness of cultured MD cells and ameliorate disease in a mouse model of Leigh syndrome. To identify small molecules that prevent cellular damage and death under nutrient stress conditions, we conduct a chemical high-throughput screen with cells carrying human MD mutations and discover a series of antibiotics that maintain survival of various MD cells. We subsequently show that a sub-library of tetracycline analogues, including doxycycline, rescues cell death and inflammatory signatures in mutant cells through partial and selective inhibition of mitochondrial translation, resulting in an ATF4-independent mitohormetic response. Doxycycline treatment strongly promotes fitness and survival of Ndufs4-/- mice, a preclinical Leigh syndrome mouse model8. A proteomic analysis of brain tissue reveals that doxycycline treatment largely prevents neuronal death and the accumulation of neuroimmune and inflammatory proteins in Ndufs4-/- mice, indicating a potential causal role for these proteins in the brain pathology. Our findings suggest that tetracyclines deserve further evaluation as potential drugs for the treatment of MDs.


Asunto(s)
Antibacterianos/uso terapéutico , Enfermedades Mitocondriales/tratamiento farmacológico , Tetraciclinas/uso terapéutico , Factor de Transcripción Activador 4/metabolismo , Animales , Encéfalo/patología , Células Cultivadas , Modelos Animales de Enfermedad , Complejo I de Transporte de Electrón/genética , Complejo I de Transporte de Electrón/metabolismo , Ensayos Analíticos de Alto Rendimiento , Humanos , Enfermedad de Leigh/tratamiento farmacológico , Enfermedad de Leigh/patología , Esperanza de Vida , Metabolómica , Ratones , Ratones Noqueados , Enfermedades Mitocondriales/mortalidad , Enfermedades Mitocondriales/patología , Aptitud Física , Análisis de Supervivencia
14.
J Proteome Res ; 20(2): 1415-1423, 2021 02 05.
Artículo en Inglés | MEDLINE | ID: mdl-33491459

RESUMEN

The utility of low sample volume in vitro diagnostic (IVDr) proton nuclear magnetic resonance (1H NMR) spectroscopic experiments on blood plasma for information recovery from limited availability or high value samples was exemplified using plasma from patients with SARS-CoV-2 infection and normal controls. 1H NMR spectra were obtained using solvent-suppressed 1D, spin-echo (CPMG), and 2-dimensional J-resolved (JRES) spectroscopy using both 3 mm outer diameter SampleJet NMR tubes (100 µL plasma) and 5 mm SampleJet NMR tubes (300 µL plasma) under in vitro diagnostic conditions. We noted near identical diagnostic models in both standard and low volume IVDr lipoprotein analysis (measuring 112 lipoprotein parameters) with a comparison of the two tubes yielding R2 values ranging between 0.82 and 0.99 for the 40 paired lipoprotein parameters samples. Lipoprotein measurements for the 3 mm tubes were achieved without time penalty over the 5 mm tubes as defined by biomarker recovery for SARS-CoV-2. Overall, biomarker pattern recovery for the lipoproteins was extremely similar, but there were some small positive offsets in the linear equations for several variables due to small shimming artifacts, but there was minimal degradation of the biological information. For the standard untargeted 1D, CPMG, and JRES NMR experiments on the same samples, the reduced signal-to-noise was more constraining and required greater scanning times to achieve similar differential diagnostic performance (15 min per sample per experiment for 3 mm 1D and CPMG, compared to 4 min for the 5 mm tubes). We conclude that the 3 mm IVDr method is fit-for-purpose for quantitative lipoprotein measurements, allowing the preparation of smaller volumes for high value or limited volume samples that is common in clinical studies. If there are no analytical time constraints, the lower volume experiments are equally informative for untargeted profiling.


Asunto(s)
/diagnóstico , Lipoproteínas/metabolismo , Metabolómica/métodos , Proteómica/métodos , Espectroscopía de Protones por Resonancia Magnética/métodos , /metabolismo , Adulto , Anciano , Biomarcadores/sangre , Biomarcadores/metabolismo , /virología , Femenino , Humanos , Lipoproteínas/sangre , Masculino , Persona de Mediana Edad , Mapas de Interacción de Proteínas , /fisiología
15.
Nat Commun ; 12(1): 530, 2021 01 22.
Artículo en Inglés | MEDLINE | ID: mdl-33483501

RESUMEN

The emergence and spread of artemisinin resistance, driven by mutations in Plasmodium falciparum K13, has compromised antimalarial efficacy and threatens the global malaria elimination campaign. By applying systems-based quantitative transcriptomics, proteomics, and metabolomics to a panel of isogenic K13 mutant or wild-type P. falciparum lines, we provide evidence that K13 mutations alter multiple aspects of the parasite's intra-erythrocytic developmental program. These changes impact cell-cycle periodicity, the unfolded protein response, protein degradation, vesicular trafficking, and mitochondrial metabolism. K13-mediated artemisinin resistance in the Cambodian Cam3.II line was reversed by atovaquone, a mitochondrial electron transport chain inhibitor. These results suggest that mitochondrial processes including damage sensing and anti-oxidant properties might augment the ability of mutant K13 to protect P. falciparum against artemisinin action by helping these parasites undergo temporary quiescence and accelerated growth recovery post drug elimination.


Asunto(s)
Artemisininas/farmacología , Resistencia a Medicamentos/genética , Eritrocitos/metabolismo , Mutación , Plasmodium falciparum/genética , Antimaláricos/farmacología , Atovacuona/farmacología , Puntos de Control del Ciclo Celular/genética , Eritrocitos/parasitología , Perfilación de la Expresión Génica/métodos , Humanos , Metabolómica/métodos , Mitocondrias/genética , Mitocondrias/metabolismo , Modelos Genéticos , Plasmodium falciparum/metabolismo , Plasmodium falciparum/fisiología , Proteómica/métodos , Proteínas Protozoarias/genética , Proteínas Protozoarias/metabolismo
16.
J Chromatogr A ; 1638: 461862, 2021 Feb 08.
Artículo en Inglés | MEDLINE | ID: mdl-33433374

RESUMEN

This work presents an evaluation of solid-phase microextraction (SPME) SPME in combination with liquid chromatography-high resolution mass spectrometry (LC-HRMS) as an analytical approach for untargeted brain analysis. The study included a characterization of the metabolite coverage provided by C18, mixed-mode (MM, with benzene sulfonic acid and C18 functionalities), and hydrophilic lipophilic balanced (HLB) particles as sorbents in SPME coatings after extraction from cow brain homogenate at static conditions. The effects of desorption solvent, extraction time, and chromatographic modes on the metabolite features detected were investigated. Method precision and absolute matrix effects were also assessed. Among the main findings of this work, it was observed that all three tested coating chemistries were able to provide comparable brain tissue information. HLB provided higher responses for polar metabolites; however, as these fibers were prepared in-house, higher inter-fiber relative standard deviations were also observed. C18 and HLB coatings offered similar responses with respect to lipid-related features, whereas MM and C18 provided the best results in terms of method precision. Our results also showed that the use of methanol is essential for effective desorption of non-polar metabolites. Using a reversed-phase chromatographic method, an average of 800 and 1200 brain metabolite features detected in positive and negative modes, respectively, met inter-fibre RSD values below 30% (n=4) after removal of fibre and solvent artefacts from the associated datasets. For features detected using a lipidomics method, a total of 900 and 1800 features detected using C18 fibers in positive and negative mode, respectively, met the same criteria. In terms of absolute matrix effects, the majority of the model metabolites tested showed values between 80 and 120%, which are within the acceptable range. Overall, the findings of this work lay the foundation for further optimization of parameters for SPME-LC-HRMS methods suitable for in vivo and ex vivo brain (and other tissue) untargeted studies, and support the applicability of this approach for non-destructive tissue metabolomics.


Asunto(s)
Encéfalo/metabolismo , Cromatografía Liquida , Espectrometría de Masas , Microextracción en Fase Sólida , Animales , Bovinos , Interacciones Hidrofóbicas e Hidrofílicas , Metabolómica/métodos , Solventes/química , Manejo de Especímenes
17.
Chemosphere ; 268: 129362, 2021 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-33385834

RESUMEN

This study investigated the effects of accidental contamination of soils with phenol, toluene, nitric acid, and hydrogen fluoride (HF) by simulating chemical leakage in the soil with/without rain and characterizing the resulting metabolites and microbial. In the case of acid leakage, pH and cation exchange capacity were decreased, and the content of fluoride ion was increased in case of HF leakage. Using mass spectrometry-based metabolomics analysis, phytosphingosine was detected as a distinguishing metabolite in soils contaminated with phenol and HF in rain conditions. Microbial communities were identified by 16s rRNA metagenome sequencing. Sphingomonas was one of the dominant species in soils contaminated with phenol and HF. These results suggest that phytosphingosine and Sphingomonas might be used as biomarkers to evaluate the status of soils contaminated with phenol or HF. Under simulated rain conditions, the species alpha-diversity index of soil microbes and the physicochemical properties of the soil indicated values close to those of the uncontaminated soil. Rain played an important role in the recovery of microbial and metabolic profiles after chemical accidents. Metabolic profiling and microbial community analysis can serve as a diagnostic tool for ecotoxicological research at chemical accident sites.


Asunto(s)
Liberación de Peligros Químicos , Microbiota , Contaminantes del Suelo , Metabolómica , ARN Ribosómico 16S/genética , Suelo , Microbiología del Suelo , Contaminantes del Suelo/análisis , Contaminantes del Suelo/toxicidad
18.
Anal Chem ; 93(4): 2471-2479, 2021 02 02.
Artículo en Inglés | MEDLINE | ID: mdl-33471512

RESUMEN

COVID-19 is still placing a heavy health and financial burden worldwide. Impairment in patient screening and risk management plays a fundamental role on how governments and authorities are directing resources, planning reopening, as well as sanitary countermeasures, especially in regions where poverty is a major component in the equation. An efficient diagnostic method must be highly accurate, while having a cost-effective profile. We combined a machine learning-based algorithm with mass spectrometry to create an expeditious platform that discriminate COVID-19 in plasma samples within minutes, while also providing tools for risk assessment, to assist healthcare professionals in patient management and decision-making. A cross-sectional study enrolled 815 patients (442 COVID-19, 350 controls and 23 COVID-19 suspicious) from three Brazilian epicenters from April to July 2020. We were able to elect and identify 19 molecules related to the disease's pathophysiology and several discriminating features to patient's health-related outcomes. The method applied for COVID-19 diagnosis showed specificity >96% and sensitivity >83%, and specificity >80% and sensitivity >85% during risk assessment, both from blinded data. Our method introduced a new approach for COVID-19 screening, providing the indirect detection of infection through metabolites and contextualizing the findings with the disease's pathophysiology. The pairwise analysis of biomarkers brought robustness to the model developed using machine learning algorithms, transforming this screening approach in a tool with great potential for real-world application.


Asunto(s)
/diagnóstico , Aprendizaje Automático , Metabolómica , Adulto , Anciano , Automatización , Biomarcadores/metabolismo , Brasil , Femenino , Humanos , Masculino , Persona de Mediana Edad , Medición de Riesgo , /aislamiento & purificación
19.
J Vis Exp ; (167)2021 01 05.
Artículo en Inglés | MEDLINE | ID: mdl-33491678

RESUMEN

Metabolomics is a methodology used for the identification and quantification of many low-molecular-weight intermediates and products of metabolism within a cell, tissue, organ, biological fluid, or organism. Metabolomics traditionally focuses on water-soluble metabolites. The water-soluble metabolome is the final product of a complex cellular network that integrates various genomic, epigenomic, transcriptomic, proteomic, and environmental factors. Hence, the metabolomic analysis directly assesses the outcome of the action for all these factors in a plethora of biological processes within various organisms. One of these organisms is the budding yeast Saccharomyces cerevisiae, a unicellular eukaryote with the fully sequenced genome. Because S. cerevisiae is amenable to comprehensive molecular analyses, it is used as a model for dissecting mechanisms underlying many biological processes within the eukaryotic cell. A versatile analytical method for the robust, sensitive, and accurate quantitative assessment of the water-soluble metabolome would provide the essential methodology for dissecting these mechanisms. Here we present a protocol for the optimized conditions of metabolic activity quenching in and water-soluble metabolite extraction from S. cerevisiae cells. The protocol also describes the use of liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) for the quantitative analysis of the extracted water-soluble metabolites. The LC-MS/MS method of non-targeted metabolomics described here is versatile and robust. It enables the identification and quantification of more than 370 water-soluble metabolites with diverse structural, physical, and chemical properties, including different structural isomers and stereoisomeric forms of these metabolites. These metabolites include various energy carrier molecules, nucleotides, amino acids, monosaccharides, intermediates of glycolysis, and tricarboxylic cycle intermediates. The LC-MS/MS method of non-targeted metabolomics is sensitive and allows the identification and quantitation of some water-soluble metabolites at concentrations as low as 0.05 pmol/µL. The method has been successfully used for assessing water-soluble metabolomes of wild-type and mutant yeast cells cultured under different conditions.


Asunto(s)
Metabolómica , Saccharomyces cerevisiae/metabolismo , Espectrometría de Masas en Tándem/métodos , Cromatografía Liquida , Fluorescencia , Glucólisis , Metaboloma , Proteómica , Solubilidad , Agua
20.
Nat Commun ; 12(1): 187, 2021 01 08.
Artículo en Inglés | MEDLINE | ID: mdl-33420074

RESUMEN

The gut microbiota is reported to modulate the immune response in hepatocellular carcinoma (HCC). Here, we employ metagenomic and metabolomic studies to characterise gut microbiota in patients with non-alcoholic fatty liver disease (NAFLD) related cirrhosis, with or without HCC, and evaluate its effect on the peripheral immune response in an ex vivo model. We find that dysbiosis characterises the microbiota of patients with NAFLD-cirrhosis, with compositional and functional shifts occurring with HCC development. Gene function of the microbiota in NAFLD-HCC supports short chain fatty acid production, and this is confirmed by metabolomic studies. Ex vivo studies show that bacterial extracts from the NAFLD-HCC microbiota, but not from the control groups, elicit a T cell immunosuppressive phenotype, characterised by expansion of regulatory T cells and attenuation of CD8 + T cells. Our study suggest that the gut microbiota in NAFLD-HCC is characterised by a distinctive microbiome/metabolomic profile, and can modulate the peripheral immune response.


Asunto(s)
Carcinoma Hepatocelular/inmunología , Microbioma Gastrointestinal/inmunología , Microbioma Gastrointestinal/fisiología , Inmunidad , Neoplasias Hepáticas/inmunología , Enfermedad del Hígado Graso no Alcohólico/inmunología , Anciano , Bacterias/genética , Linfocitos T CD8-positivos , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/patología , Citocinas , Fibras de la Dieta , Disbiosis/inmunología , Ácidos Grasos Volátiles/sangre , Ácidos Grasos Volátiles/metabolismo , Heces/química , Femenino , Humanos , Hígado/patología , Cirrosis Hepática , Neoplasias Hepáticas/patología , Masculino , Metabolómica , Metagenómica , Persona de Mediana Edad , Enfermedad del Hígado Graso no Alcohólico/genética , Enfermedad del Hígado Graso no Alcohólico/patología , Fenotipo
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