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Oxylipins are potent lipid mediators with increasing interest in clinical research. They are usually measured in systemic circulation and can provide a wealth of information regarding key biological processes such as inflammation, vascular tone, or blood coagulation. Although procedures still require harmonization to generate comparable oxylipin datasets, performing comprehensive profiling of circulating oxylipins in large studies is feasible and no longer restricted by technical barriers. However, it is essential to improve and facilitate the biological interpretation of complex oxylipin profiles to truly leverage their potential in clinical research. This requires regular updating of our knowledge about the metabolism and the mode of action of oxylipins, and consideration of all factors that may influence circulating oxylipin profiles independently of the studied disease or condition. This review aims to provide the readers with updated and necessary information regarding oxylipin metabolism, their different forms in systemic circulation, the current limitations in deducing oxylipin cellular effects from in vitro bioactivity studies, the biological and technical confounding factors needed to consider for a proper interpretation of oxylipin profiles.
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Oxilipinas , Oxilipinas/sangre , Oxilipinas/metabolismo , Humanos , Animales , Investigación BiomédicaRESUMEN
Inflammation is initiated and driven by a mixture of mediators, which modify effects of each other. This study analysed in vitro pro-inflammatory activity of inflammatory cytokines (TNFα and IL-1ß) in a combination with a lipid DAMP molecule, oxidized palmitoyl-arachidonoyl-phosphatidylcholine (OxPAPC). The study was performed on endothelial and monocytic cell lines. The cells were treated with different concentrations of TNFα or IL-1ß, OxPAPC and their combinations, either in the presence or absence of drugs regulating inflammation. Pro-inflammatory effects of TNFα/IL-1ß and OxPAPC were estimated by analysis of chemokines CXCL8, CXCL2 and CXCL3 by ELISA and RT-PCR. Toxicity was determined by analysis of metabolic activity. Statistical significance was estimated by ANOVA and Dunnett's test. OxPAPC was a much weaker chemokine inducer as compared to TNFα or IL-1ß. However, OxPAPC and TNFα/IL-1ß together induced effects that were significantly stronger than the arithmetical sum of individual effects. This cooperative action of OxPAPC and TNFα was reversed by inhibitors of p38 MAPK. We hypothesise that the boosting of TNFα and IL-1ß effects by OxPAPC may be more pathologically important than the action of the lipid alone. Inhibitors of p38 MAPK may become a tool for analysis of pathological role of oxidized phospholipids.
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Oxidised phospholipids such as oxidised palmitoyl-arachidonoyl-phosphatidylcholine (OxPAPC) are increasingly recognised as danger-associated molecular patterns (DAMPs) inducing cyto- and chemokines. The pathological impact of oxidised phosphatidylcholine in vivo has been demonstrated in several animal models, as well as in human association studies. In this work, we have tested a number of small molecules with known or potential anti-inflammatory properties for their ability to inhibit secretion of interleukin-8 by OxPAPC-treated endothelial cells. Six compounds capable of inhibiting the induction of IL-8 were selected. Analysis of gene expression has shown that all these substances reduced the OxPAPC-induced elevation of IL-8 mRNA but potentiated induction of heat-shock proteins (HSPs). We further found that drug-like HSP inducers also prevented the induction of IL-8 by OxPAPC. Similar inhibitory action was demonstrated by two chemical chaperones, which stabilise proteins through physicochemical mechanisms thus mimicking effects of HSPs. Our data suggest that proteostatic stress plays an important mechanistic role in the pro-inflammatory effects of OxPAPC and that stabilisation of proteome by overexpression of HSPs or by chemical chaperones can reduce the pro-inflammatory effects of OxPLs.
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Interleucina-8 , Fosfolípidos , Animales , Humanos , Fosfolípidos/farmacología , Fosfolípidos/química , Fosfolípidos/metabolismo , Interleucina-8/metabolismo , Regulación hacia Arriba , Células Endoteliales/metabolismo , Proteínas de Choque Térmico/metabolismo , Fosfatidilcolinas/farmacología , Fosfatidilcolinas/metabolismoRESUMEN
Oxidized phospholipids (OxPLs) are generated by enzymatic or autooxidation of esterified polyunsaturated fatty acids (PUFAs) residues. OxPLs are present in circulation and atherosclerotic plaques where they are thought to induce predominantly proinflammatory and toxic changes in endothelial (ECs) and other cell types. Unexpectedly, we found that low concentrations of OxPLs were not toxic but protected ECs from stress induced by serum deprivation or cytostatic drugs. The protective effect was observed in ECs obtained from different vessels and was monitored using a variety of readouts based on different biological and chemical principles. Analysis of the structure−activity relationship identified oxidized or missing fatty acid residue (OxPLs or Lyso-PLs, respectively) as a prerequisite for the protective action of a PL. Protective OxPLs or Lyso-PLs acquired detergent-like properties and formed in solution aggregates <10 nm in diameter (likely micelles), which were in striking contrast with large aggregates (>1000 nm, likely multilayer liposomes) produced by nonoxidized precursor PLs. Because surfactants, OxPLs, and Lyso-PLs are known to extract membrane cholesterol, we tested if this effect might trigger the protection of endothelial cells. The protective action of OxPLs and Lyso-PLs was inhibited by cotreatment with cholesterol and mimicked by cholesterol-binding beta-cyclodextrin but not inactive α-cyclodextrin. Wide-scale mRNA expression analysis in four types of ECs showed the induction of genes encoding for heat shock proteins (HSPs) and secreted prosurvival peptides and proteins. Inducers of HSPs, chemical chaperones, and pure prosurvival factors mimicked the protective action of OxPLs/Lyso-PLs. We hypothesize that oxidation changes the physicochemical properties of PLs, thus promoting membrane cholesterol redistribution or extraction leading to the expression of intra- and extracellular prosurvival factors.
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Oxidized phospholipids (OxPLs) containing enzymatically or non-enzymatically oxidized fatty acids (oxylipins) are increasingly recognized as lipid mediators involved in pathogenesis of diseases. Further understanding of structure-activity relationship and molecular mechanisms activated by OxPLs is hampered by the complexity of synthesis of individual molecular species. Although dozens of individual free oxylipins are commercially available, their attachment to the phospholipid scaffold requires relatively harsh conditions during activation of carboxy-group, which may lead to decomposition of unstable oxylipins. Furthermore, additional protection-deprotection steps are required for oxylipins containing hydroxy-groups. In this work we describe synthesis of OxPLs containing oxylipins bound at the sn-2-position via an amide-bond that is characteristic of sphingophospholipids. Activation of oxylipins and attachment to the phospholipid scaffold are performed under mild conditions and characterized by high yield. Hydroxy-groups of oxylipins do not interfere with reactions and therefore no protection/deprotection steps are needed. In order to prevent oxylipin migration, a fatty acid residue at the sn-1 was bound through an alkyl bond, which is a common bond present in a large proportion of naturally occurring phospholipids. An additional advantage of combining alkyl and amide bonds in a single phospholipid molecule is that both types of bonds are phospholipase A1/A2-resistant, which may be expected to improve biological stability of OxPLs and thus simplify analysis of their effects. As proof of principle, several alkyl-amide oxidized phosphatidylcholines (OxPCs) containing either linear or prostane ring oxylipins have been synthesized. Importantly, we show here that alkyl-amide-OxPCs demonstrated biological activities similar to those of di-acyl-OxPCs. Alkyl-amide-OxPCs inhibited pro-inflammatory action of LPS and increased endothelial cellular barrier in vitro and in mouse models. The effects of alkyl-amide and di-acyl-OxPCs developed in a similar range of concentrations. We hypothesize that alkyl-amide-OxPLs may become a useful tool for deeper analysis of the structure-activity relationship of OxPLs.
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Endotoxinas , Fosfolípidos , Amidas , Animales , Ratones , Oxidación-Reducción , FosfatidilcolinasRESUMEN
Processed and ready-to-eat foods become routinely consumed resulting in a sharp rise of sugar intake in people's daily diets. The inclusion of fresh fruits and vegetables rich in (poly)phenols has been encouraged by the World Health Organization (WHO) as part of the daily choices to ameliorate endothelial dysfunction and ease the socio-economic burden of diabetes. Research in Food, Nutrition, and Cell Metabolism areas is revealing that the health benefits of (poly)phenol-rich foods go beyond their antioxidant properties and are in fact key modulators of redox and glycaemia status, and inflammatory response contributing to improved endothelial function and vascular health in diabetes. Other beneficial aspects include appetite modulation, regulation of hydrolytic enzymes involved in sugar and lipid metabolism, and mediation of cell-cell aggregation events. This work overviews the current knowledge on the biological properties of ingested (poly)phenols in cultured endothelial cells with emphasis on the circulating (poly)phenols, providing support to (poly)phenol-rich diets as alternatives to drug-based therapies in the prevention, treatment, and management of diabetes. A critical evaluation on the caveats and challenges involve in current experimental cell-based designs and approaches adopted is also discussed.
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Diabetes Mellitus/fisiopatología , Dieta , Endotelio Vascular/efectos de los fármacos , Endotelio Vascular/fisiopatología , Polifenoles/farmacología , Animales , Apetito/efectos de los fármacos , Células Cultivadas , Diabetes Mellitus/dietoterapia , Células Endoteliales , Humanos , Obesidad/dietoterapia , Oxidación-Reducción , Polifenoles/sangre , Polifenoles/metabolismo , Trombosis/prevención & controlRESUMEN
A comprehensive and standardized system to report lipid structures analyzed by MS is essential for the communication and storage of lipidomics data. Herein, an update on both the LIPID MAPS classification system and shorthand notation of lipid structures is presented for lipid categories Fatty Acyls (FA), Glycerolipids (GL), Glycerophospholipids (GP), Sphingolipids (SP), and Sterols (ST). With its major changes, i.e., annotation of ring double bond equivalents and number of oxygens, the updated shorthand notation facilitates reporting of newly delineated oxygenated lipid species as well. For standardized reporting in lipidomics, the hierarchical architecture of shorthand notation reflects the diverse structural resolution powers provided by mass spectrometric assays. Moreover, shorthand notation is expanded beyond mammalian phyla to lipids from plant and yeast phyla. Finally, annotation of atoms is included for the use of stable isotope-labeled compounds in metabolic labeling experiments or as internal standards. This update on lipid classification, nomenclature, and shorthand annotation for lipid mass spectra is considered a standard for lipid data presentation.
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Lípidos/química , Espectrometría de Masas , Terminología como AsuntoRESUMEN
Circulating oxidized phospholipids are increasingly recognized as biomarkers of atherosclerosis. Clinical association studies have been mainly performed using an immune assay based on monoclonal antibody E06, which recognizes a variety of molecular species of oxidized phosphatidylcholine (OxPC) in lipoproteins, cell membranes or covalently bound to plasma proteins. Accumulating evidence shows that individual molecular species of OxPC demonstrate different biological activities and have different half-life times. Therefore, it is likely that certain molecular species can be associated with pathology more strongly than others. This hypothesis can only be tested using LC-MS/MS allowing quantification of individual molecular species of OxPCs. In order to ensure that laborious LC-MS/MS methods do not simply replicate the results of a technically simpler E06-OxPCs assay, we have performed relative quantification of 8 truncated molecular species of OxPCs in plasma of 132 probands and compared the data with the results of the E06-OxPCs and OxLDL assays. We have found a strong correlation between individual molecular species of OxPCs but only a weak correlation of LC-MS/MS-OxPCs data with the E06-OxPCs assay and no correlation with the OxLDL assay. Furthermore, in contrast to the results of E06-OxPCs or OxLDL assays, 7 out of 8 OxPC species were associated with hypertension. The data suggest that the results of the LC-MS/MS-OxPCs assay do not replicate the results of two ELISA-based lipid oxidation tests and therefore may produce additional diagnostic information. These findings necessitate development of simplified mass spectrometric procedures for high-throughput and affordable analysis of selected molecular species of OxPCs.
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Enfermedad de la Arteria Coronaria/sangre , Dislipidemias/sangre , Hipertensión/sangre , Fosfatidilcolinas/sangre , Adulto , Biomarcadores/sangre , Colesterol/sangre , Cromatografía Liquida , Estudios de Cohortes , Enfermedad de la Arteria Coronaria/diagnóstico , Enfermedad de la Arteria Coronaria/fisiopatología , Creatinina/sangre , Dislipidemias/diagnóstico , Dislipidemias/fisiopatología , Ensayo de Inmunoadsorción Enzimática , Femenino , Humanos , Hipertensión/diagnóstico , Hipertensión/fisiopatología , Masculino , Persona de Mediana Edad , Oxidación-Reducción , Fosfatidilcolinas/clasificación , Espectrometría de Masas en Tándem , Triglicéridos/sangreRESUMEN
Ultraviolet light is the dominant environmental oxidative skin stressor and a major skin aging factor. We studied which oxidized phospholipid (OxPL) mediators would be generated in primary human keratinocytes (KC) upon exposure to ultraviolet A light (UVA) and investigated the contribution of OxPL to UVA responses. Mass spectrometric analysis immediately or 24â¯h post UV stress revealed significant changes in abundance of 173 and 84 lipid species, respectively. We identified known and novel lipid species including known bioactive and also potentially reactive carbonyl containing species. We found indication for selective metabolism and degradation of selected reactive lipids. Exposure to both UVA and to in vitro UVA - oxidized phospholipids activated, on transcriptome and proteome level, NRF2/antioxidant response signaling, lipid metabolizing enzyme expression and unfolded protein response (UPR) signaling. We identified NUPR1 as an upstream regulator of UVA/OxPL transcriptional stress responses and found this protein to be expressed in the epidermis. Silencing of NUPR1 resulted in augmented expression of antioxidant and lipid detoxification genes and disturbed the cell cycle, making it a potential key factor in skin reactive oxygen species (ROS) responses intimately involved in aging and pathology.
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Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/genética , Queratinocitos/metabolismo , Queratinocitos/efectos de la radiación , Proteínas de Neoplasias/genética , Oxidación-Reducción/efectos de la radiación , Fosfolípidos/metabolismo , Estrés Fisiológico/genética , Estrés Fisiológico/efectos de la radiación , Rayos Ultravioleta , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/metabolismo , Perfilación de la Expresión Génica , Regulación de la Expresión Génica , Humanos , Metabolismo de los Lípidos , Metaboloma , Metabolómica/métodos , Modelos Biológicos , Proteínas de Neoplasias/metabolismo , TranscriptomaRESUMEN
We have reported recently that inactivation of the essential autophagy-related gene 7 (Atg7) in keratinocytes has little or no impact on morphology and function of the epidermal barrier in experimental animals. When these mice aged, mutant males, (Atg7 ΔKC), developed an oily coat. As the keratin 14 promoter driven cre/LoxP system inactivates floxed Atg7 in all keratin 14 (K14) expressing cells, including sebocytes, we investigated whether the oily hair phenotype was the consequence of changes in function of the skin sebaceous glands. Using an antibody to the GFP-LC3 fusion protein, autophagosomes were detected at the border of sebocyte disintegration in control but not in mutant animals, suggesting that autophagy was (a) active in normal sebaceous glands and (b) was inactivated in the mutant mice. Detailed analysis established that dorsal sebaceous glands were about twice as large in all Atg7 ΔKC mice compared to those of controls (Atg7 F/F), and their rate of sebocyte proliferation was increased. In addition, male mutant mice yielded twice as much lipid per unit hair as age-matched controls. Analysis of sebum lipids by thin layer chromatography revealed a 40% reduction in the proportion of free fatty acids (FFA) and cholesterol, and a 5-fold increase in the proportion of fatty acid methyl esters (FAME). In addition, the most common diester wax species (58-60 carbon atoms) were increased, while shorter species (54-55 carbon atoms) were under-represented in mutant sebum. Our data show that autophagy contributes to sebaceous gland function and to the control of sebum composition.
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Proteína 7 Relacionada con la Autofagia/genética , Autofagia/genética , Glándulas Sebáceas/patología , Glándulas Sebáceas/fisiopatología , Sebo/química , Animales , Autofagosomas , Proliferación Celular/genética , Colesterol/análisis , Ácidos Grasos no Esterificados/análisis , Cabello , Masculino , Ratones , Fenotipo , Ceras/análisisRESUMEN
Receptor tyrosine kinase c-Kit and its ligand stem cell factor (SCF) regulate resident vascular wall cells and recruit circulating progenitors. We tested whether SCF may be induced by oxidized palmitoyl-arachidonoyl-phosphatidylcholine (OxPAPC) known to accumulate in atherosclerotic vessels. Gene expression analysis demonstrated OxPAPC-induced upregulation of SCF mRNA and protein in different types of endothelial cells (ECs). Elevated levels of SCF mRNA were observed in aortas of ApoE-/- knockout mice. ECs produced biologically active SCF because conditioned medium from OxPAPC-treated cells stimulated activation (phosphorylation) of c-Kit in naïve ECs. Induction of SCF by OxPAPC was inhibited by knocking down transcription factor NRF2. Inhibition or stimulation of NRF2 by pharmacological or molecular tools induced corresponding changes in SCF expression. Finally, we observed decreased levels of SCF mRNA in aortas of NRF2 knockout mice. We characterize OxPLs as a novel pathology-associated stimulus inducing expression of SCF in endothelial cells. Furthermore, our data point to transcription factor NRF2 as a major mediator of OxPL-induced upregulation of SCF. This mechanism may represent one of the facets of pleiotropic action of NRF2 in vascular wall.
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Aorta/metabolismo , Regulación de la Expresión Génica , Factor 2 Relacionado con NF-E2/metabolismo , Fosfatidilcolinas/metabolismo , Factor de Células Madre/biosíntesis , Animales , Aorta/patología , Apolipoproteínas E/deficiencia , Células Endoteliales de la Vena Umbilical Humana/metabolismo , Células Endoteliales de la Vena Umbilical Humana/patología , Humanos , Masculino , Ratones , Ratones Noqueados para ApoE , Factor 2 Relacionado con NF-E2/genética , Oxidación-Reducción , Fosfatidilcolinas/genética , Factor de Células Madre/genéticaRESUMEN
Correction of barrier dysfunction and inflammation in acute lung injury (ALI) represents an important problem. Previous studies demonstrate barrier-protective and anti-inflammatory effects of bioactive lipid prostacyclin and its stable analog iloprost (ILO). We generated a phospholipase resistant synthetic phospholipid with iloprost attached at the sn-2 position (ILO-PC) and investigated its biological effects. In comparison to free ILO, ILO-PC caused sustained endothelial cell (EC) barrier enhancement, linked to more prolonged activation of Rap1 and Rac1 GTPases and their cytoskeletal and cell junction effectors: cortactin, PAK1, p120-catenin and VE-cadherin. ILO and ILO-PC equally efficiently suppressed acute, Rho GTPase-dependent EC hyper-permeability caused by thrombin. However, ILO-PC exhibited more sustained barrier-protective and anti-inflammatory effects in the model of chronic EC dysfunction caused by bacterial wall lipopolysacharide (LPS). ILO-PC was also more potent inhibitor of NFκB signaling and lung vascular leak in the murine model of LPS-induced ALI. Treatment with ILO-PC showed more efficient ALI recovery over 3 days after LPS challenge than free ILO. In conclusion, this study describes a novel synthetic phospholipid with barrier-enhancing and anti-inflammatory properties superior to existing prostacyclin analogs, which may be used as a prototype for future development of more efficient treatment for ALI and other vascular leak syndromes.
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Células Endoteliales/efectos de los fármacos , Iloprost/farmacología , Pulmón/efectos de los fármacos , Fosfolipasas/metabolismo , Fosfolípidos/metabolismo , Sustancias Protectoras/farmacología , Lesión Pulmonar Aguda/tratamiento farmacológico , Lesión Pulmonar Aguda/metabolismo , Uniones Adherentes/efectos de los fármacos , Uniones Adherentes/metabolismo , Animales , Antígenos CD/metabolismo , Cadherinas/metabolismo , Permeabilidad Capilar/efectos de los fármacos , Línea Celular , Citoesqueleto/efectos de los fármacos , Citoesqueleto/metabolismo , Células Endoteliales/metabolismo , Endotelio Vascular/efectos de los fármacos , Endotelio Vascular/metabolismo , Epoprostenol/metabolismo , Humanos , Lipopolisacáridos/farmacología , Pulmón/metabolismo , Ratones , Ratones Endogámicos C57BL , FN-kappa B/metabolismo , Transducción de Señal/efectos de los fármacos , Trombina/metabolismo , Proteínas de Unión al GTP rap1/metabolismoRESUMEN
Prostaglandins (PG), the products of cyclooxygenase-mediated conversion of arachidonic acid, become upregulated in many situations including allergic response, inflammation, and injury, and exhibit a variety of biological activities. Previous studies described barrier-enhancing and anti-inflammatory effects of PGE2 and PGI2 on vascular endothelial cells (EC). Yet, the effects of other PG members on EC barrier and inflammatory activation have not been systematically analyzed. This study compared effects of PGE2, PGI2, PGF2α, PGA2, PGJ2, and PGD2 on human pulmonary EC. EC permeability was assessed by measurements of transendothelial electrical resistance and cell monolayer permeability for FITC-labeled tracer. Anti-inflammatory effects of PGs were evaluated by analysis of expression of adhesion molecule ICAM1 and secretion of soluble ICAM1 and cytokines by EC. PGE2, PGI2, and PGA2 exhibited the most potent barrier-enhancing effects and most efficient attenuation of thrombin-induced EC permeability and contractile response, whereas PGI2 effectively suppressed thrombin-induced permeability but was less efficient in the attenuation of prolonged EC hyperpermeability caused by interleukin-6 or bacterial wall lipopolysaccharide, LPS. PGD2 showed a modest protective effect on the EC inflammatory response, whereas PGF2α and PGJ2 were without effect on agonist-induced EC barrier dysfunction. In vivo, PGE2, PGI2, and PGA2 attenuated LPS-induced lung inflammation, whereas PGF2α and PGJ2 were without effect. Interestingly, PGD2 exhibited a protective effect in the in vivo model of LPS-induced lung injury. This study provides a comprehensive analysis of barrier-protective and anti-inflammatory effects of different prostaglandins on lung EC in vitro and in vivo and identifies PGE2, PGI2, and PGA2 as prostaglandins with the most potent protective properties.
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Permeabilidad de la Membrana Celular/efectos de los fármacos , Endotelio Vascular/efectos de los fármacos , Inflamación/tratamiento farmacológico , Lesión Pulmonar/tratamiento farmacológico , Prostaglandinas/farmacología , Animales , Endotelio Vascular/citología , Endotelio Vascular/metabolismo , Hemostáticos/efectos adversos , Humanos , Inflamación/inducido químicamente , Inflamación/patología , Molécula 1 de Adhesión Intercelular/metabolismo , Interleucina-6/metabolismo , Lipopolisacáridos/efectos adversos , Lesión Pulmonar/inducido químicamente , Lesión Pulmonar/patología , Ratones , Trombina/efectos adversosRESUMEN
Unlike other agonists that cause transient endothelial cell (EC) response, the products of 1-palmitoyl-2-arachidonoyl-sn-glycero-3-phosphocholine (PAPC) oxidation that contain cyclopenthenone groups, which recapitulate prostaglandin-like structure, cause sustained enhancement of the pulmonary EC barrier. The mechanisms that drive the sustained effects by oxidized PAPC (OxPAPC) remain unexplored. On the basis of the structural similarity of isoprostanoid moieties that are present in full-length oxygenated PAPC species, we used an inhibitory approach to perform the screening of prostanoid receptors as potential candidates that mediate OxPAPC effects. Results show that only prostaglandin E receptor-4 (EP4) was involved and mediated the sustained phase of the barrier-enhancing effects of OxPAPC that are associated with the activation of Rac GTPase and its cytoskeletal targets. EC incubation with OxPAPC also induced EP4 mRNA expression in pulmonary ECs and lung tissue. EP4 knockdown using gene-specific small interfering RNA did not affect the rapid phase of OxPAPC-induced EC barrier enhancement or the protective effects against thrombin-induced EC permeability, but abolished the advanced barrier enhancement phase and suppressed the protective effects of OxPAPC against more sustained EC barrier dysfunction and cell inflammatory response caused by TNF-α. Endothelial-specific knockout of the EP4 receptor in mice attenuated the protective effect of intravenous OxPAPC administration in the model of acute lung injury caused by intratracheal injection of LPS. Taken together, these results demonstrate a novel role for prostaglandin receptor EP4 in the mediation of barrier-enhancing and anti-inflammatory effects caused by oxidized phospholipids.-Oskolkova, O., Gawlak, G., Tian, Y., Ke, Y., Sarich, N., Son, S., Andreasson, K., Bochkov, V. N., Birukova, A. A., Birukov, K. G. Prostaglandin E receptor-4 receptor mediates endothelial barrier-enhancing and anti-inflammatory effects of oxidized phospholipids.
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Células Endoteliales/fisiología , Fosfatidilcolinas/metabolismo , Subtipo EP4 de Receptores de Prostaglandina E/metabolismo , Uniones Adherentes/fisiología , Animales , Citoesqueleto , Impedancia Eléctrica , Humanos , Inflamación/metabolismo , Lesión Pulmonar , Ratones , Ratones Noqueados , Oxidación-Reducción , Fosfatidilcolinas/química , Fosfolípidos , Subtipo EP4 de Receptores de Prostaglandina E/genética , Trombina , Factor de Necrosis Tumoral alfa , Proteína de Unión al GTP rac1/genética , Proteína de Unión al GTP rac1/metabolismoRESUMEN
RATIONALE: Oxidation of 1-palmitoyl-2-arachidonoyl-sn-glycero-3-phosphorylcholine (OxPAPC) generates a group of bioactive oxidized phospholipid products with a broad range of biological activities. Barrier-enhancing and anti-inflammatory effects of OxPAPC on pulmonary endothelial cells are critical for prevention of acute lung injury caused by bacterial pathogens or excessive mechanical ventilation. Anti-inflammatory properties of OxPAPC are associated with its antagonistic effects on Toll-like receptors and suppression of RhoA GTPase signaling. OBJECTIVE: Because OxPAPC exhibits long-lasting anti-inflammatory and lung-protective effects even after single administration in vivo, we tested the hypothesis that these effects may be mediated by additional mechanisms, such as OxPAPC-dependent production of anti-inflammatory and proresolving lipid mediator, lipoxin A4 (LXA4). METHODS AND RESULTS: Mass spectrometry and ELISA assays detected significant accumulation of LXA4 in the lungs of OxPAPC-treated mice and in conditioned medium of OxPAPC-exposed pulmonary endothelial cells. Administration of LXA4 reproduced anti-inflammatory effect of OxPAPC against tumor necrosis factor-α in vitro and in the animal model of lipopolysaccharide-induced lung injury. The potent barrier-protective and anti-inflammatory effects of OxPAPC against tumor necrosis factor-α and lipopolysaccharide challenge were suppressed in human pulmonary endothelial cells with small interfering RNA-induced knockdown of LXA4 formyl peptide receptor-2 (FPR2/ALX) and in mFPR2-/- (mouse formyl peptide receptor 2) mice lacking the mouse homolog of human FPR2/ALX. CONCLUSIONS: This is the first demonstration that inflammation- and injury-associated phospholipid oxidation triggers production of anti-inflammatory and proresolution molecules, such as LXA4. This lipid mediator switch represents a novel mechanism of OxPAPC-assisted recovery of inflamed lung endothelium.
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Lesión Pulmonar Aguda/metabolismo , Antiinflamatorios no Esteroideos/uso terapéutico , Células Endoteliales/efectos de los fármacos , Células Endoteliales/metabolismo , Lipoxinas/metabolismo , Fosfatidilcolinas/uso terapéutico , Lesión Pulmonar Aguda/prevención & control , Animales , Antiinflamatorios no Esteroideos/farmacología , Células Cultivadas , Humanos , Lipoxinas/farmacología , Lipoxinas/uso terapéutico , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Fosfatidilcolinas/farmacología , Resultado del TratamientoRESUMEN
We have found that a well-characterized P2X7 receptor antagonist AZ11645373 blocked production of pro-inflammatory chemokine IL-8 in endothelial cells treated with OxPAPC. The effect was not due to toxicity of AZ11645373 as documented by cellular metabolic activity assay. The mechanism of inhibition by AZ11645373 was apparently independent of the P2X7 receptor because this receptor was not involved in induction of IL-8 under our experimental conditions. In support of this notion, two P2X7 agonists ATP and BzATP did not upregulate IL-8. On the other hand, a chemically different P2X7 receptor antagonist A740003 did not inhibit OxPAPC-induced production of IL-8. The inhibitory action of AZ11645373 was observed at the level of IL-8 protein and messenger RNA (mRNA) induction. Furthermore, AZ11645373 inhibited induction of mRNA encoding for COX-2 (PTGS2) suggesting that its anti-inflammatory potential is not limited to suppression of IL-8 production. In addition to inhibiting stimulation by OxPAPC, AZ11645373 suppressed induction of IL-8 by TNFα and LPS. To summarize, AZ11645373 inhibits in a P2X7-independent manner action of chemically different inflammatory agonists such as OxPLs, LPS, and TNFα. Thus, AZ11645373 may be especially effective for treatment of inflammatory disorders due to a beneficial combination of P2X7 receptor-dependent effects (inhibition of inflammasome activation, antinociceptive effects) with P2X7-independent general anti-inflammatory action described in this paper.
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Antiinflamatorios/farmacología , Interleucina-8/genética , Antagonistas del Receptor Purinérgico P2X/farmacología , Tiazoles/farmacología , Células Endoteliales de la Vena Umbilical Humana , Humanos , Inflamasomas/metabolismo , Interleucina-8/antagonistas & inhibidores , Interleucina-8/efectos de los fármacos , Fosfatidilcolinas/farmacología , ARN Mensajero , Receptores Purinérgicos P2X7 , Activación Transcripcional/efectos de los fármacosRESUMEN
Oxidized phospholipids (OxPLs) are increasingly recognized to play a role in a variety of normal and pathological states. OxPLs were implicated in regulation of inflammation, thrombosis, angiogenesis, endothelial barrier function, immune tolerance and other important processes. Rapidly accumulating evidence suggests that OxPLs are biomarkers of atherosclerosis and other pathologies. In addition, successful application of experimental drugs based on structural scaffold of OxPLs in animal models of inflammation was recently reported. This review briefly summarizes current knowledge on generation, methods of quantification and biological activities of OxPLs. Furthermore, receptor and cellular mechanisms of these effects are discussed. The goal of the review is to give a broad overview of this class of lipid mediators inducing pleiotropic biological effects.
Asunto(s)
Aterosclerosis/metabolismo , Endotelio/metabolismo , Neovascularización Patológica/metabolismo , Fosfolípidos/metabolismo , Trombosis/metabolismo , Animales , Aterosclerosis/inmunología , Aterosclerosis/patología , Biomarcadores/metabolismo , Endotelio/inmunología , Regulación de la Expresión Génica , Humanos , Tolerancia Inmunológica , Inflamación , Neovascularización Patológica/inmunología , Neovascularización Patológica/patología , Oxidación-Reducción , Permeabilidad , Fosfolípidos/química , Fosfolípidos/clasificación , Fosfolípidos/inmunología , Receptores Lisofosfolípidos/genética , Receptores Lisofosfolípidos/inmunología , Trombosis/inmunología , Trombosis/patologíaRESUMEN
AIM: Oxidized phospholipids (OxPL) are the major pathogenic component of oxidized low-density lipoproteins (OxLDL). Endogenous anti-OxPL activity, defined as the ability to neutralize adverse effects of oxidized lipids, may have biomarker potential. METHODS & RESULTS: Using two anti-OxPL monoclonal antibodies (commercial mAB-E06 and custom mAB-509) we developed a novel ELISA that measures the global capacity of plasma to inactivate OxPL. Preincubation of OxLDL with plasma inhibits its binding of anti-OxPL mABs. This phenomenon ('masking') reflects anti-OxPL plasma activity. A pilot clinical application of the assay revealed reduced anti-OxPL activity in hypertension, coronary artery disease, acute coronary syndrome and diabetes. CONCLUSION: Inadequate anti-OxPL protection may contribute to cardiovascular disease and have biomarker potential in conditions associated with abnormal lipid peroxidation.
Asunto(s)
Análisis Químico de la Sangre/métodos , Ensayo de Inmunoadsorción Enzimática , Fosfolípidos/sangre , Síndrome Coronario Agudo/sangre , Síndrome Coronario Agudo/patología , Adulto , Anciano , Anticuerpos Monoclonales/inmunología , Enfermedad de la Arteria Coronaria/sangre , Enfermedad de la Arteria Coronaria/patología , Diabetes Mellitus/sangre , Diabetes Mellitus/patología , Femenino , Humanos , Hipertensión/sangre , Hipertensión/patología , Lipoproteínas LDL/sangre , Masculino , Persona de Mediana Edad , Oxidación-Reducción , Fosfolípidos/inmunologíaRESUMEN
Oxidized phospholipids are generally recognized as deleterious factors involved in disease pathogenesis. This review summarizes the data suggesting that under certain biological conditions the opposite is correct, namely that OxPLs can also induce protective effects. Examples that are discussed in the review include upregulation of antioxidant genes, inhibition of inflammatory signaling pathways through Nrf2-dependent and -independent mechanisms, antagonism of Toll-like receptors, immuno-modulating and immuno-suppressive action of OxPLs in adaptive immunity and autoimmune disease, activation of PPARs known for their anti-inflammatory action, as well as protective action against lung edema in acute lung inflammation. The data support the notion that oxidation of phospholipids provides a negative feedback preventing damage to host tissues due to uncontrolled inflammation and oxidative stress.
Asunto(s)
Antiinflamatorios/metabolismo , Hormesis , Oxidación-Reducción , Fosfolípidos/metabolismo , Inmunidad Adaptativa/efectos de los fármacos , Animales , Antiinflamatorios/química , Antiinflamatorios/farmacología , Citocinas/metabolismo , Humanos , Inmunomodulación/efectos de los fármacos , Inflamación/inmunología , Inflamación/metabolismo , Inflamación/patología , Mediadores de Inflamación/metabolismo , Redes y Vías Metabólicas , Estructura Molecular , Estrés Oxidativo , Receptores Activados del Proliferador del Peroxisoma/metabolismo , Fosfolípidos/química , Fosfolípidos/farmacología , Estallido Respiratorio/efectos de los fármacos , Estallido Respiratorio/inmunología , Transducción de Señal , Receptores Toll-Like/antagonistas & inhibidores , Receptores Toll-Like/metabolismoRESUMEN
DCs are able to undergo rapid maturation, which subsequently allows them to initiate and orchestrate T cell-driven immune responses. DC maturation must be tightly controlled in order to avoid random T cell activation and development of autoimmunity. Here, we determined that 12/15-lipoxygenase-meditated (12/15-LO-mediated) enzymatic lipid oxidation regulates DC activation and fine-tunes consecutive T cell responses. Specifically, 12/15-LO activity determined the DC activation threshold via generation of phospholipid oxidation products that induced an antioxidative response dependent on the transcription factor NRF2. Deletion of the 12/15-LO-encoding gene or pharmacologic inhibition of 12/15-LO in murine or human DCs accelerated maturation and shifted the cytokine profile, thereby favoring the differentiation of Th17 cells. Exposure of 12/15-LO-deficient DCs to 12/15-LO-derived oxidized phospholipids attenuated both DC activation and the development of Th17 cells. Analysis of lymphatic tissues from 12/15-LO-deficient mice confirmed enhanced maturation of DCs as well as an increased differentiation of Th17 cells. Moreover, experimental autoimmune encephalomyelitis in mice lacking 12/15-LO resulted in an exacerbated Th17-driven autoimmune disease. Together, our data reveal that 12/15-LO controls maturation of DCs and implicate enzymatic lipid oxidation in shaping the adaptive immune response.