RESUMEN
Modulation of the plant defense response by bioactive molecules is of increasing interest. However, despite plant cell lipids being one of the major cellular components, their role in plant immunity remains elusive. We found that the exogenous application of the cell-membrane localized phospholipid lyso-phosphatidylethanolamine (LPE) reprograms the plant transcript profile in favor of defense-associated genes thereby priming the plant immune system. Exogenous LPE application to different Arabidopsis accessions increases resistance against the necrotrophic pathogens, Botrytis cinerea and Cochliobolus heterostrophus. We found that the immunity-promoting effect of LPE is repealed in the jasmonic acid (JA) receptor mutant coi1, but multiplied in the JA-hypersensitive mutant feronia (fer-4). The JA-signaling repressor JAZ1 is degraded following LPE administration, suggesting that JA-signaling is promoted by LPE. Following LPE-treatment, reactive oxygen species (ROS) accumulation is affected in coi1 and fer-4. Moreover, FER signaling inhibitors of the RALF family are strongly expressed after LPE application, and RALF23 is internalized in stress granules, suggesting the LPE-mediated repression of FER-signaling by promoting RALF function. The in-situ increase of LPE-abundance in the LPE-catabolic mutants lpeat1 and lpeat2 elevates plant resistance to B. cinerea, in contrast to the endogenous LPE-deficient mutant pla2-alpha. We show that LPE increases plant resistance against necrotrophs by promoting JA-signaling and ROS-homeostasis, thereby paving the way for the LPE-targeted genomic engineering of crops to raise their ability to resist biotic threats.
Asunto(s)
Proteínas de Arabidopsis , Arabidopsis , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Fosfatidiletanolaminas/metabolismo , Fosfatidiletanolaminas/farmacología , Arabidopsis/metabolismo , Oxilipinas/metabolismo , Ciclopentanos/metabolismo , Homeostasis , Enfermedades de las Plantas/genética , Botrytis/metabolismo , Regulación de la Expresión Génica de las PlantasRESUMEN
Oxidized low-density lipoprotein (OX-LDL)-induced inflammation and autophagy dysregulation are important events in the progression of atherosclerosis. Phosphatidylethanolamine (PE), a multifunctional phospholipid that is enriched in cells, has been proven to be directly involved in autophagy which is closely associated with inflammation. However, whether PE can influence OX-LDL-induced autophagy dysregulation and inflammation has not been reported. In the present study, we revealed that OX-LDL significantly induced macrophage inflammation through the CD36-NLRP1-caspase-1 signaling pathway in fish. Meanwhile, cellular PE levels were significantly decreased in response to OX-LDL induction. Based on the relationship between PE and autophagy, we then examined the effect of PE supplementation on OX-LDL-mediated autophagy impairment and inflammation induction in macrophages. As expected, exogenous PE restored impaired autophagy and alleviated inflammation in OX-LDL-stimulated cells. Notably, autophagy inhibitors reversed the inhibitory effect of PE on OX-LDL-induced maturation of IL-1ß, indicating that the regulation of PE on OX-LDL-induced inflammation is dependent on autophagy. Furthermore, the positive effect of PE on OX-LDL-induced inflammation was relatively conserved in mouse and fish macrophages. In conclusion, we elucidated the role of the CD36-NLRP1-caspase-1 signaling pathway in OX-LDL-induced inflammation in fish and revealed for the first time that altering PE abundance in OX-LDL-treated cells could alleviate inflammasome-mediated inflammation by inducing autophagy. Given the relationship between OX-LDL-induced inflammation and atherosclerosis, this study prompts that the use of PE-rich foods promises to be a new strategy for atherosclerosis treatment in vertebrates.
Asunto(s)
Aterosclerosis , Inflamasomas , Fosfatidiletanolaminas , Animales , Ratones , Aterosclerosis/tratamiento farmacológico , Aterosclerosis/genética , Aterosclerosis/metabolismo , Autofagia , Caspasa 1/genética , Caspasa 1/metabolismo , Inflamasomas/genética , Inflamasomas/metabolismo , Inflamación/tratamiento farmacológico , Inflamación/metabolismo , Lipoproteínas LDL/metabolismo , Macrófagos/metabolismo , Fosfatidiletanolaminas/farmacologíaRESUMEN
Modulation of bile acid (BA) structure is a potential strategy for obesity and metabolic disease treatment. BAs act not only as signaling molecules involved in energy expenditure and glucose homeostasis, but also as regulators of food intake. The structure of BAs, particularly the position of the hydroxyl groups of BAs, impacts food intake partly by intestinal effects: (1) modulating the activity of N-acyl phosphatidylethanolamine phospholipase D, which produces the anorexigenic bioactive lipid oleoylethanolamide (OEA) or (2) regulating lipid absorption and the gastric emptying-satiation pathway. We hypothesized that 16α-hydroxylated BAs uniquely regulate food intake because of the long intermeal intervals in snake species in which these BAs are abundant. However, the effects of 16α-hydroxylated BAs in mammals are completely unknown because they are not naturally found in mammals. To test the effect of 16α-hydroxylated BAs on food intake, we isolated the 16α-hydroxylated BA pythocholic acid from ball pythons (Python regius). Pythocholic acid or deoxycholic acid (DCA) was given by oral gavage in mice. DCA is known to increase N-acyl phosphatidylethanolamine phospholipase D activity better than other mammalian BAs. We evaluated food intake, OEA levels, and gastric emptying in mice. We successfully isolated pythocholic acid from ball pythons for experimental use. Pythocholic acid treatment significantly decreased food intake in comparison to DCA treatment, and this was associated with increased jejunal OEA, but resulted in no change in gastric emptying or lipid absorption. The exogenous BA pythocholic acid is a novel regulator of food intake and the satiety signal for OEA in the mouse intestine.
Asunto(s)
Ácidos y Sales Biliares , Fosfolipasa D , Ratones , Masculino , Animales , Fosfolipasa D/metabolismo , Fosfolipasa D/farmacología , Fosfatidiletanolaminas/farmacología , Ingestión de Alimentos , Mamíferos/metabolismoRESUMEN
Decarboxylation of phosphatidylserine (PS) to form phosphatidylethanolamine by PS decarboxylases (PSDs) is an essential process in most eukaryotes. Processing of a malarial PSD proenzyme into its active alpha and beta subunits is by an autoendoproteolytic mechanism regulated by anionic phospholipids, with PS serving as an activator and phosphatidylglycerol (PG), phosphatidylinositol, and phosphatidic acid acting as inhibitors. The biophysical mechanism underlying this regulation remains unknown. We used solid phase lipid binding, liposome-binding assays, and surface plasmon resonance to examine the binding specificity of a processing-deficient Plasmodium PSD (PkPSDS308A) mutant enzyme and demonstrated that the PSD proenzyme binds strongly to PS and PG but not to phosphatidylethanolamine and phosphatidylcholine. The equilibrium dissociation constants (Kd) of PkPSD with PS and PG were 80.4 nM and 66.4 nM, respectively. The interaction of PSD with PS is inhibited by calcium, suggesting that the binding mechanism involves ionic interactions. In vitro processing of WT PkPSD proenzyme was also inhibited by calcium, consistent with the conclusion that PS binding to PkPSD through ionic interactions is required for the proenzyme processing. Peptide mapping identified polybasic amino acid motifs in the proenzyme responsible for binding to PS. Altogether, the data demonstrate that malarial PSD maturation is regulated through a strong physical association between PkPSD proenzyme and anionic lipids. Inhibition of the specific interaction between the proenzyme and the lipids can provide a novel mechanism to disrupt PSD enzyme activity, which has been suggested as a target for antimicrobials, and anticancer therapies.
Asunto(s)
Carboxiliasas , Malaria , Fosfolípidos , Plasmodium , Secuencias de Aminoácidos , Calcio/metabolismo , Calcio/farmacología , Carboxiliasas/antagonistas & inhibidores , Carboxiliasas/química , Carboxiliasas/metabolismo , Precursores Enzimáticos/metabolismo , Liposomas , Ácidos Fosfatidicos/metabolismo , Ácidos Fosfatidicos/farmacología , Fosfatidilcolinas/metabolismo , Fosfatidilcolinas/farmacología , Fosfatidiletanolaminas/metabolismo , Fosfatidiletanolaminas/farmacología , Fosfatidilgliceroles/metabolismo , Fosfatidilgliceroles/farmacología , Fosfatidilinositoles/metabolismo , Fosfatidilinositoles/farmacología , Fosfatidilserinas/metabolismo , Fosfatidilserinas/farmacología , Fosfolípidos/química , Fosfolípidos/metabolismo , Fosfolípidos/farmacología , Unión Proteica , Malaria/parasitología , Proteolisis/efectos de los fármacos , Resonancia por Plasmón de Superficie , Plasmodium/enzimologíaRESUMEN
Eight lactating cows were used to determine the effects of citrus peel extract (CPE) on milk performance, antioxidant properties, and milk lipids composition. CPE supplementation up to 150 g/d (CPE150) increased milk yield and the proportions of unsaturated fatty acids of conjugated linoleic acid. CPE with abundant polyphenol and flavonoids can transfer these bioactive substances to mammary gland and improve the antioxidant properties of milk obtained from cows. Lipidomics revealed that 56 lipid species were altered between CON vs CPE150, and there were five key differential metabolic pathways. In particular, milk phosphatidylethanolamine and phosphatidylcholine were significantly increased with dietary CPE supplementation. In summary, our results provide insights into the modifications in the milk components and milk quality of dairy cows received CPE. The inclusion of CPE in the diet of dairy cows may be an effective and natural way to increase the antioxidant amounts and beneficial lipids in milk.
Asunto(s)
Citrus , Ácidos Linoleicos Conjugados , Alimentación Animal/análisis , Animales , Antioxidantes/farmacología , Bovinos , Cromatografía Liquida , Suplementos Dietéticos , Femenino , Lactancia , Ácidos Linoleicos Conjugados/metabolismo , Lipidómica , Leche/metabolismo , Fosfatidilcolinas , Fosfatidiletanolaminas/metabolismo , Fosfatidiletanolaminas/farmacología , Extractos Vegetales/metabolismo , Extractos Vegetales/farmacología , Polifenoles/farmacología , Espectrometría de Masas en TándemRESUMEN
Serious environmental pollution of heavy metals has attracted people's attention in recent years and halophiles seem to be potential bioremediation in the controlling of heavy metals contamination. In this study, the adaptive mechanism of halophilic Brachybacterium muris (B. muris) in response to salt stress and its mitigation of copper (Cu) toxicity in hydroponic plants were investigated. The cell morphology was observed using transmission electron microscopy. The cell membrane composition and fluidity were examined by the combination of gas chromatography, gas chromatography-mass spectrometry, ultra-high performance liquid chromatography-mass spectrometry, and fluorescence spectrophotometry. Moreover, the metabolic pathways of B. muris in response to salt stress were analyzed using the prokaryotic transcriptomics approach. A hydroponic co-culture model was further conducted to explore the effects of B. muris on wheat seedlings subjected to Cu toxicity. It was found that B. muris can respond to high osmotic pressure by improving the cell membrane fluidity, altering the cell morphology and cell membrane compositions. The proportion of unsaturated fatty acids, phosphatidylethanolamine, and phosphatidylinositol in B. muris cell membranes increased significantly, while zymosterol, fecosterol, and ergosterol contents decreased under a high salinity situation. Further transcriptomic analysis showed that genes encoding L-glutamate synthase, glutamate ABC transporter ATP-binding protein, and sodium cotransporter were up-regulated, indicating that both the synthesis and transport of glutamate were significantly enhanced under high osmotic pressure. Additionally, B. muris alleviated the inhibitory effect of Cu2+ on wheat seedlings' growth, causing a 30.14% decrease in H2O2 content and a significant increase of 83.86% and 45.96% in POD activity and GSH content in wheat roots, respectively. The findings of this study suggested that the salt-tolerant B. muris may serve as a promising strategy for improving the bioremediation of metal-contaminated saline water and soils.
Asunto(s)
Cobre , Metales Pesados , Transportadoras de Casetes de Unión a ATP/metabolismo , Actinobacteria , Adenosina Trifosfato/metabolismo , Cobre/toxicidad , Ergosterol/metabolismo , Ergosterol/farmacología , Cromatografía de Gases y Espectrometría de Masas , Glutamato Sintasa/metabolismo , Glutamato Sintasa/farmacología , Ácido Glutámico/metabolismo , Ácido Glutámico/farmacología , Humanos , Peróxido de Hidrógeno/metabolismo , Hidroponía , Metales Pesados/toxicidad , Fosfatidiletanolaminas/metabolismo , Fosfatidiletanolaminas/farmacología , Fosfatidilinositoles/metabolismo , Fosfatidilinositoles/farmacología , Raíces de Plantas/metabolismo , Estrés Salino , Plantones , Sodio/metabolismo , Suelo , Triticum/metabolismoRESUMEN
Microplastic exposure leads to various toxic effects in Daphnia magna; however, the effects of microplastics on the metabolic processes in D. magna and the corresponding molecular toxicity mechanisms remain unclear. In the present study, the effects of acute exposure to polyethylene microplastics with different particle sizes (20 µm [MPs-20] and 30 µm [MPs-30]) on metabolites in D. magna and the mechanisms of toxicity were investigated by combining metabolomics and traditional toxicology techniques. Exposure to both MPs-20 and MPs-30 resulted in significant accumulation of microplastics in the gut of D. magna and significantly reduced D. magna survival and heart rate. Metabolomics analysis revealed that MPs-20 and MPs-30 induced significant changes in up to 88 and 91 differential metabolites, respectively, and collectively induced significant changes in 75 metabolites in D. magna. Among lipid metabolites, MPs-20 specifically downregulated phosphatidylcholine and upregulated phosphatidylethanolamine, which mainly affected phospholipid metabolism, whereas MPs-30 specifically downregulated amino acid metabolites l-glutamine, l-glutamate and malic acid, which mainly interfered with energy metabolism. The results of this study provide novel insights into the mechanism of effects of microplastics on metabolic processes in D. magna.
Asunto(s)
Microplásticos , Contaminantes Químicos del Agua , Animales , Daphnia , Ácido Glutámico , Glutamina/farmacología , Fosfatidilcolinas/farmacología , Fosfatidiletanolaminas/farmacología , Plásticos/toxicidad , Polietileno/toxicidad , Contaminantes Químicos del Agua/análisisRESUMEN
Health risks caused by crucial environmental carcinogens N-nitrosamines triggered ubiquitous attention. As the liver exerted vital function through metabolic process, lipid metabolism disorders have been confirmed as potential drivers for toxicological effects, and the mechanisms of lipid regulation related to hepatotoxicity induced by N-nitrosamines remained largely unclear. In this study, we comprehensively explored the disturbance of hepatic lipid homeostasis in mice induced by nitrosamines. The results implied that nitrosamines exposure induced hepatotoxicity accompanied by liver injury, inflammatory infiltration, and hepatic edema. Lipidomics profiling analysis indicated the decreased levels of phosphatidic acids (PA), phosphatidylcholines (PC), phosphatidylethanolamines (PE), lyso-phosphatidylcholines (LPC), lyso-phosphatidylethanolamines (LPE), diacylglycerols (DAG) and triacylglycerols (TAG), the elevation of ceramides (Cer) and decomposition of free fatty acids (FFA) in high-dose nitrosamines exposure group. Importantly, nitrosamines exposure promoted fatty acid oxidation (FAO) by facilitating fatty acid uptake and decomposition, together with the upregulation of genes associated with FAO accompanied by the activation of inflammatory cytokines TNF-α, IL-1ß and NLRP3. Furthermore, fatty acid translocase CD36-mediated fatty acid oxidation was correlated with the enhancement of oxidative stress in the liver caused by nitrosamines exposure. Overall, our results contributed to the new strategies to interpret the early toxic effects of nitrosamines exposure.
Asunto(s)
Enfermedad Hepática Inducida por Sustancias y Drogas , Trastornos del Metabolismo de los Lípidos , Nitrosaminas , Animales , Enfermedad Hepática Inducida por Sustancias y Drogas/etiología , Enfermedad Hepática Inducida por Sustancias y Drogas/metabolismo , Ácidos Grasos/metabolismo , Metabolismo de los Lípidos , Trastornos del Metabolismo de los Lípidos/metabolismo , Hígado , Ratones , Ratones Endogámicos ICR , Nitrosaminas/toxicidad , Fosfatidilcolinas/metabolismo , Fosfatidilcolinas/farmacología , Fosfatidiletanolaminas/metabolismo , Fosfatidiletanolaminas/farmacologíaRESUMEN
Brain ageing has been related to a decrease in cellular metabolism, to an accumulation of misfolded proteins and to an alteration of the lipid membrane composition. These alterations act as contributive aspects of age-related memory decline by reducing membrane excitability and neurotransmitter release. In this sense, precursors of phospholipids (PLs) can restore the physiological composition of cellular membranes and ameliorate the cellular defects associated with brain ageing. In particular, phosphatidylcholine (PC) and phosphatidylethanolamine (PE) have been shown to restore mitochondrial function, reduce the accumulation of amyloid beta (Aß) and, at the same time, provide the amount of acetylcholine needed to reduce memory deficit. Among PL precursors, alpha-glycerylphosphorylethanolamine (GPE) has shown to protect astrocytes from Aß injuries and to slow-down ageing of human neural stem cells. GPE has been evaluated in aged human hippocampal neurons, which are implicated in learning and memory, and constitute a good in vitro model to investigate the beneficial properties of GPE. In order to mimic cellular ageing, the cells have been maintained 21 days in vitro and challenged with GPE. Results of the present paper showed GPE ability to increase PE and PC content, glucose uptake and the activity of the chain respiratory complex I and of the GSK-3ß pathway. Moreover, the nootropic compound showed an increase in the transcriptional/protein levels of neurotrophic and well-being related genes. Finally, GPE counteracted the accumulation of ageing-related misfolded proteins (a-synuclein and tau). Overall, our data underline promising effects of GPE in counteracting cellular alterations related to brain ageing and cognitive decline.
Asunto(s)
Péptidos beta-Amiloides , Fosfatidiletanolaminas , Anciano , Péptidos beta-Amiloides/metabolismo , Etanolaminas/metabolismo , Etanolaminas/farmacología , Glucógeno Sintasa Quinasa 3 beta/metabolismo , Hipocampo/metabolismo , Humanos , Neuronas/metabolismo , Fosfatidilcolinas/metabolismo , Fosfatidiletanolaminas/metabolismo , Fosfatidiletanolaminas/farmacologíaRESUMEN
Antitumor activities of L-MTP-PE (Liposome entrapped myuramyl tripeptide phosphatidylethanolamine) in the combination treatment with chemo- or immune-therapeutic antitumor agents against various syngeneic tumors were tested.Against Meth A fibrosarcoma solid tumor system, L-MTP-PE showed slight but statistically significant elongation of survival days against 5-FU monotherapy in spite of its non-effect on tumor growth, when combined with 5-FU. Against liver metastasis model of M5076 carcinoma, L-MTP-PE showed a tendency of elongation of survival days by its single drug treatment, however, elongation with statistical significance was observed in the combination treatment with 5-FU in comparison with control group.These data suggest that L-MTP-PE seems to elongate the survival days of the solid tumor bearing mice and the liver metastasis model basically due to its saving effect on chemotherapeutic drug-induced immunosuppression. In the combination with an immunotherapeutic agent in mice, TNF production induced by another biological response modifier OK-432 was potentiated when primed with L-MTP-PE. L-MTP-PE also potentiate the antitumor effect of OK-432 possibly through the enhanced production of TNF-α. Combination of L-MTP-PE and OK-432 is considered to be a candidate for a new treatment model for cancer.
Asunto(s)
Neoplasias Hepáticas , Fosfatidiletanolaminas , Acetilmuramil-Alanil-Isoglutamina/análogos & derivados , Acetilmuramil-Alanil-Isoglutamina/farmacología , Acetilmuramil-Alanil-Isoglutamina/uso terapéutico , Adyuvantes Inmunológicos , Animales , Portadores de Fármacos , Fluorouracilo , Factores Inmunológicos , Agentes Inmunomoduladores , Liposomas , Neoplasias Hepáticas/tratamiento farmacológico , Ratones , Fosfatidiletanolaminas/farmacología , Fosfatidiletanolaminas/uso terapéutico , PicibanilRESUMEN
The aim of this study was to develop a green method to fabricate a novel CS modified N-(4-hydroxyphenyl)- methacrylamide conjugate (CSNHMA) and to evaluate its biomedical potential. CSNHMA has been prepared by a simple method via aza Michael addition reaction between CS and N- (4-hydroxyphenyl)-methacrylamide (NHMA) in ethanol. Its structural and morphological properties were characterized by various analysis techniques. The obtained results confirmed that a highly porous network structure of CSNHMA was successfully synthesized via aza Michael addition reaction. Consequently, it was analyzed as a drug and gene carrier. CSNHMA/pGL3 showed an enhanced buffering capacity due to the presence of NHMA moiety leading to higher transfection efficiency in all cancer cells (A549, HeLa and HepG2) as compared to native CS and Lipofectamine®. Therefore, these findings clearly support the possibility of using CSNHMA as a good transfection agent. For in vitro drug release study, we prepared CSNHMA nanoparticles (NPs) and curcumin loaded CSNHMA NPs of size <230 nm respectively via the non-toxic ionic gelation route and the encapsulation efficiency of drug was found to be 77.03%. In vitro drug release studies demonstrated a faster and sustained release of curcumin loaded CSNHMA NPs at pH 5.0 compared to physiological pH.
Asunto(s)
Acrilamidas/química , Quitosano/síntesis química , Curcumina/farmacología , Luciferasas/genética , Células A549 , Secuencia de Carbohidratos , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Quitosano/química , Curcumina/química , Preparaciones de Acción Retardada , Portadores de Fármacos , Tecnología Química Verde , Células HeLa , Células Hep G2 , Humanos , Tamaño de la Partícula , Fosfatidiletanolaminas/farmacología , Porosidad , TransfecciónRESUMEN
Mesenchymal stem cell-derived small extracellular vesicles (MSC-sEVs) possess a great therapeutical potential for osteoarthritis (OA) treatment. However, the steric and electrostatic hindrance of cartilage matrix leads to very limited distribution of MSC-sEVs in cartilage and low bioavailability of MSC-sEVs after intra-articular injection. To overcome this, a strategy to reverse the surface charge of MSC-sEVs by modifying the MSC-sEVs with a novel cationic amphiphilic macromolecule namely ε-polylysine-polyethylene-distearyl phosphatidylethanolamine (PPD) was developed in this study. Through incubation with 100 µg/ml PPD, positively charged MSC-sEVs (PPD-sEVs) were obtained, and the modification process showed nearly no disturbance to the integrity and contents of sEVs and exhibited good stability under the interference of anionic macromolecules. A more effective cellular uptake and homeostasis modulation ability of PPD-sEVs than unmodified MSC-sEVs to chondrocytes was demonstrated. More importantly, PPD-sEVs demonstrated significantly enhanced cartilage uptake, cartilage penetration, and joint retention capacity as compared to MSC-sEVs. Intra-articular injection of PPD-sEVs into a mouse OA model showed significantly improved bioavailability than MSC-sEVs, which resulted in enhanced therapeutic efficacy with reduced injection frequency. In general, this study provides a facile and effective strategy to improve the intra-articular bioavailability of MSC-sEVs and has a great potential to accelerate the clinical practice of MSC-sEVs based OA therapy.
Asunto(s)
Tratamiento Basado en Trasplante de Células y Tejidos/métodos , Vesículas Extracelulares/efectos de los fármacos , Células Madre Mesenquimatosas/citología , Osteoartritis/terapia , Fosfatidiletanolaminas/química , Fosfatidiletanolaminas/farmacología , Polietilenglicoles/química , Polietilenglicoles/farmacología , Polilisina/química , Polilisina/farmacología , Adolescente , Animales , Cartílago/citología , Células Cultivadas , Condrocitos/metabolismo , Modelos Animales de Enfermedad , Humanos , Células Madre Pluripotentes Inducidas/citología , Inyecciones Intraarticulares , Masculino , Ratones , Ratones Endogámicos C57BL , Porcinos , Resultado del TratamientoRESUMEN
Delivering nucleic acids into the endothelium has great potential in treating vascular diseases. However, endothelial cells, which line the vasculature, are considered as sensitive in nature and hard to transfect. Low transfection efficacies in endothelial cells limit their potential therapeutic applications. Towards improving the transfection efficiency, we made an effort to understand the internalization of lipoplexes into the cells, which is the first and most critical step in nucleic acid transfections. In this study, we demonstrated that the transient modulation of caveolae/lipid rafts mediated endocytosis with the cholesterol-sequestrating agents, nystatin, filipin III, and siRNA against Cav-1, which significantly increased the transfection properties of cationic lipid-(2-hydroxy-N-methyl-N,N-bis(2-tetradecanamidoethyl)ethanaminium chloride), namely, amide liposomes in combination with 1,2-Dioleoyl-sn-glycero-3-phosphoethanolamine (DOPE) (AD Liposomes) in liver sinusoidal endothelial cells (SK-Hep1). In particular, nystatin was found to be highly effective with 2-3-fold enhanced transfection efficacy when compared with amide liposomes in combination with Cholesterol (AC), by switching lipoplex internalization predominantly through clathrin-mediated endocytosis and macropinocytosis.
Asunto(s)
Caveolas/efectos de los fármacos , Colesterol/química , Células Endoteliales/efectos de los fármacos , Liposomas/química , Microdominios de Membrana/efectos de los fármacos , Transfección/métodos , Animales , Caveolas/química , Caveolas/metabolismo , Caveolina 1/antagonistas & inhibidores , Caveolina 1/genética , Caveolina 1/metabolismo , Línea Celular Transformada , Colesterol/metabolismo , Clatrina/metabolismo , ADN/química , ADN/metabolismo , Endocitosis/efectos de los fármacos , Células Endoteliales/citología , Células Endoteliales/metabolismo , Filipina/química , Filipina/farmacología , Expresión Génica , Liposomas/metabolismo , Microdominios de Membrana/química , Microdominios de Membrana/metabolismo , Nistatina/química , Nistatina/farmacología , Fosfatidiletanolaminas/química , Fosfatidiletanolaminas/farmacología , Pinocitosis/efectos de los fármacos , Plásmidos/química , Plásmidos/metabolismo , ARN Interferente Pequeño/genética , ARN Interferente Pequeño/metabolismo , RatasRESUMEN
Extracellular vesicles (EVs) contain biological molecules and are secreted by cells into the extracellular milieu. The endothelial sodium channel (EnNaC) plays an important role in modulating endothelial cell stiffness. We hypothesized EVs secreted from human aortic endothelial cells (hAoECs) positively regulate EnNaC in an autocrine-dependent manner. A comprehensive lipidomic analysis using targeted mass spectrometry was performed on multiple preparations of EVs isolated from the conditioned media of hAoECs or complete growth media of these cells. Cultured hAoECs challenged with EVs isolated from the conditioned media of these cells resulted in an increase in EnNaC activity when compared with the same concentration of media-derived EVs or vehicle alone. EVs isolated from the conditioned media of hAoECs but not human fibroblast cells were enriched in MARCKS-like protein 1 (MLP1). The pharmacological inhibition of the negative regulator of MLP1, protein kinase C, in cultured hAoECs resulted in an increase in EV size and release compared with vehicle or pharmacological inhibition of protein kinase D. The MLP1-enriched EVs increased the density of actin filaments in cultured hAoECs compared with EVs isolated from human fibroblast cells lacking MLP1. We quantified 141 lipids from glycerolipids, glycerophospholipids, and sphingolipids in conditioned media EVs that represented twice the number found in control media EVs. The concentrations of sphingomyelin, lysophosphatidylcholine and phosphatidylethanolamine were higher in conditioned media EVs. These results provide the first evidence for EnNaC regulation in hAoECs by EVs and provide insight into a possible mechanism involving MLP1, unsaturated lipids, and bioactive lipids.
Asunto(s)
Proteínas de Unión a Calmodulina/genética , Medios de Cultivo Condicionados/farmacología , Células Endoteliales/metabolismo , Vesículas Extracelulares/metabolismo , Lisofosfatidilcolinas/metabolismo , Proteínas de Microfilamentos/genética , Fosfatidiletanolaminas/metabolismo , Esfingomielinas/metabolismo , Citoesqueleto de Actina/genética , Citoesqueleto de Actina/metabolismo , Citoesqueleto de Actina/ultraestructura , Aorta/citología , Aorta/metabolismo , Comunicación Autocrina , Proteínas de Unión a Calmodulina/metabolismo , Medios de Cultivo Condicionados/química , Medios de Cultivo Condicionados/metabolismo , Células Endoteliales/citología , Células Endoteliales/efectos de los fármacos , Vesículas Extracelulares/química , Expresión Génica , Glicerofosfolípidos/metabolismo , Humanos , Lipidómica/métodos , Lisofosfatidilcolinas/farmacología , Proteínas de Microfilamentos/metabolismo , Fosfatidiletanolaminas/farmacología , Cultivo Primario de Células , Proteína Quinasa C/antagonistas & inhibidores , Proteína Quinasa C/genética , Proteína Quinasa C/metabolismo , Inhibidores de Proteínas Quinasas/farmacología , Transducción de Señal , Esfingomielinas/farmacologíaRESUMEN
Phosphatidylethanolamine is a major component of phospholipids with both structural and metabolic functions in cells. Previous studies have revealed that phosphatidylethanolamine can modulate autophagy with a protective effect against age-related diseases. We examined the effect of dietary supplementation with phosphatidylethanolamine on stress response and aging in Caenorhabditis elegans. Phosphatidylethanolamine increased resistance to oxidative stress without effect on heat stress or ultraviolet irradiation. Both mean and maximum lifespans were significantly increased by phosphatidylethanolamine while fertility was reduced as a trade-off. Age-related decline of muscle function was delayed in animals treated with phosphatidylethanolamine. Supplementation with phosphatidylethanolamine suppressed toxic effect of amyloid ß and high-glucose diet. Increased ROS levels and induction of stress-responsive genes after dietary supplementation with phosphatidylethanolamine suggest that anti-oxidative stress and anti-aging effects of phosphatidylethanolamine might be though hormesis. Genetic analysis using long-lived mutants and knockdown by RNAi revealed that the lifespan-extending effect of phosphatidylethanolamine overlapped with that of reduced insulin/IGF-1-like signaling and required DAF-16, a downstream transcription factor known to regulate the expression of many stress-responsive genes. These findings indicate that phosphatidylethanolamine has anti-oxidative stress and anti-aging activities with its underlying mechanisms involving hormesis and reduced insulin/IGF-1-like signaling in C. elegans.
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Envejecimiento/efectos de los fármacos , Antioxidantes/farmacología , Proteínas de Caenorhabditis elegans/metabolismo , Caenorhabditis elegans/metabolismo , Factor I del Crecimiento Similar a la Insulina/metabolismo , Insulina/metabolismo , Fosfatidiletanolaminas/farmacología , Envejecimiento/genética , Envejecimiento/metabolismo , Animales , Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/genética , Insulina/genética , Factor I del Crecimiento Similar a la Insulina/genéticaRESUMEN
We have recently reported that phosphatidylethanolamine (PE)-containing liposomes are endocytosed and then induce lipid droplets (LDs) in HEK293T cells. In this study, we elucidated a mechanism responsible for endocytosis of PE-containing liposomes and induction of LDs. By using fluorescence-labeled liposomes and flow cytometry, we found that PE-containing liposomes were very efficiently internalized in HEK293T cells. However, Block lipid transporter-1 (BLT-1) only marginally suppressed the uptake of these liposomes, indicating that entire liposomes were mostly taken up in these cells. They were therefore inferred to express abundant PE receptors responsible for endocytosis of PE-containing liposomes. We examined the expression of 52 candidate genes through transcriptomic analyses and eventually narrowed it down to four candidate genes, which were abundantly expressed in HEK293T cells. Among siRNAs targeting these candidates, scavenger receptor class B type 1 (SR-B1) siRNA showed the most profound reduction in PE liposomal uptake. Conversely, the expression of SR-B1 by transfection of an expression plasmid enhanced the uptake of PE-containing liposomes. After the internalization of PE-containing liposomes, they were colocalized with endosomes/lysosomes and SR-B1, which indicates that these liposomes are taken up in HEK293T cells at least partially through the endosomal/lysosomal pathway. A specific anti-SR-B1-antibody blocked the uptake of PE-containing liposomes in HEK293T cells while LD formation in these cells induced by PE-containing liposomes was suppressed by treatment with SR-B1 siRNA. These results demonstrate that SR-B1 functions as a receptor for the endocytosis of PE-containing liposomes and regulates the formation of LDs induced by PE-containing liposomes in HEK293T cells.
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Endocitosis/genética , Gotas Lipídicas/metabolismo , Receptores de Leucotrieno B4/genética , Receptores Depuradores de Clase B/genética , Animales , Transporte Biológico/genética , Cricetinae , Células HEK293 , Humanos , Liposomas/metabolismo , Fosfatidiletanolaminas/química , Fosfatidiletanolaminas/farmacología , ARN Interferente Pequeño/químicaRESUMEN
The structures of bioactive polar lipids (PLs) of Irish ale with potent antithrombotic and cardioprotective properties were elucidated. Ale PL was fractionated by preparative thin layer chromatography (TLC) into subclasses, and their antithrombotic effect was assessed against human platelet aggregation induced by the pro-inflammatory mediator, platelet-activating factor (PAF). The fatty acid content and the overall structures of ale PL were elucidated by liquid chromatography mass spectrometry (LC-MS). Phosphatidylcholines (PC) and molecules of the sphingomyelin (SM) family exhibited the strongest anti-PAF effects, followed by phosphatidylethanolamines (PE). PC contained higher amounts of omega-3 polyunsaturated fatty acids (n-3 PUFA) and thus the lowest n-6/n-3 ratio. Bioactive diacyl and alkyl-acyl PC and PE molecules bearing n-3 PUFA at their sn-2 position, especially docosahexaenoic acid (DHA) and α-linolenic acid (ALA) but mostly oleic acid (OA), were identified in both PC and PE subclasses. Eicosapentaenoic acid (EPA) was present only in bioactive PC molecules and not in PE, explaining the lower anti-PAF effects of PE. Bioactive sphingolipid and glycolipid molecules with reported anti-inflammatory and anti-tumour properties, such as specific ceramides and glucosylcerebrosides with sphingosine, phytosphingosine and dihydrosphingosine bases but also specific monogalactodiglycerides and SM species bearing ALA at their sn-2 position, were identified in the SM subclass, providing a rational for its strong bioactivities against the PAF pathway. Further studies are required on the health benefits of bioactive PL from beer and brewery by-products.
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Cerveza/análisis , Factor de Activación Plaquetaria/antagonistas & inhibidores , Inhibidores de Agregación Plaquetaria/análisis , Inhibidores de Agregación Plaquetaria/farmacología , Ácidos Grasos Omega-3/análisis , Ácidos Grasos Omega-3/farmacología , Humanos , Fosfatidilcolinas/análisis , Fosfatidilcolinas/farmacología , Fosfatidiletanolaminas/análisis , Fosfatidiletanolaminas/farmacología , Agregación Plaquetaria/efectos de los fármacos , Esfingomielinas/análisis , Esfingomielinas/farmacologíaRESUMEN
An efficient three-step strategy for the convenient synthesis of Sn-glycero-3-phosphoethanolamine (GroPEtn) from a commercially available 1,2-dipalmitoyl-sn-glycero-3-phosphoethanolamine (DPPE) is reported. Direct hydrolysis of DPPE produces a complex inseparable mixture, hence a protection and deprotection strategy is employed to prepare GroPEtn. The primary amine of DPPE is protected with a highly stable acid-labile trityl group, followed by strong base hydrolysis of N-trityl-DPPE gives N-trityl-GroPEtn. Further a mild, rapid, and efficient deprotection method is established using trifluoroacetic acid to remove N-trityl moiety, affords GroPEtn as a single product. This is the first semisynthetic approach and efficient method to produce GroPEtn with a total yield of 66% in three steps. GroPEtn did not show any cytotoxicity against human kidney (HK-2) cells and reporter gene assay for activation of Keap1-Nrf2-mediated antioxidant defense mechanism showed no significant effects.
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Fosfatidiletanolaminas/química , Fosfatidiletanolaminas/síntesis química , Línea Celular , Proliferación Celular , Humanos , Proteína 1 Asociada A ECH Tipo Kelch/metabolismo , Factor 2 Relacionado con NF-E2/metabolismo , Fosfatidiletanolaminas/farmacología , Ácido Trifluoroacético/químicaRESUMEN
Brain aging involves changes in the lipid membrane composition that lead to a decrease in membrane excitability and neurotransmitter release. These membrane modifications have been identified as contributing factors in age-related memory decline. In this sense, precursors of phospholipids (PLs) can restore the physiological composition of cellular membranes and produce valuable therapeutic effects in brain aging. Among promising drugs, alpha-glycerylphosphorylethanolamine (GPE) has demonstrated protective effects in amyloid-injured astrocytes and in an aging model of human neural stem cells. However, the compound properties on mature neuronal cells remain unexplored. Herein, GPE was tested in human hippocampal neurons, which are involved in learning and memory, and characterized by a functional cholinergic transmission, thus representing a valuable cellular model to explore the beneficial properties of GPE. GPE induced the release of the main membrane phospholipids and of the acetylcholine neurotransmitter. Moreover, the compound reduced lipid peroxidation and enhanced membrane fluidity of human brain cells. GPE counteracted the DNA damage and viability decrease observed in in vitro aged neurons. Among GPE treatment effects, the autophagy was found positively upregulated. Overall, these results confirm the beneficial effects of GPE treatment and suggest the compound as a promising drug to preserve hippocampal neurons and virtually memory performances.
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Hipocampo/citología , Fármacos Neuroprotectores/farmacología , Nootrópicos/farmacología , Fosfatidiletanolaminas/farmacología , Línea Celular , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Hipocampo/efectos de los fármacos , Humanos , Peroxidación de Lípido/efectos de los fármacos , Modelos Biológicos , Fosfatidilcolinas/metabolismo , Fosfatidiletanolaminas/metabolismoRESUMEN
Therapeutic peptides (TPs) are biological macromolecules which can act as neurotransmitters, hormones, ion channel ligands and growth factors. Undoubtedly, TPs are crucial in modern medicine. But low bio-stability and some special adverse reactions reduce their places to the application. With the development of nanotechnology, nanoparticles (NPs) in pharmaceutical science gained much attention. They can encapsulate the TPs into their membrane or shell. Therefore, they can protect the TPs against degradation and then increase the bioavailability, which was thought to be the biggest advantage of them. Additionally, targeting was also studied to improve the effect of TPs. However, there were some drawbacks of nano TPs like low loading efficiency and difficulty to manufacture. Nowadays, lots of studies focused on improving effect of TPs by preparing nanoparticles. In this review, we presented a brief analysis of peptide-combined nanoparticles. Their advantages and disadvantages were listed in terms of mechanism. And several examples of applications were summarized.