RESUMEN
Pulmonary surfactant replacement therapy is a promising improvement in neonatal care for infants with respiratory distress syndrome. Lysophosphatidylcholine (LPC) is an undesirable component that can hinder surfactant proteins from enhancing the adsorption of surfactant lipids to balance surface tensions by creating a saturated coating on the interior of the lungs. A novel normal-phase liquid chromatography method utilizing UV detection and non-toxic solvents was developed and validated for the first time to analyze LPC in the complex matrix of pulmonary surfactant medication. The analytical method validation included evaluation of system suitability, repeatability, intermediate precision, linearity, accuracy, limit of detection (LOD), limit of quantification (LOQ), stability and robustness. The method yielded detection and quantification limits of 4.4 and 14.5 µg/ml, respectively. The calibration curve was modified linearly within the LOQ to 1.44 mg/ml range, with a determination coefficient of 0.9999 for standards and 0.9997 for sample solutions. Given the lack of reliable published data on LPC analysis in pulmonary surfactant medications, this newly developed method demonstrates promising results and offers advantages of HPLC methodology, including simplicity, accuracy, specificity, sensitivity and an exceptionally low LOD and LOQ. These attributes contribute to considering this achievement as an innovative method.
Asunto(s)
Límite de Detección , Lisofosfatidilcolinas , Surfactantes Pulmonares , Cromatografía Líquida de Alta Presión/métodos , Surfactantes Pulmonares/análisis , Surfactantes Pulmonares/química , Lisofosfatidilcolinas/análisis , Lisofosfatidilcolinas/química , Reproducibilidad de los Resultados , Animales , Bovinos , Modelos LinealesRESUMEN
Multiplexing of phosphatidylcholine analysis is hindered by a lack of appropriate derivatization. Presented here is a tagging scheme that uses a quaternary amine tag and targets the hydroxy group of the phosphate, which switches the net charge from neutral to +2. Quantitative yields were achieved from >99% reaction completion derived by dimethoxymethyl morpholinium (DMTMM) activation. Fragmentation of phosphatidylcholines (PCs) and lysophosphatidylcholines (LPCs) releases two trimethylamines and the acyl chains through neutral loss and generates a unique double cyclization constant mass reporter. Selective incorporation of isotopes onto the tag produces a six-plex set of isobaric reagents. For equivalent six-plex-labeled samples, <14% RSD was achieved, followed by a dynamic range of 1:10 without signal compression. Quantification of PCs/LPCs in human hepatic cancer cells was conducted as six-plex using data-dependent analysis tandem MS. We report a six-plex qualitative and quantitative isobaric tagging strategy expanding the limits of analyzing PCs/LPCs.
Asunto(s)
Fosfatidilcolinas , Espectrometría de Masas en Tándem , Humanos , Fosfatidilcolinas/química , Fosfatidilcolinas/análisis , Espectrometría de Masas en Tándem/métodos , Cromatografía Liquida/métodos , Ciclización , Línea Celular Tumoral , Células Hep G2 , Lisofosfatidilcolinas/análisis , Lisofosfatidilcolinas/químicaRESUMEN
Mfsd2a is the transporter for docosahexaenoic acid (DHA), an omega-3 fatty acid, across the blood brain barrier (BBB). Defects in Mfsd2a are linked to ailments from behavioral and motor dysfunctions to microcephaly. Mfsd2a transports long-chain unsaturated fatty-acids, including DHA and α-linolenic acid (ALA), that are attached to the zwitterionic lysophosphatidylcholine (LPC) headgroup. Even with the recently determined structures of Mfsd2a, the molecular details of how this transporter performs the energetically unfavorable task of translocating and flipping lysolipids across the lipid bilayer remains unclear. Here, we report five single-particle cryo-EM structures of Danio rerio Mfsd2a (drMfsd2a): in the inward-open conformation in the ligand-free state and displaying lipid-like densities modeled as ALA-LPC at four distinct positions. These Mfsd2a snapshots detail the flipping mechanism for lipid-LPC from outer to inner membrane leaflet and release for membrane integration on the cytoplasmic side. These results also map Mfsd2a mutants that disrupt lipid-LPC transport and are associated with disease.
Asunto(s)
Ácidos Grasos Omega-3 , Simportadores , Simportadores/metabolismo , Proteínas de Transporte de Membrana/metabolismo , Barrera Hematoencefálica/metabolismo , Transporte Biológico , Ácidos Docosahexaenoicos , Lisofosfatidilcolinas/químicaRESUMEN
Long-chain omega-3 fatty acids esterified in lysophosphatidylcholine (LPC-omega-3) are the most bioavailable omega-3 fatty acid form and are considered important for brain health. Lysophosphatidylcholine is a hydrolyzed phospholipid that is generated from the action of either phospholipase PLA1 or PLA2. There are two types of LPC; 1-LPC (where the omega-3 fatty acid at the sn-2 position is acylated) and 2-LPC (where the omega-3 fatty acid at the sn-1 position is acylated). The 2-LPC type is more highly bioavailable to the brain than the 1-LPC type. Given the biological and health aspects of LPC types, it is important to understand the structure, properties, extraction, quantification, functional role, and effect of the processing of LPC. This review examines various aspects involved in the extraction, characterization, and quantification of LPC. Further, the effects of processing methods on LPC and the potential biological roles of LPC in health and wellbeing are discussed. DHA-rich-LysoPLs, including LPC, can be enzymatically produced using lipases and phospholipases from wide microbial strains, and the highest yields were obtained by Lipozyme RM-IM®, Lipozyme TL-IM®, and Novozym 435®. Terrestrial-based phospholipids generally contain lower levels of long-chain omega-3 PUFAs, and therefore, they are considered less effective in providing the same health benefits as marine-based LPC. Processing (e.g., thermal, fermentation, and freezing) reduces the PL in fish. LPC containing omega-3 PUFA, mainly DHA (C22:6 omega-3) and eicosapentaenoic acid EPA (C20:5 omega-3) play important role in brain development and neuronal cell growth. Additionally, they have been implicated in supporting treatment programs for depression and Alzheimer's. These activities appear to be facilitated by the acute function of a major facilitator superfamily domain-containing protein 2 (Mfsd2a), expressed in BBB endothelium, as a chief transporter for LPC-DHA uptake to the brain. LPC-based delivery systems also provide the opportunity to improve the properties of some bioactive compounds during storage and absorption. Overall, LPCs have great potential for improving brain health, but their safety and potentially negative effects should also be taken into consideration.
Asunto(s)
Ácidos Grasos Omega-3 , Lisofosfatidilcolinas , Animales , Lisofosfatidilcolinas/química , Encéfalo/metabolismo , Ácidos Grasos Omega-3/metabolismo , Proteínas de Transporte de Membrana/metabolismo , Transporte Biológico , Ácido Eicosapentaenoico/metabolismo , Fosfolípidos/metabolismo , Ácidos Grasos/metabolismo , Ácidos Docosahexaenoicos/metabolismoRESUMEN
Lipid-porphyrin conjugates are versatile compounds which can self-assemble into liposome-like structures with multifunctional properties. Most of the conjugates that have been described so far, consisted in grafting pyropheophorbide-a (Pyro-a) or other porphyrin derivatives through the esterification of the hydroxyl group in the sn-2 position of a lysophosphatidylcholine. However, despite the versatility of these conjugates, less is known about the impact of the lipid backbone structure on their 2D phase behavior at the air/water interface and more precisely on their fine structures normal to the interface as well as on their in-plane organization. Herein, we synthesized a new lipid-porphyrin conjugate (PyroLSM) based on the amide coupling of Pyro-a to a lysosphingomyelin backbone (LSM) and we compared its interfacial behavior to that of Pyro-a and Pyro-a conjugated lysophosphatidylcholine (PyroLPC) using Langmuir balance combined to a variety of other physical techniques. Our results provided evidence on the significant impact of the lipid backbone on the lateral packing of the conjugates as well as on the shape and size of the formed domains. Compared to Pyro-a and PyroLPC monolayers, PyroLSM exhibited the highest lateral packing which highlights the role of the lipid backbone in controlling their 2D organization which in turn may impact the photophysical properties of their assemblies.
Asunto(s)
Lisofosfatidilcolinas , Porfirinas , Porfirinas/química , Lisofosfatidilcolinas/química , Agua , Aire , Estructura Molecular , Temperatura , Microscopía de Fuerza AtómicaRESUMEN
A huge amount of phospholipids or lecithin is produced as a byproduct in the vegetable oil industry. However, most are just used as a feed additive. This study has focused on enzymatic valorization of lecithin. This was exploited by enzymatic transformation of soy lecithin into lysolecithin liposomes, including functional free fatty acids, hydroxy fatty acids, hydrocarbons, or secondary fatty alcohols. One of the representative examples was the preparation of lysolecithin liposomes containing secondary fatty alcohols [e.g., 9-Hydroxyheptadec-11-ene (9) and 9-heptadecanol (10)] by using a phospholipase A2 from Streptomyces violaceoruber, a fatty acid double-bond hydratase from Stenotrophomonas maltophilia, and a photoactivated decarboxylase from Chlorella variabilis NC64A. The engineered liposomes turned out to range ca. 144 nm in diameter by dynamic light scattering analysis. Thereby, this study will contribute to application of functional fatty acids and their derivatives as well as valorization of lecithin for the food and cosmetic industries.
Asunto(s)
Carboxiliasas , Chlorella , Ácidos Grasos , Alcoholes Grasos , Lecitinas , Liposomas , Lisofosfatidilcolinas/química , Fosfatidilcolinas/química , Fosfolipasas A2RESUMEN
Lysophosphatidylcholine (LPC) is an essential mediator in human lipid metabolism and is associated with a variety of diseases, but the exact identity of LPC receptors remains controversial. Through extensive biochemical and structural analyses, we have identified the orphan receptor GPR119 as the receptor for LPC. The structure of the GPR119-G-protein complex without any added ligands reveals a density map that fits well with LPC, which is further confirmed by mass spectrometry and functional studies. As LPCs are abundant on the cell membrane, their preoccupancy in the receptor may lead to 'constitutive activity' of GPR119. The structure of GPR119 bound to APD668, a clinical drug candidate for type 2 diabetes, reveals an exceedingly similar binding mode to LPC. Together, these data highlight structural evidence for LPC function in regulating glucose-dependent insulin secretion through direct binding and activation of GPR119, and provide structural templates for drug design targeting GPR119.
Asunto(s)
Diabetes Mellitus Tipo 2 , Lisofosfatidilcolinas , Glucosa/metabolismo , Humanos , Ligandos , Lisofosfatidilcolinas/química , Lisofosfatidilcolinas/metabolismo , Receptores Acoplados a Proteínas G/químicaRESUMEN
Brain development and function require uptake of essential omega-3 fatty acids in the form of lysophosphatidylcholine via major-facilitator superfamily transporter MFSD2A, a potential pharmaceutical target to modulate blood-brain barrier (BBB) permeability. MFSD2A is also the receptor of endogenous retroviral envelope syncytin-2 (SYNC2) in human placenta, where it mediates cell-cell fusion and formation of the maternal-fetal interface. Here, we report a cryo-electron microscopy structure of the human MFSD2A-SYNC2 complex that reveals a large hydrophobic cavity in the transporter C-terminal domain to occlude long aliphatic chains. The transporter architecture suggests an alternating-access transport mechanism for lipid substrates in mammalian MFS transporters. SYNC2 establishes an extensive binding interface with MFSD2A, and a SYNC2-soluble fragment acts as a long-sought-after inhibitor of MFSD2A transport. Our work uncovers molecular mechanisms important to brain and placenta development and function, and SYNC2-mediated inhibition of MFSD2A transport suggests strategies to aid delivery of therapeutic macromolecules across the BBB.
Asunto(s)
Proteínas Gestacionales/química , Simportadores/química , Animales , Encéfalo/metabolismo , Microscopía por Crioelectrón , Femenino , Humanos , Lisofosfatidilcolinas/química , Lisofosfatidilcolinas/metabolismo , Mamíferos/metabolismo , Proteínas de Transporte de Membrana/metabolismo , Embarazo , Proteínas Gestacionales/metabolismo , Simportadores/metabolismoRESUMEN
Lysophosphatidylcholine (LPC) and phosphatidylcholine (PC) are important membrane constituents implicated in signaling and immune regulation. Synthesis of LPCs is challenging due to rapid acyl migration, e.g., induced by chromatography. We here report a highly regioselective synthesis of LPC and mixed PC via an intermediate allowing specific terminal acyl introduction, yielding the pure LPC without chromatography by an exceedingly mild TBS deprotection, using 1 equiv of TFA in aqueous solution. The method enabled the synthesis of glycerol-, acyl-, and choline-labeled LPC.
Asunto(s)
Lisofosfatidilcolinas , Fosfatidilcolinas , Lisofosfatidilcolinas/química , Lisofosfatidilcolinas/farmacología , Fosfatidilcolinas/química , AguaRESUMEN
As the major component of cell membranes, phosphatidylcholine (PC) is synthesized de novo in the Kennedy pathway and then undergoes extensive deacylation-reacylation remodeling via Lands' cycle. The re-acylation is catalyzed by lysophosphatidylcholine acyltransferase (LPCAT) and among the four LPCAT members in human, the LPCAT3 preferentially introduces polyunsaturated acyl onto the sn-2 position of lysophosphatidylcholine, thereby modulating the membrane fluidity and membrane protein functions therein. Combining the x-ray crystallography and the cryo-electron microscopy, we determined the structures of LPCAT3 in apo-, acyl donor-bound, and acyl receptor-bound states. A reaction chamber was revealed in the LPCAT3 structure where the lysophosphatidylcholine and arachidonoyl-CoA were positioned in two tunnels connected near to the catalytic center. A side pocket was found expanding the tunnel for the arachidonoyl CoA and holding the main body of arachidonoyl. The structural and functional analysis provides the basis for the re-acylation of lysophosphatidylcholine and the substrate preference during the reactions.
Asunto(s)
1-Acilglicerofosfocolina O-Aciltransferasa/química , Fosfolípidos/química , 1-Acilglicerofosfocolina O-Aciltransferasa/metabolismo , Acilcoenzima A/química , Acilcoenzima A/metabolismo , Acilación , Animales , Dominio Catalítico , Pollos , Microscopía por Crioelectrón , Cristalografía por Rayos X , Lisofosfatidilcolinas/química , Lisofosfatidilcolinas/metabolismo , Modelos Moleculares , Fosfolípidos/metabolismo , Multimerización de Proteína , Relación Estructura-Actividad , Especificidad por SustratoRESUMEN
The molecular details of the passive water flux across the hydrophobic membrane interior are still a matter of debate. One of the postulated mechanisms is the spontaneous, water-filled pore opening, which facilitates the hydrophilic connection between aqueous phases separated by the membrane. In the paper, we provide experimental evidence showing that the spontaneous lipid pore formation correlates with the membrane mechanics; hence, it depends on the composition of the lipid bilayer and the concentration of the osmotically active compound. Using liposomes as an experimental membrane model, osmotically induced water efflux was measured with the stopped-flow technique. Shapes of kinetic curves obtained at low osmotic pressure differences are interpreted in terms of two events: the lipid pore opening and water flow across the aqueous channel. The biological significance of the dependence of the lipid pore formation on the concentration difference of an osmotically active compound was illustrated by the demonstration that osmotically driven water flow can be accompanied by the dissipation of the pH gradient. The application of the Helfrich model to describe the probability of lipid pore opening was validated by demonstrating that the probability of pore opening correlates with the membrane bending rigidity. The correlation was determined by experimentally derived bending rigidity coefficients and probabilities of lipid pores opening.
Asunto(s)
Membrana Celular/química , Membrana Dobles de Lípidos/química , Membrana Dobles de Lípidos/metabolismo , Agua/metabolismo , Huevos , Concentración de Iones de Hidrógeno , Cinética , Luz , Liposomas/química , Lisofosfatidilcolinas/química , Lípidos de la Membrana/química , Modelos Químicos , Presión Osmótica , Fosfatidilcolinas/química , Fosfatidiletanolaminas/química , Protones , Agua/químicaRESUMEN
Insulin plays a significant role in carbohydrate homeostasis as the blood glucose lowering hormone. Glucose-induced insulin secretion (GSIS) is augmented by glucagon-like peptide (GLP-1), a gastrointestinal peptide released in response to ingesting nutriments. The secretion of insulin and GLP-1 is mediated by the binding of nutrients to G protein-coupled receptors (GPCRs) expressed by pancreatic ß-cells and enteroendocrine cells, respectively. Therefore, insulin secretagogues and incretin mimetics currently serve as antidiabetic treatments. This study demonstrates the potency of synthetic isoprenoid derivatives of lysophosphatidylcholines (LPCs) to stimulate GSIS and GLP-1 release. Murine insulinoma cell line (MIN6) and enteroendocrinal L cells (GLUTag) were incubated with LPCs bearing geranic acid (1-GA-LPC), citronellic acid (1-CA-LPC), 3,7-dimethyl-3-vinyloct-6-enoic acid (GERA-LPC), and (E)-3,7,11-trimethyl- 3-vinyldodeca-6,10-dienoic acid (1-FARA-LPC). Respective free terpene acids were also tested for comparison. Besides their insulin- and GLP-1-secreting capabilities, we also investigated the cytotoxicity of tested compounds, the ability to intracellular calcium ion mobilization, and targeted GPCRs involved in maintaining lipid and carbohydrate homeostasis. We observed the high cytotoxicity of 1-GERA-LPC and 1-FARA-LPC in contrast 1-CA-LPC and 1-GA-LPC. Moreover, 1-CA-LPC and 1-GA-LPC demonstrated the stimulatory effect on GSIS and 1-CA-LPC augmented GLP-1 secretion. Insulin and GLP-1 release appeared to be GPR40-, GPR55-, GPR119- and GPR120-dependent.
Asunto(s)
Péptido 1 Similar al Glucagón/biosíntesis , Secreción de Insulina/efectos de los fármacos , Insulina/biosíntesis , Metabolismo de los Lípidos/efectos de los fármacos , Lisofosfatidilcolinas/farmacología , Receptores Acoplados a Proteínas G/metabolismo , Terpenos/farmacología , Calcio/metabolismo , Humanos , Espacio Intracelular/metabolismo , Lisofosfatidilcolinas/química , Estructura Molecular , Terpenos/químicaRESUMEN
The lipid mediators, platelet-activating factor (PAF) and lysophosphatidylcholine (LPC), play relevant pathophysiological roles in Trypanosoma cruzi infection. Several species of LPC, including C18:1 LPC, which mimics the effects of PAF, are synthesized by T. cruzi. The present study identified a receptor in T. cruzi, which was predicted to bind to PAF, and found it to be homologous to members of the progestin and adiponectin family of receptors (PAQRs). We constructed a three-dimensional model of the T. cruzi PAQR (TcPAQR) and performed molecular docking to predict the interactions of the TcPAQR model with C16:0 PAF and C18:1 LPC. We knocked out T. cruzi PAQR (TcPAQR) gene and confirmed the identity of the expressed protein through immunoblotting and immunofluorescence assays using an anti-human PAQR antibody. Wild-type and knockout (KO) parasites were also used to investigate the in vitro cell differentiation and interactions with peritoneal mouse macrophages; TcPAQR KO parasites were unable to react to C16:0 PAF or C18:1 LPC. Our data are highly suggestive that PAF and LPC act through TcPAQR in T. cruzi, triggering its cellular differentiation and ability to infect macrophages.
Asunto(s)
Lisofosfatidilcolinas/metabolismo , Factor de Activación Plaquetaria/metabolismo , Proteínas Protozoarias/genética , Proteínas Protozoarias/metabolismo , Trypanosoma cruzi/genética , Trypanosoma cruzi/metabolismo , Secuencia de Aminoácidos , Animales , Diferenciación Celular , Enfermedad de Chagas/parasitología , Técnicas de Inactivación de Genes/métodos , Interacciones Huésped-Parásitos , Humanos , Lisofosfatidilcolinas/química , Macrófagos , Ratones , Simulación del Acoplamiento Molecular , Filogenia , Factor de Activación Plaquetaria/química , Conformación Proteica , Proteínas Protozoarias/química , Receptores de Adiponectina/química , Receptores de Adiponectina/genética , Receptores de Adiponectina/metabolismo , Receptores de Progesterona/química , Receptores de Progesterona/genética , Receptores de Progesterona/metabolismo , Trypanosoma cruzi/químicaRESUMEN
Phosphatidylcholines (PCs) are the major structural components of the plasma membrane of mammalian cells, while lysophosphatidylcholines (LPCs) are critical intermediates in lipid remodeling. Conventional tandem mass spectrometric (MSn) methods via collision-induced dissociation (CID) are blind to intrachain modifications such as the location of the carbon-carbon double bond (CâC) and methyl branching point. In this study, we demonstrate that almost complete structural information can be inferred from a single MS2 CID spectrum of the bicarbonate anion adducts of PC or LPC ([M + HCO3]-), including the identity of the headgroup, composition of fatty acyl chains, their sn-positions, the location of CâC, and the point of methyl branching in fatty acyls. We have integrated this MS2 CID method onto liquid chromatography for the analysis LPCs in human plasma, revealing the existence of multiple sn-isomers, branched chain isomers, and CâC location isomers of LPC.
Asunto(s)
Lisofosfatidilcolinas/química , Fosfatidilcolinas/química , Espectrometría de Masas en Tándem/métodos , Cromatografía Liquida , Humanos , Isomerismo , Lisofosfatidilcolinas/sangreRESUMEN
Docosahexaenoic acid (DHA) is one of the most important fatty acids that plays a critical role in maintaining proper brain function and cognitive development. Deficiency of DHA leads to several neurodegenerative disorders and, therefore, dietary supplementations of these fatty acids are essential to maintain cognitive health. However, the complete picture of how DHA is incorporated into the brain is yet to be explored. In general, the de novo synthesis of DHA is poor, and targeting the brain with specific phospholipid carriers provides novel insights into the process of reduction of disease progression. Recent studies have suggested that compared to triacylglycerol form of DHA, esterified form of DHA (i.e., lysophosphatidylcholine [lysoPC]) is better incorporated into the brain. Free DHA is transported across the outer membrane leaflet of the blood-brain barrier via APOE4 receptors, whereas DHA-lysoPC is transported across the inner membrane leaflet of the blood-brain barrier via a specific protein called Mfsd2a. Dietary supplementation of this lysoPC specific form of DHA is a novel therapy and is used to decrease the risk of various neurodegenerative disorders. Currently, structured glycerides of DHA - novel nutraceutical agents - are being widely used for the prevention and treatment of various neurological diseases. However, it is important to fully understand their metabolic regulation and mechanism of transportation to the brain. This article comprehensively reviews various studies that have evaluated the bioavailability of DHA, mechanisms of DHA transport, and role of DHA in preventing neurodegenerative disorders, which provides better insight into the pathophysiology of these disorders and use of structured DHA in improving neurological health.
Asunto(s)
Suplementos Dietéticos , Ácidos Docosahexaenoicos/administración & dosificación , Ácidos Docosahexaenoicos/metabolismo , Lisofosfatidilcolinas/administración & dosificación , Lisofosfatidilcolinas/metabolismo , Enfermedades Neurodegenerativas/prevención & control , Animales , Disponibilidad Biológica , Transporte Biológico , Barrera Hematoencefálica/metabolismo , Encéfalo/metabolismo , Grasas Insaturadas en la Dieta/administración & dosificación , Ácidos Docosahexaenoicos/biosíntesis , Ácidos Docosahexaenoicos/química , Ácidos Grasos Insaturados/administración & dosificación , Humanos , Lisofosfatidilcolinas/química , Enfermedades Neurodegenerativas/fisiopatología , Obesidad/metabolismoRESUMEN
Studies on neutrophil-based nanotherapeutic engineering have shown great potentials in treating infection and inflammation disorders. Conventional neutrophil labeling methods are time-consuming and often result in undesired contamination and activation since neutrophils are terminal-differentiated cells with a half-life span of only 7â h. A simple, fast, and biocompatible strategy to construct engineered neutrophils is highly desirable but remains difficult to achieve. In this study, we present an AIEgen-lipid conjugate, which can efficiently label harvested neutrophils in 30â s with no washing step required. This fast labeling method does not affect the activation and transmigration property of neutrophils, which has been successfully used to monitor neutrophil behaviors such as the chemotaxis process and migrating function towards inflammation sites both inâ vitro and inâ vivo, offering a tantalizing prospect for neutrophil-based nanotherapeutics studies.
Asunto(s)
Lípidos/química , Neutrófilos/metabolismo , Animales , Quimiotaxis , Membrana Dobles de Lípidos/química , Lisofosfatidilcolinas/química , Ratones , Nanopartículas/química , Neutrófilos/química , Neutrófilos/inmunología , Imagen Óptica , Especies de Nitrógeno Reactivo/metabolismo , Especies Reactivas de Oxígeno/metabolismoRESUMEN
Alimentary atherosclerosis is associated with a significant decrease in the content of phosphatidylcholine, the phospholipid that provides antioxidant protection, in the plasma membrane of liver cells, while the level of phosphatidic acid that initiates generation of superoxides, on the contrary, increases. The level of membrane phosphatidylserine, a target of the scavenger receptors, which initiates removal of damaged cells and modified lipoproteins from the circulation was also elevated. In the blood serum of rabbits receiving an atherogenic diet, the content of cardiolipin involved in the immune mechanisms of atherosclerosis development and a risk factor for thrombosis, sharply increased. The level of lysophosphatidylcholine that mediates initiation and progression of atherosclerosis increased. The content of phosphatidylinositol that is involved in the mechanisms protecting from exposure to excess cholesterol was significantly reduced. Treatment of alimentary atherosclerosis with "empty" phosphatidylcholine nanosomes eliminates the key factors initiating atherosclerosis development.
Asunto(s)
Aterosclerosis/sangre , Membrana Celular/metabolismo , Hígado/metabolismo , Nanopartículas/química , Fosfatidilcolinas/química , Fosfolípidos/química , Animales , Antioxidantes/química , Cardiolipinas/metabolismo , Dieta Aterogénica , Sistema Inmunológico , Lipoproteínas/metabolismo , Lisofosfatidilcolinas/química , Conejos , Factores de Riesgo , Trombosis/inmunologíaRESUMEN
Nonshivering thermogenesis occurs in brown adipose tissue to generate heat in response to cold ambient temperatures. Thioesterase superfamily member 1 (Them1) is transcriptionally up-regulated in brown adipose tissue upon exposure to the cold and suppresses thermogenesis in order to conserve energy reserves. It hydrolyzes long-chain fatty acyl-CoAs that are derived from lipid droplets, preventing their use as fuel for thermogenesis. In addition to its enzymatic domains, Them1 contains a C-terminal StAR-related lipid transfer (START) domain with unknown ligand or function. By complementary biophysical approaches, we show that the START domain binds to long-chain fatty acids, products of Them1's enzymatic reaction, as well as lysophosphatidylcholine (LPC), lipids shown to activate thermogenesis in brown adipocytes. Certain fatty acids stabilize the START domain and allosterically enhance Them1 catalysis of acyl-CoA, whereas 18:1 LPC destabilizes and inhibits activity, which we verify in cell culture. Additionally, we demonstrate that the START domain functions to localize Them1 near lipid droplets. These findings define the role of the START domain as a lipid sensor that allosterically regulates Them1 activity and spatially localizes it in proximity to the lipid droplet.
Asunto(s)
Ácidos Grasos/metabolismo , Lisofosfatidilcolinas/metabolismo , Palmitoil-CoA Hidrolasa/química , Palmitoil-CoA Hidrolasa/metabolismo , Acilcoenzima A/metabolismo , Tejido Adiposo Pardo/enzimología , Tejido Adiposo Pardo/metabolismo , Regulación Alostérica , Ácidos Grasos/química , Humanos , Cinética , Gotas Lipídicas/enzimología , Gotas Lipídicas/metabolismo , Lisofosfatidilcolinas/química , Palmitoil-CoA Hidrolasa/genética , Dominios ProteicosRESUMEN
Untargeted metabolomics can be a great tool for exploring new scientific areas; however, wrong metabolite annotation questions the credibility and puts the success of the entire research at risk. Therefore, an effort should be made to improve the quality and robustness of the annotation despite of the challenges, especially when final identification with standards is not possible. Through non-targeted analysis of human plasma samples, from a large cancer cohort study using RP-LC-ESI-QTOF-MS/MS, we have resolved MS/MS annotation through spectral matching, directed to hydroxyeicosatetraenoic acids (HETEs) and, MS/MS structural elucidation for newly annotated oxidized lyso-phosphatidylcholines (oxLPCs). The annotation of unknowns is supported with structural information from fragmentation spectra as well as the fragmentation mechanisms involved, necessarily including data from both polarity modes and different collision energies. In this work, we present evidences that various oxidation products show significant differences between cancer patients and control individuals and we establish a workflow to help identify such modifications. We report here the upregulation of HETEs and oxLPCs in patients with neuroendocrine tumors (NETs). To our knowledge, this is the first attempt to determine HETEs in NETs and one of very few studies where oxLPCs are annotated. The obtained results provide an important insight regarding lipid oxidation in NETs, although their physiological functions still have to be established and require further research.
Asunto(s)
Lípidos/sangre , Metaboloma , Adulto , Anciano , Anciano de 80 o más Años , Axitinib/uso terapéutico , Cromatografía Líquida de Alta Presión , Cromatografía de Fase Inversa , Femenino , Humanos , Peroxidación de Lípido , Lípidos/química , Lípidos/aislamiento & purificación , Lisofosfatidilcolinas/sangre , Lisofosfatidilcolinas/química , Lisofosfatidilcolinas/aislamiento & purificación , Masculino , Persona de Mediana Edad , Tumores Neuroendocrinos/tratamiento farmacológico , Tumores Neuroendocrinos/metabolismo , Tumores Neuroendocrinos/patología , Análisis de Componente Principal , Espectrometría de Masas en Tándem/métodosRESUMEN
Recently, the parylene-matrix chip was developed for quantitative analysis of small molecules less than 1 kDa. In this study, MALDI-TOF MS based on the parylene-matrix chip was performed to clinically diagnose intrahepatic cholangiocarcinoma (IHCC) and colorectal cancer (CRC). The parylene-matrix chip was applied for the detection of small cancer biomarkers, including N-methyl-2-pyridone-5-carboxamide (2PY), glutamine, lysophosphatidylcholine (LPC) 16:0, and LPC 18:0. The feasibility of MALDI-TOF MS based on the parylene-matrix chip was confirmed via analysis of spot-to-spot and shot-to-shot reproducibility. Serum metabolite markers of IHCC, N-methyl-2-pyridone-5-carboxamide (2PY), and glutamine were quantified using MALDI-TOF MS based on the parylene-matrix chip. For clinical diagnosis of CRC, two water-insoluble (barely soluble) biomarkers, lysophosphatidylcholine (LPC) 16:0 and LPC 18:0, were quantified. Finally, glutamine and LPC 16:0 were simultaneously detected at a range of concentrations in sera from colon cancer patients using the parylene-matrix chip. Thus, this method yielded high-throughput detection of cancer biomarkers for the mixture samples of water-soluble analytes (2PY and glutamine) and water-insoluble analytes (LPC 16:0 and LPC 18:0).