RESUMO
Toll-like receptors (TLR) are crucial for recognizing bacterial, viral or fungal pathogens and to orchestrate the appropriate immune response. The widely expressed TLR2 and TLR4 differentially recognize various pathogens to initiate partly overlapping immune cascades. To better understand the physiological consequences of both immune responses, we performed comparative lipidomic analyses of local paw inflammation in mice induced by the TLR2 and TLR4 agonists, zymosan and lipopolysaccharide (LPS), respectively, which are commonly used in models for inflammation and inflammatory pain. Doses for both agonists were chosen to cause mechanical hypersensitivity with identical strength and duration. Lipidomic analysis showed 5 h after LPS or zymosan injection in both models an increase of ether-phosphatidylcholines (PC O) and their corresponding lyso species with additional lipids being increased only in response to LPS. However, zymosan induced stronger immune cell recruitment and edema formation as compared to LPS. Importantly, only in LPS-induced inflammation the lipid profile in the contralateral paw was altered. Fittingly, the plasma level of various cytokines and chemokines, including IL-1ß and IL-6, were significantly increased only in LPS-treated mice. Accordingly LPS induced distinct changes in the lipid profiles of ipsilateral and contralateral paws. Here, oxydized fatty acids, phosphatidylcholines and phosphatidylethanolamines were uniquely upregulated on the contralateral side. Thus, both models cause increased levels of PC O and lyso-PC O lipids at the site of inflammation pointing at a common role in inflammation. Also, LPS initiates systemic changes, which can be detected by changes in the lipid profiles.
Assuntos
Reação de Fase Aguda/sangue , Edema/sangue , Lipopolissacarídeos/administração & dosagem , Fosfatidilcolinas/sangue , Fosfatidiletanolaminas/sangue , Zimosan/administração & dosagem , Reação de Fase Aguda/induzido quimicamente , Reação de Fase Aguda/genética , Reação de Fase Aguda/patologia , Animais , Edema/induzido quimicamente , Edema/genética , Edema/patologia , Ácidos Graxos/sangue , Ácidos Graxos/classificação , Regulação da Expressão Gênica , Membro Posterior/irrigação sanguínea , Membro Posterior/efeitos dos fármacos , Membro Posterior/metabolismo , Interleucina-1beta/sangue , Interleucina-1beta/genética , Interleucina-6/sangue , Interleucina-6/genética , Lipidômica/métodos , Camundongos , Camundongos Endogâmicos C57BL , Fosfatidilcolinas/classificação , Fosfatidiletanolaminas/classificação , Transdução de Sinais , Receptor 2 Toll-Like/sangue , Receptor 2 Toll-Like/genética , Receptor 4 Toll-Like/sangue , Receptor 4 Toll-Like/genéticaRESUMO
Obtaining fast screening information on molecular composition of a tissue sample is of great importance for a disease biomarkers search and for online surgery control. In this study, high resolution mass spectrometry analysis of eutopic and ectopic endometrium tissues (90 samples) is done using direct tissue spray mass spectrometry in both positive and negative ion modes. The most abundant peaks in the both ion modes are those corresponding to lipids. Species of three lipid classes are observed, phosphatidylcholines (PC), sphingomyelins (SM) and phosphoethanolamines (PE). Direct tissue analysis gives mainly information on PC and SM lipids (29 species) in positive ion mode and PC, SM and PE lipids (50 species) in negative ion mode which gives complementary data for endometriosis foci differentiation. The biggest differences were found for phospholipids with polyunsaturated acyls and alkils. Although, tissue spray shows itself as appropriate tool for tissue investigation, caution should be paid to the interpretation of mass spectra because of their higher complexity with more possible adducts formation and multiple interferences must be taken into account. The present work extends the application of direct tissue analysis for the rapid differentiation between endometriotic tissues of different foci.
Assuntos
Endometriose/diagnóstico , Cistos Ovarianos/diagnóstico , Fosfatidilcolinas/isolamento & purificação , Fosfatidiletanolaminas/isolamento & purificação , Esfingomielinas/isolamento & purificação , Adulto , Estudos de Casos e Controles , Diagnóstico Diferencial , Endometriose/metabolismo , Endometriose/patologia , Endometriose/cirurgia , Endométrio/metabolismo , Endométrio/patologia , Endométrio/cirurgia , Feminino , Humanos , Metabolismo dos Lipídeos , Metaboloma , Pessoa de Meia-Idade , Cistos Ovarianos/metabolismo , Cistos Ovarianos/patologia , Cistos Ovarianos/cirurgia , Fosfatidilcolinas/classificação , Fosfatidilcolinas/metabolismo , Fosfatidiletanolaminas/classificação , Fosfatidiletanolaminas/metabolismo , Espectrometria de Massas por Ionização por Electrospray , Esfingomielinas/classificação , Esfingomielinas/metabolismoRESUMO
Significant sensitivity enhancements in the tandem mass spectrometry-based analysis of complex mixtures of several phospholipid classes has been achieved via (13)C-TrEnDi. (13)C-TrEnDi-modified phosphatidylethanolamine (PE), phosphatidylserine (PS), and phosphatidylcholine (PC) lipids extracted from HeLa cells demonstrated greater sensitivity via precursor ion scans (PISs) than their unmodified counterparts. Sphingomyelin (SM) species exhibited neither an increased nor decreased sensitivity following modification. The use of isotopically labeled diazomethane enabled the distinction of modified PE and modified PC species that would yield isobaric species with unlabeled diazomethane. (13)C-TrEnDi created a PE-exclusive PIS of m/z 202.1, two PS-exclusive PISs of m/z 148.1 and m/z 261.1, and a PIS of m/z 199.1 for PC species (observed at odd m/z values) and SM species (observed at even m/z values). The standardized average area increase after TrEnDi modification was 10.72-fold for PE species, 2.36-fold for PC, and 1.05-fold for SM species. The sensitivity increase of PS species was not quantifiable, as there were no unmodified PS species identified prior to derivatization. (13)C-TrEnDi allowed for the identification of 4 PE and 7 PS species as well as the identification and quantitation of an additional 4 PE and 4 PS species that were below the limit of detection (LoD) prior to modification. (13)C-TrEnDi also pushed 24 PE and 6 PC lipids over the limit of quantitation (LoQ) that prior to modification were above the LoD only.
Assuntos
Diazometano/química , Fosfatidilcolinas/análise , Fosfatidiletanolaminas/análise , Fosfatidilserinas/análise , Isótopos de Carbono , Células HeLa , Humanos , Limite de Detecção , Metilação , Fosfatidilcolinas/química , Fosfatidilcolinas/classificação , Fosfatidiletanolaminas/química , Fosfatidiletanolaminas/classificação , Fosfatidilserinas/química , Fosfatidilserinas/classificação , Sensibilidade e Especificidade , Espectrometria de Massas por Ionização por Electrospray/métodosRESUMO
The FA composition of glycerophospholipid (GPL) classes and subclasses was investigated in whole animals of three marine bivalve mollusks: the Japanese oyster Crassostrea gigas, the blue mussel Mytilus edulis, and the Manila clam Ruditapes philippinarum. Individual organs (gills, mantle, foot, siphon, and muscle) of the Manila clam also were examined. The PS plasmalogen (PSplsm), PE plasmalogen (PEplsm), and PC plasmalogen (PCplsm) subclasses were isolated by HPLC, and their individual FA compositions were examined using GC. Plasmalogen forms of PS and PE, when compared to their respective diacyl forms, were found to be specifically enriched with non-methylene-interrupted (NMI) FA (7,15-22:2, 7,13-22:2, and their precursors) and 20:1 n-11 FA. Such a clear specific association was not found for PCplsm. Interestingly, this trend was most apparent in PSplsm, and the above FA were found to be, respectively, the predominant PUFA and monounsaturated FA in the PSplsm isolated from the three species. This specificity was maintained in all the analyzed organs of the Manila clam but varied in proportions: The highest level of plasmalogens, NMI FA, and 20:1 n-11 was measured in gills and the lowest was in muscle. These results represent the first comprehensive report on a FA composition of the PSplsm subclass isolated from mollusks. The fact that NMI FA and 20:1 n-11, which are thought to be biosynthesized FA, were mainly associated with aminophospholipid plasmalogens (PE and PS) is likely to have a functional significance in bivalve membranes.
Assuntos
Bivalves/química , Ácidos Graxos/análise , Ostreidae/química , Plasmalogênios/química , Frutos do Mar/análise , Animais , Fosfatidilcolinas/química , Fosfatidilcolinas/classificação , Fosfatidiletanolaminas/química , Fosfatidiletanolaminas/classificação , Fosfatidilserinas/química , Fosfatidilserinas/classificação , Plasmalogênios/classificaçãoRESUMO
BACKGROUND: The aim of this study was to assess the short-term effect of IV infusion of fish oil emulsion on the fatty acid profiles of platelet phosphatidylcholine and phosphatidylethanolamine and on platelet function in postoperative patients. METHODS: Over a 7-day period, 10 patients received a 20% soybean fat emulsion with an added 10% marine fish oil emulsion, whereas 9 controls received only 20% soybean fat emulsion. RESULTS: By comparison with controls, in patients receiving fish oil, (1) a large increase in eicosapentaenoic acid (20:5n-3) was observed in both platelet phosphatidylcholine (1.55% +/- 0.17% vs 0.38% +/- 0.06% by weight, p < .01) and phosphatidylethanolamine 2.21% +/- 0.18% vs 0.66% +/- 0.08% by weight, p < .01); (2) eicosapentaenoic acid (20:5n-3)/arachidonic acid (20:4n-6) ratios doubled in both platelet phosphatidylcholine (p < .01) and phosphatidylethanolamine (p < .05); (3) with collagen as aggregating factor, maximal reaction speed decreased (p < .02) and latency increased (p < .002); and (4) no toxic effect, in particular no increase of postoperative bleeding and no perturbation of hepatic and renal function, was observed during the fish oil infusion. CONCLUSIONS: A short-term IV infusion of fish oil clearly modifies the platelet composition and changes some parameters of platelet function.
Assuntos
Plaquetas/química , Plaquetas/fisiologia , Emulsões Gordurosas Intravenosas/administração & dosagem , Óleos de Peixe/administração & dosagem , Nutrição Parenteral/métodos , Fosfolipídeos/análise , Estudos de Coortes , Humanos , Infusões Intravenosas , Pessoa de Meia-Idade , Fosfatidilcolinas/análise , Fosfatidilcolinas/química , Fosfatidilcolinas/classificação , Fosfatidiletanolaminas/análise , Fosfatidiletanolaminas/química , Fosfatidiletanolaminas/classificação , Fosfolipídeos/química , Fosfolipídeos/classificação , Período Pós-Operatório , Fatores de Tempo , Estados UnidosRESUMO
We show that phosphorus-31 nuclear magnetic resonance spectroscopy can be used to distinguish and to quantify the alk-1-enylacyl, alkylacyl, and diacyl glycerophosphoethanolamine (GPE) subclasses, and the respective glycerophosphocholine (GPC) subclasses, in their native form without prior degradation or derivatization, provided the phospholipids are observed in the nonaggregated state. Monomeric phospholipid distribution is ascertained by recording the spectra, after removal of metal ions, on CDCl3/CD3OD/D2O (50:50:15, by vol) solutions. The utility of this approach is exemplified for the ethanolamine glycerophospholipids (EPL) from bovine brain and the choline glycerophospholipids (CPL) from bovine heart. Sharp and well-resolved resonances are obtained for alkylacylGPE (+0.395 ppm; re 1% H3PO4), alkenylacylGPE (+0.353 ppm), and diacylGPE (+0.315 ppm), and for alkylacylGPC (-0.383 ppm), alkenyl-acylGPC (-0.436 ppm) and diacylGPC (-0.451 ppm). Integrated peak areas are shown to closely correlate with dose. Accurate quantitation of EPL and CPL subclasses at submicromolar levels can further be facilitated by use of synthetic dialkylGPE (+0.602 ppm) and dialkylGPC (-0.196 ppm) as internal standards. The method is simple, rapid, sensitive and reproducible, and permits the complete resolution and direct quantitation of all ethanolamine and choline glycerophospholipid subclasses quite independent of fatty chain length and degree of unsaturation.
Assuntos
Espectroscopia de Ressonância Magnética/métodos , Fosfatidilcolinas/análise , Fosfatidilcolinas/química , Fosfatidiletanolaminas/análise , Fosfatidiletanolaminas/química , Animais , Química Encefálica , Bovinos , Estrutura Molecular , Miocárdio/química , Fosfatidilcolinas/classificação , Fosfatidiletanolaminas/classificação , Fósforo/químicaRESUMO
A rapid method for the separation of the individual phospholipid classes phosphatidylcholine (PC), phosphatidylethanolamine (PE), phosphatidylserine (PS) and phosphatidylinositol (PI) by a single solid-phase extraction was developed. PC, PE, PS and PI were sequentially eluted from aminopropyl bonded silica with acetonitrile/n-propanol (2â¶1, vol/vol), methanol, isopropanol/methanolic HCl (4â¶1, vol/vol) and methanol/methanolic HCl (9â¶1, vol/vol). Standard recoveries were over 95% for PC and PE and over 85% for PS and PI with undistorted fatty acid composition. The separation of complex lipid mixtures on aminopropyl minicolumns can be refined to the level of individual phospholipid classes.
Assuntos
Fosfolipídeos/análise , Extração em Fase Sólida/métodos , Cromatografia Líquida de Alta Pressão , Cromatografia em Camada Fina , Fosfatidilcolinas/análise , Fosfatidilcolinas/classificação , Fosfatidiletanolaminas/análise , Fosfatidiletanolaminas/classificação , Fosfatidilinositóis/análise , Fosfatidilinositóis/classificação , Fosfatidilserinas/análise , Fosfatidilserinas/classificação , Fosfolipídeos/classificaçãoRESUMO
Four subfractions of phosphatidycholine and phosphyatidylethanolamine according to the degree of unsaturation of their fatty acids have been separated from lipid extracts of microsomes, and inner and outer mitochondrial membranes. The predominant species found in the three membranes contained one saturated and one unsaturated fatty acid. In microsomes completely saturated species of both phosphatidylcholine and phosphatideylethanolamine were practically nonexistent. In outer mitochondrial membranes species with two unsaturated fatty acids were absent. In the inner mitochondrial membranes, however, disaturated species and those with two unsaturated fatty acids were found.