Lipidomics and metabolomics communities comprise various informatics tools; however, software programs handling multimodal mass spectrometry (MS) data with structural annotations guided by the Lipidomics Standards Initiative are limited. Here, we provide MS-DIAL 5 for in-depth lipidome structural elucidation through electron-activated dissociation (EAD)-based tandem MS and determining their molecular localization through MS imaging (MSI) data using a species/tissue-specific lipidome database containing the predicted collision-cross section values. With the optimized EAD settings using 14 eV kinetic energy, the program correctly delineated lipid structures for 96.4% of authentic standards, among which 78.0% had the sn-, OH-, and/or C = C positions correctly assigned at concentrations exceeding 1 μM. We showcased our workflow by annotating the sn- and double-bond positions of eye-specific phosphatidylcholines containing very-long-chain polyunsaturated fatty acids (VLC-PUFAs), characterized as PC n-3-VLC-PUFA/FA. Using MSI data from the eye and n-3-VLC-PUFA-supplemented HeLa cells, we identified glycerol 3-phosphate acyltransferase as an enzyme candidate responsible for incorporating n-3 VLC-PUFAs into the sn1 position of phospholipids in mammalian cells, which was confirmed using EAD-MS/MS and recombinant proteins in a cell-free system. Therefore, the MS-DIAL 5 environment, combined with optimized MS data acquisition methods, facilitates a better understanding of lipid structures and their localization, offering insights into lipid biology.
- MeSH
- data mining * metody MeSH
- fosfatidylcholiny metabolismus chemie MeSH
- HeLa buňky MeSH
- hmotnostní spektrometrie metody MeSH
- lidé MeSH
- lipidomika * metody MeSH
- lipidy chemie analýza MeSH
- metabolomika metody MeSH
- nenasycené mastné kyseliny metabolismus chemie MeSH
- software MeSH
- tandemová hmotnostní spektrometrie metody MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
BACKGROUND: Heart failure (HF) is becoming an increasingly prevalent issue, particularly among the elderly population. Lipids are closely associated with cardiovascular disease (CVD) pathology. Lipidomics as a comprehensive profiling tool is showing to be promising in the prediction of events and mortality due to CVD as well as identifying novel biomarkers. MATERIALS AND METHODS: In this study, eicosanoids and lipid profiles were measured in order to predict survival in patients with de novo or acute decompensated HF. Our study included 50 patients (16 females, mean age 73 years and 34 males, mean age 71 years) with de novo or acute decompensated chronic HF with a median follow-up of 7 months. Lipids were semiquantified using targeted lipidomic liquid chromatography-mass spectrometry (LC-MS/MS) analysis. Eicosanoid concentrations were determined using a commercially available sandwich ELISA assay. RESULTS: From 736 lipids and 3 eicosanoids, 39 significant lipids were selected (by using the Mann-Whitney U test after Benjamini-Hochberg correction) with the highest number of representatives belonging to the polyunsaturated (PUFA) phosphatidylcholines (PC). PC 42:10 (p = 1.44 × 10-4) was found to be the most statistically significantly elevated in the surviving group with receiver operating characteristics of AUC = 0.84 (p = 3.24 × 10-7). A multivariate supervised discriminant analysis based on the aforementioned lipid panel enabled the classification of the groups of surviving and non-surviving patients with 90 % accuracy. CONCLUSIONS: In the present study we describe a trend in PUFA esterified in PC that were systematically increased in surviving patients with HF. This trend in low-abundant and rarely identified PUFA PC (mainly very long chain PUFA containing PC such as PC 42:10 or PC 40:9 containing FA 22:6, FA 20:5 and FA 20:4) suggests candidate biomarkers.
- Publikační typ
- časopisecké články MeSH
BACKGROUND: Accumulation of tau leads to neuroinflammation and neuronal cell death in tauopathies, including Alzheimer's disease. As the disease progresses, there is a decline in brain energy metabolism. However, the role of tau protein in regulating lipid metabolism remains less characterized and poorly understood. METHODS: We used a transgenic rat model for tauopathy to reveal metabolic alterations induced by neurofibrillary pathology. Transgenic rats express a tau fragment truncated at the N- and C-terminals. For phenotypic profiling, we performed targeted metabolomic and lipidomic analysis of brain tissue, CSF, and plasma, based on the LC-MS platform. To monitor disease progression, we employed samples from transgenic and control rats aged 4, 6, 8, 10, 12, and 14 months. To study neuron-glia interplay in lipidome changes induced by pathological tau we used well well-established multicomponent cell model system. Univariate and multivariate statistical approaches were used for data evaluation. RESULTS: We showed that tau has an important role in the deregulation of lipid metabolism. In the lipidomic study, pathological tau was associated with higher production of lipids participating in protein fibrillization, membrane reorganization, and inflammation. Interestingly, significant changes have been found in the early stages of tauopathy before the formation of high-molecular-weight tau aggregates and neurofibrillary pathology. Increased secretion of pathological tau protein in vivo and in vitro induced upregulated production of phospholipids and sphingolipids and accumulation of lipid droplets in microglia. We also found that this process depended on the amount of extracellular tau. During the later stages of tauopathy, we found a connection between the transition of tau into an insoluble fraction and changes in brain metabolism. CONCLUSION: Our results revealed that lipid metabolism is significantly affected during different stages of tau pathology. Thus, our results demonstrate that the dysregulation of lipid composition by pathological tau disrupts the microenvironment, further contributing to the propagation of pathology.
- MeSH
- Alzheimerova nemoc * patologie MeSH
- krysa rodu rattus MeSH
- metabolismus lipidů MeSH
- modely nemocí na zvířatech MeSH
- mozek metabolismus MeSH
- myši transgenní MeSH
- myši MeSH
- neurofibrilární klubka metabolismus MeSH
- potkani transgenní MeSH
- proteiny tau genetika metabolismus MeSH
- tauopatie * patologie MeSH
- zvířata MeSH
- Check Tag
- krysa rodu rattus MeSH
- myši MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
Glutamate carboxypeptidase II (GCPII, also known as PSMA or FOLH1) is responsible for the cleavage of N-acetyl-aspartyl-glutamate (NAAG) to N-acetyl-aspartate and glutamate in the central nervous system and facilitates the intestinal absorption of folate by processing dietary folyl-poly-γ-glutamate in the small intestine. The physiological function of GCPII in other organs like kidneys is still not known. GCPII inhibitors are neuroprotective in various conditions (e.g., ischemic brain injury) in vivo; however, their utilization as potential drug candidates has not been investigated in regard to not yet known GCPII activities. To explore the GCPII role and possible side effects of GCPII inhibitors, we performed parallel metabolomic and lipidomic analysis of the cerebrospinal fluid (CSF), urine, plasma, and brain tissue of mice with varying degrees of GCPII deficiency (fully deficient in Folh1, -/-; one allele deficient in Folh1, +/-; and wild type, +/+). Multivariate analysis of metabolites showed no significant differences between wild-type and GCPII-deficient mice (except for NAAG), although changes were observed between the sex and age. NAAG levels were statistically significantly increased in the CSF, urine, and plasma of GCPII-deficient mice. However, no difference in NAAG concentrations was found in the whole brain lysate likely because GCPII, as an extracellular enzyme, can affect only extracellular and not intracellular NAAG concentrations. Regarding the lipidome, the most pronounced genotype-linked changes were found in the brain tissue. In brains of GCPII-deficient mice, we observed statistically significant enrichment in phosphatidylcholine-based lipids and reduction of sphingolipids and phosphatidylethanolamine plasmalogens. We hypothesize that the alteration of the NAA-NAAG axis by absent GCPII activity affected myelin composition. In summary, the absence of GCPII and thus similarly its inhibition do not have detrimental effects on metabolism, with just minor changes in the brain lipidome.
- MeSH
- dipeptidy metabolismus MeSH
- glutamátkarboxypeptidasa II * genetika metabolismus MeSH
- kyselina glutamová MeSH
- lipidomika * MeSH
- lipidy chemie MeSH
- metabolomika * MeSH
- mozek metabolismus MeSH
- myši MeSH
- zvířata MeSH
- Check Tag
- myši MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
OBJECTIVE: The laboratory diagnosis of inherited metabolic disorders (IMD) has undergone significant development in recent decades, mainly due to the use of mass spectrometry, which allows rapid multicomponent analysis of a wide range of metabolites. Combined with advanced software tools, the diagnosis becomes more efficient as a benefit for both physicians and patients. METHODS: A hydrophilic interaction liquid chromatography coupled with tandem mass spectrometry assay for determination of urinary purines, pyrimidines, N-acylglycines, N-acetylated amino acids, sugars, sugar alcohols and other diagnostically important biomarkers was developed and validated. Evaluation of the results consisting of utilisation of robust scaling and advanced visualization tools is simple and even suitable for urgent requirements. RESULTS: The developed method, covering 65 biomarkers, provides a comprehensive diagnostic platform for 51 IMD. For most analytes, linearity with R2 > 0.99, intra and inter-day accuracy between 80 and 120 % and precision lower than 20 % were achieved. Diagnostic workflow was evaluated on 47 patients and External Quality Assurance samples involving a total of 24 different IMD. Over seven years, more than 2300 urine samples from patients suspected for IMD have been routinely analysed. CONCLUSIONS: This method offers the advantage of a broad coverage of intermediate metabolites of interest and therefore may be a potential alternative and simplification for clinical laboratories that use multiple methods for screening these markers.
Some pathological conditions affecting the human body can also disrupt metabolic pathways and thus alter the overall metabolic profile. Knowledge of metabolic disturbances in specific diseases could thus enable the differential diagnosis of otherwise similar conditions. This work therefore aimed to comprehensively characterize changes in tryptophan metabolism in selected neurodegenerative diseases. Levels of 18 tryptophan-related neuroactive substances were determined by high throughput and sensitive ultrahigh-performance liquid chromatography-tandem mass spectrometry in time-linked blood serum and cerebrospinal fluid samples from 100 age-matched participants belonging to five cohorts: healthy volunteers (n = 21) and patients with Lewy body disease (Parkinson's disease and dementia with Lewy bodies; n = 31), four-repeat tauopathy (progressive supranuclear palsy and corticobasal syndrome; n = 10), multiple system atrophy (n = 13), and Alzheimer's disease (n = 25). Although these conditions have different pathologies and clinical symptoms, the discovery of new biomarkers is still important. The most statistically significant differences (with p-values of ≤0.05 to ≤0.0001) between the study cohorts were observed for three tryptophan metabolites: l-kynurenine in cerebrospinal fluid and 3-hydroxy-l-kynurenine and 5-hydroxy-l-tryptophan in blood serum. This led to the discovery of distinctive correlation patterns between the profiled cerebrospinal fluid and serum metabolites that could provide a basis for the differential diagnosis of neurodegenerative tauopathies and synucleinopathies. However, further large-scale studies are needed to determine the direct involvement of these metabolites in the studied neuropathologies, their response to medication, and their potential therapeutic relevance.
- MeSH
- Alzheimerova nemoc * diagnóza MeSH
- biologické markery MeSH
- kynurenin MeSH
- lidé MeSH
- poruchy proteostázy * MeSH
- sérum MeSH
- tauopatie * MeSH
- tryptofan MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
BACKGROUND: Currently, it is not possible to predict whether patients with hyperuricemia (HUA) will develop gout and how this progression may be affected by urate-lowering treatment (ULT). Our study aimed to evaluate differences in plasma lipidome between patients with asymptomatic HUA detected ≤ 40 years (HUA ≤ 40) and > 40 years, gout patients with disease onset ≤ 40 years (Gout ≤ 40) and > 40 years, and normouricemic healthy controls (HC). METHODS: Plasma samples were collected from 94 asymptomatic HUA (77% HUA ≤ 40) subjects, 196 gout patients (59% Gout ≤ 40), and 53 HC. A comprehensive targeted lipidomic analysis was performed to semi-quantify 608 lipids in plasma. Univariate and multivariate statistics and advanced visualizations were applied. RESULTS: Both HUA and gout patients showed alterations in lipid profiles with the most significant upregulation of phosphatidylethanolamines and downregulation of lysophosphatidylcholine plasmalogens/plasmanyls. More profound changes were observed in HUA ≤ 40 and Gout ≤ 40 without ULT. Multivariate statistics differentiated HUA ≤ 40 and Gout ≤ 40 groups from HC with an overall accuracy of > 95%. CONCLUSION: Alterations in the lipidome of HUA and Gout patients show a significant impact on lipid metabolism. The most significant glycerophospholipid dysregulation was found in HUA ≤ 40 and Gout ≤ 40 patients, together with a correction of this imbalance with ULT.
- MeSH
- antiuratika terapeutické užití MeSH
- dna (nemoc) * diagnóza farmakoterapie MeSH
- hyperurikemie * diagnóza farmakoterapie MeSH
- kyselina močová MeSH
- lidé MeSH
- lipidomika MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
OBJECTIVES: The analysis of organic acids in urine is an important part of the diagnosis of inherited metabolic disorders (IMDs), for which gas chromatography coupled with mass spectrometry is still predominantly used. METHODS: Ultra-performance liquid chromatography-tandem mass spectrometry (LC-MS/MS) assay for urinary organic acids, acylcarnitines and acylglycines was developed and validated. Sample preparation consists only of dilution and the addition of internal standards. Raw data processing is quick and easy using selective scheduled multiple reaction monitoring mode. A robust standardised value calculation as a data transformation together with advanced automatic visualisation tools are applied for easy evaluation of complex data. RESULTS: The developed method covers 146 biomarkers consisting of organic acids (n=99), acylglycines (n=15) and acylcarnitines (n=32) including all clinically important isomeric compounds present. Linearity with r2>0.98 for 118 analytes, inter-day accuracy between 80 and 120 % and imprecision under 15 % for 120 analytes were achieved. Over 2 years, more than 800 urine samples from children tested for IMDs were analysed. The workflow was evaluated on 93 patient samples and ERNDIM External Quality Assurance samples involving a total of 34 different IMDs. CONCLUSIONS: The established LC-MS/MS workflow offers a comprehensive analysis of a wide range of organic acids, acylcarnitines and acylglycines in urine to perform effective, rapid and sensitive semi-automated diagnosis of more than 80 IMDs.
- MeSH
- chromatografie kapalinová metody MeSH
- dítě MeSH
- lidé MeSH
- metabolické nemoci * MeSH
- organické látky MeSH
- průběh práce MeSH
- tandemová hmotnostní spektrometrie * metody MeSH
- Check Tag
- dítě MeSH
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
Stanovení hladin lipidů představuje rutinní servis poskytovaný laboratořemi klinické biochemie. Zahrnuje však jen několik parametrů jako jsou triacylglyceroly, cholesterol a lipoproteinové částice. Díky rozvoji hmotnostní spektrometrie vznikl samostatný odbor lipidomika, který umožňuje komplexní popis a kvantifikaci lipidomu skýtajícího stovky individuálních molekul. Nadějným směrem využití lipidomiky v rutinní laboratorní diagnostice se v posledních letech ukazuje oblast kardiovaskulárních chorob. Poměry hladin vybraných ceramidů a fosfatidylcholinů v plasmě jsou kombinovány k vyhodnocení skóre CERT1, CERT2 a dalších, které slouží k predikci komplikací a úmrtí pacientů s kardiovaskulárními chorobami. V tomto přehledovém článku bude blíže popsána výše uvedená problematika a diskutován potenciál lipidomiky v rutinní laboratorní diagnostice.
The determination of lipid levels is a routine service provided by clinical biochemistry laboratories. It commonly includes several parameters such as triacylglycerols, cholesterol and lipoprotein particles. However, with the development of mass spectrometry, a separate field of lipidomics has emerged, which allows the comprehensive description and quantification of a lipidome containing hundreds of individual molecules. A promising direction for the use of lipidomics in routine laboratory diagnostics has emerged in recent years in the field of cardiovascular disease. Ratios of plasma levels of selected ceramides and phosphatidylcholines are combined to evaluate CERT1, CERT2 and other scores to predict events and death in patients with cardiovascular disease. This review article will elaborate on the above issues and discuss the potential of lipidomics in routine laboratory diagnostics.
- Publikační typ
- abstrakt z konference MeSH
