RESUMEN

Aquaporin-0 (AQP0) tetramers form square arrays in lens membranes through a yet unknown mechanism, but lens membranes are enriched in sphingomyelin and cholesterol. Here, we determined electron crystallographic structures of AQP0 in sphingomyelin/cholesterol membranes and performed molecular dynamics (MD) simulations to establish that the observed cholesterol positions represent those seen around an isolated AQP0 tetramer and that the AQP0 tetramer largely defines the location and orientation of most of its associated cholesterol molecules. At a high concentration, cholesterol increases the hydrophobic thickness of the annular lipid shell around AQP0 tetramers, which may thus cluster to mitigate the resulting hydrophobic mismatch. Moreover, neighboring AQP0 tetramers sandwich a cholesterol deep in the center of the membrane. MD simulations show that the association of two AQP0 tetramers is necessary to maintain the deep cholesterol in its position and that the deep cholesterol increases the force required to laterally detach two AQP0 tetramers, not only due to protein-protein contacts but also due to increased lipid-protein complementarity. Since each tetramer interacts with four such 'glue' cholesterols, avidity effects may stabilize larger arrays. The principles proposed to drive AQP0 array formation could also underlie protein clustering in lipid rafts.

Asunto(s)

Acuaporinas , Colesterol , Microdominios de Membrana , Simulación de Dinámica Molecular , Esfingomielinas , Colesterol/metabolismo , Colesterol/química , Acuaporinas/química , Acuaporinas/metabolismo , Microdominios de Membrana/metabolismo , Microdominios de Membrana/química , Esfingomielinas/química , Esfingomielinas/metabolismo , Animales , Proteínas del Ojo/química , Proteínas del Ojo/metabolismo , Multimerización de Proteína , Cristalino/química , Cristalino/metabolismo , Conformación Proteica

RESUMEN

Pancreatic cancer is a very aggressive disease with a dismal prognosis. The tumor microenvironment exerts immunosuppressive activities through the secretion of several cytokines, including interleukin (IL)-1. The IL-1/IL-1 receptor (IL-1R) axis is a key regulator in tumor-promoting T helper (Th)2- and Th17-type inflammation. Th2 cells are differentiated by dendritic cells endowed with Th2-polarizing capability by the thymic stromal lymphopoietin (TSLP) that is secreted by IL-1-activated cancer-associated fibroblasts (CAFs). Th17 cells are differentiated in the presence of IL-1 and other IL-1-regulated cytokines. In pancreatic cancer, the use of a recombinant IL-1R antagonist (IL1RA, anakinra, ANK) in in vitro and in vivo models has shown efficacy in targeting the IL-1/IL-1R pathway. In this study, we have developed sphingomyelin nanosystems (SNs) loaded with ANK (ANK-SNs) to compare their ability to inhibit Th2- and Th17-type inflammation with that of the free drug in vitro. We found that ANK-SNs inhibited TSLP and other pro-tumor cytokines released by CAFs at levels similar to ANK. Importantly, inhibition of IL-17 secretion by Th17 cells, but not of interferon-γ, was significantly higher, and at lower concentrations, with ANK-SNs compared to ANK. Collectively, the use of ANK-SNs might be beneficial in reducing the effective dose of the drug and its toxic effects.

Asunto(s)

Proteína Antagonista del Receptor de Interleucina 1 , Interleucina-1 , Neoplasias Pancreáticas , Esfingomielinas , Neoplasias Pancreáticas/tratamiento farmacológico , Neoplasias Pancreáticas/metabolismo , Neoplasias Pancreáticas/patología , Humanos , Proteína Antagonista del Receptor de Interleucina 1/farmacología , Interleucina-1/metabolismo , Esfingomielinas/metabolismo , Citocinas/metabolismo , Línea Celular Tumoral , Inflamación/tratamiento farmacológico , Inflamación/metabolismo , Inflamación/patología , Células Th17/inmunología , Células Th17/efectos de los fármacos , Células Th17/metabolismo , Células Th2/inmunología , Células Th2/efectos de los fármacos , Células Th2/metabolismo , Microambiente Tumoral/efectos de los fármacos , Nanopartículas/química , Fibroblastos Asociados al Cáncer/metabolismo , Fibroblastos Asociados al Cáncer/efectos de los fármacos

RESUMEN

Azoospermia, the absence of sperm cells in semen, affects around 15% of infertile males. Sertoli cell-only syndrome (SCOS) is the most common pathological lesion in the background of non-obstructive azoospermia and is characterised by the complete absence of germinal epithelium, with Sertoli cells exclusively present in the seminiferous tubules. Studies have shown a correlation between successful spermatogenesis and male fertility with lipid composition of spermatozoa, semen, seminal plasma or testis. The aim of this research was to discover the correlation between the Johnsen scoring system and phospholipid expressions in testicular cryosections of SCOS patients. MALDI imaging mass spectrometry is used to determine spatial distributions of molecular species, such as phospholipids. Phosphatidylcholines (PCs), phosphatidylethanolamines (PEs) and sphingomyelins (SMs) are the most abundant phospholipids in mammalian cells and testis. SMs, the structural components of plasma membranes, are crucial for spermatogenesis and sperm function. Plasmalogens, are unique PCs in testis with strong antioxidative properties. This study, using imaging mass spectrometry, demonstrates the local distribution of phospholipids, particularly SMs, PCs, plasmalogens and PEs in human testicular samples with SCOS for the first time. This study found a strong relationship between the Johnsen scoring system and phospholipid expression levels in human testicular tissues. Future findings could enable routine diagnostic techniques during microTESE procedures for successful sperm extraction.

Asunto(s)

Síndrome de Sólo Células de Sertoli , Espectrometría de Masa por Láser de Matriz Asistida de Ionización Desorción , Testículo , Masculino , Humanos , Espectrometría de Masa por Láser de Matriz Asistida de Ionización Desorción/métodos , Testículo/metabolismo , Testículo/patología , Síndrome de Sólo Células de Sertoli/metabolismo , Síndrome de Sólo Células de Sertoli/patología , Fosfolípidos/metabolismo , Espermatogénesis , Azoospermia/metabolismo , Azoospermia/patología , Esfingomielinas/metabolismo , Lípidos/análisis , Adulto , Espermatozoides/metabolismo , Espermatozoides/patología

RESUMEN

Numerous lipids are heterogeneously distributed among organelles. Most lipid trafficking between organelles is achieved by a group of lipid transfer proteins (LTPs) that carry lipids using their hydrophobic cavities. The human genome encodes many intracellular LTPs responsible for lipid trafficking and the function of many LTPs in defining cellular lipid levels and distributions is unclear. Here, we created a gene knockout library targeting 90 intracellular LTPs and performed whole-cell lipidomics analysis. This analysis confirmed known lipid disturbances and identified new ones caused by the loss of LTPs. Among these, we found major sphingolipid imbalances in ORP9 and ORP11 knockout cells, two proteins of previously unknown function in sphingolipid metabolism. ORP9 and ORP11 form a heterodimer to localize at the ER-trans-Golgi membrane contact sites, where the dimer exchanges phosphatidylserine (PS) for phosphatidylinositol-4-phosphate (PI(4)P) between the two organelles. Consequently, loss of either protein causes phospholipid imbalances in the Golgi apparatus that result in lowered sphingomyelin synthesis at this organelle. Overall, our LTP knockout library toolbox identifies various proteins in control of cellular lipid levels, including the ORP9-ORP11 heterodimer, which exchanges PS and PI(4)P at the ER-Golgi membrane contact site as a critical step in sphingomyelin synthesis in the Golgi apparatus.

Asunto(s)

Retículo Endoplásmico , Esfingomielinas , Esfingomielinas/metabolismo , Esfingomielinas/biosíntesis , Humanos , Retículo Endoplásmico/metabolismo , Aparato de Golgi/metabolismo , Proteínas Portadoras/metabolismo , Proteínas Portadoras/genética , Multimerización de Proteína , Receptores de Esteroides/metabolismo , Receptores de Esteroides/genética , Técnicas de Inactivación de Genes , Fosfatos de Fosfatidilinositol/metabolismo , Fosfatos de Fosfatidilinositol/biosíntesis

RESUMEN

Sphingomyelin is a key molecule of sphingolipid metabolism, and its enzymatic breakdown is associated with various infectious diseases. Here, we introduce trifunctional sphingomyelin derivatives that enable the visualization of sphingomyelin distribution and sphingomyelinase activity in infection processes. We demonstrate this by determining the activity of a bacterial sphingomyelinase on the plasma membrane of host cells using a combination of Förster resonance energy transfer and expansion microscopy. We further use our trifunctional sphingomyelin probes to visualize their metabolic state during infections with Chlamydia trachomatis and thereby show that chlamydial inclusions primarily contain the cleaved forms of the molecules. Using expansion microscopy, we observe that the proportion of metabolized molecules increases during maturation from reticulate to elementary bodies, indicating different membrane compositions between the two chlamydial developmental forms. Expansion microscopy of trifunctional sphingomyelins thus provides a powerful microscopy tool to analyze sphingomyelin metabolism in cells at nanoscale resolution.

Asunto(s)

Membrana Celular , Chlamydia trachomatis , Esfingomielina Fosfodiesterasa , Esfingomielinas , Esfingomielinas/metabolismo , Esfingomielina Fosfodiesterasa/metabolismo , Chlamydia trachomatis/metabolismo , Humanos , Membrana Celular/metabolismo , Transferencia Resonante de Energía de Fluorescencia/métodos , Células HeLa , Infecciones por Chlamydia/metabolismo , Infecciones por Chlamydia/microbiología , Microscopía/métodos

RESUMEN

BACKGROUND: Glycerophospholipids (GPLs) are essential for cell membrane structure and function. Sphingomyelin and its metabolites regulate cell growth, apoptosis, and stress responses. This study aimed to investigate lipid metabolism in patients experiencing sudden sensorineural hearing loss across all frequencies (AF-SSNHL). METHODS: The study included 60 patients diagnosed with unilateral AF-SSNHL, among whom 30 patients had a level of hearing improvement ≥ 15 dB after 6 months of follow-up. A propensity score-matched (2:1) control group was used. Liquid chromatography‒mass spectrometry based untargeted lipidomics analysis combined with multivariate statistics was performed to investigate the lipids change. The "lipidome" R package and weighted gene co-expression network analysis (WGCNA) were utilised to assess the lipids' structural features and the association between lipids and hearing. RESULTS: Lipidomics successfully differentiated the AF-SSNHL group from the control group, identifying 17 risk factors, mainly including phosphatidylcholine (PC), phosphatidylethanolamine (PE), and related metabolites. The ratios of lysophosphatidylcholine/PC, lysophosphatidylethanolamine/PE, and lysodimethylphosphatidylethanolamine/PE were upregulated, while some glycerophospholipid (GPL)-plasmalogens were downregulated in the AF-SSNHL group, indicating abnormal metabolism of GPLs. Trihexosylceramide (d34:1), PE (18:1e_22:5), and sphingomyelin (d40:3) were significantly different between responders and nonresponders, and positively correlated with hearing improvement. Additionally, the results of the WGCNA also suggested that partial GPL-plasmalogens were positively associated with hearing improvement. CONCLUSION: AF-SSNHL patients exhibited abnormally high blood lipids and pronounced GPLs metabolic abnormalities. Sphingolipids and GPL-plasmalogens had an association with the level of hearing improvement. By understanding the lipid changes, clinicians may be able to predict the prognosis of hearing recovery and personalize treatment approaches.

Asunto(s)

Biomarcadores , Pérdida Auditiva Sensorineural , Metabolismo de los Lípidos , Lipidómica , Humanos , Femenino , Masculino , Persona de Mediana Edad , Biomarcadores/sangre , Pérdida Auditiva Sensorineural/sangre , Adulto , Pérdida Auditiva Súbita/sangre , Glicerofosfolípidos/sangre , Anciano , Fosfatidiletanolaminas/sangre , Fosfatidiletanolaminas/metabolismo , Fosfatidilcolinas/sangre , Fosfatidilcolinas/metabolismo , Lisofosfatidilcolinas/sangre , Esfingomielinas/sangre , Esfingomielinas/metabolismo , Lisofosfolípidos

RESUMEN

Sphingomyelin (SM) is a major sphingolipid in mammalian cells. SM is enriched in the extracellular leaflet of the plasma membrane (PM). Besides this localization, recent electron microscopic and biochemical studies suggest the presence of SM in the cytosolic leaflet of the PM. In the present study, we generated a non-toxic SM-binding variant (NT-EqtII) based on equinatoxin-II (EqtII) from the sea anemone Actinia equina, and examined the dynamics of SM in the cytosolic leaflet of living cell PMs. NT-EqtII with two point mutations (Leu26Ala and Pro81Ala) had essentially the same specificity and affinity to SM as wild-type EqtII. NT-EqtII expressed in the cytosol was recruited to the PM in various cell lines. Super-resolution microscopic observation revealed that NT-EqtII formed tiny domains that were significantly colocalized with cholesterol and N-terminal Lyn. Meanwhile, single molecule observation at high resolutions down to 1 ms revealed that all the examined lipid probes including NT-EqtII underwent apparent fast simple Brownian diffusion, exhibiting that SM and other lipids in the cytosolic leaflet rapidly moved in and out of domains. Thus, the novel SM-binding probe demonstrated the presence of the raft-like domain in the cytosolic leaflet of living cell PMs.

Asunto(s)

Membrana Celular , Venenos de Cnidarios , Citosol , Esfingomielinas , Esfingomielinas/metabolismo , Membrana Celular/metabolismo , Citosol/metabolismo , Animales , Venenos de Cnidarios/metabolismo , Venenos de Cnidarios/genética , Humanos , Anémonas de Mar/metabolismo , Anémonas de Mar/genética , Colesterol/metabolismo

RESUMEN

Sphingomyelin is postulated to form clusters with glycosphingolipids, cholesterol and other sphingomyelin molecules in biomembranes through hydrophobic interaction and hydrogen bonds. These clusters form submicron size lipid domains. Proteins that selectively binds sphingomyelin and/or cholesterol are useful to visualize the lipid domains. Due to their small size, visualization of lipid domains requires advanced microscopy techniques in addition to lipid binding proteins. This Chapter describes the method to characterize plasma membrane sphingomyelin-rich and cholesterol-rich lipid domains by quantitative microscopy. This Chapter also compares different permeabilization methods to visualize intracellular lipid domains.

Asunto(s)

Colesterol , Esfingomielinas , Esfingomielinas/química , Esfingomielinas/metabolismo , Colesterol/química , Colesterol/metabolismo , Humanos , Animales , Microdominios de Membrana/metabolismo , Microdominios de Membrana/química , Microscopía/métodos , Proteínas Portadoras/química , Proteínas Portadoras/metabolismo , Membrana Celular/metabolismo , Membrana Celular/química

RESUMEN

Measurements of sphingolipid metabolism are most accurately performed by LC-MS. However, this technique is expensive, not widely accessible, and without the use of specific probes, it does not provide insight into metabolic flux through the pathway. Employing the fluorescent ceramide analogue NBD-C6-ceramide as a tracer in intact cells, we developed a comprehensive HPLC-based method that simultaneously measures the main nodes of ceramide metabolism in the Golgi. Hence, by quantifying the conversion of NBD-C6-ceramide to NBD-C6-sphingomyelin, NBD-C6-hexosylceramides, and NBD-C6-ceramide-1-phosphate (NBD-C1P), the activities of Golgi resident enzymes sphingomyelin synthase 1, glucosylceramide synthase, and ceramide kinase (CERK) could be measured simultaneously. Importantly, the detection of NBD-C1P allowed us to quantify CERK activity in cells, a usually difficult task. By applying this method, we evaluated the specificity of commonly used sphingolipid inhibitors and discovered that 1-phenyl-2-decanoylamino-3-morpholino-1-propanol, which targets glucosylceramide synthase, and fenretinide (4HPR), an inhibitor for dihydroceramide desaturase, also suppress CERK activity. This study demonstrates the benefit of an expanded analysis of ceramide metabolism in the Golgi, and it provides a qualitative and easy-to-implement method.

Asunto(s)

Ceramidas , Glucosiltransferasas , Aparato de Golgi , Fosfotransferasas (Aceptor de Grupo Alcohol) , Esfingolípidos , Aparato de Golgi/metabolismo , Ceramidas/metabolismo , Esfingolípidos/metabolismo , Humanos , Glucosiltransferasas/antagonistas & inhibidores , Glucosiltransferasas/metabolismo , Fosfotransferasas (Aceptor de Grupo Alcohol)/metabolismo , Fosfotransferasas (Aceptor de Grupo Alcohol)/antagonistas & inhibidores , Inhibidores Enzimáticos/farmacología , 4-Cloro-7-nitrobenzofurazano/análogos & derivados , 4-Cloro-7-nitrobenzofurazano/metabolismo , Cromatografía Líquida de Alta Presión , Células HeLa , Hexosiltransferasas/metabolismo , Hexosiltransferasas/antagonistas & inhibidores , Esfingomielinas/metabolismo , Transferasas (Grupos de Otros Fosfatos Sustitutos)

RESUMEN

Lipids from cow milk fat globule membranes (MFGMs) and extracellular vesicles (EVs) are considered beneficial for neurodevelopment, cognitive maintenance and human health in general. Nevertheless, it is largely unknown whether intake of infant formulas and medical nutrition products rich in these particles promote accretion of specific lipids and whether this affects metabolic homeostasis. To address this, we carried out a 16-week dietary intervention study where mice were supplemented with a MFGM/EV-rich concentrate, a control diet supplemented with a whey protein concentrate and devoid of milk lipids, or regular chow. Assessment of commonly used markers of metabolic health, including body weight, glucose intolerance and liver microanatomy, demonstrated no differences across the dietary regimes. In contrast, in-depth lipidomic analysis revealed accretion of milk-derived very long odd-chain sphingomyelins and ceramides in blood plasma and multiple tissues of mice fed the MFGM/EV diet. Furthermore, lipidomic flux analysis uncovered that mice fed the MFGM/EV diet have increased lipid metabolic turnover at the whole-body level. These findings help fill a long-lasting knowledge gap between the intake of MFGM/EV-containing foods and the health-promoting effects of their lipid constituents. In addition, the findings suggest that dietary sphingomyelins or ceramide-breakdown products with very long-chains can be used as structural components of cellular membranes, lipoprotein particles and signaling molecules that modulate metabolic homeostasis and health.

Asunto(s)

Vesículas Extracelulares , Glucolípidos , Glicoproteínas , Gotas Lipídicas , Metabolismo de los Lípidos , Esfingolípidos , Animales , Esfingolípidos/metabolismo , Vesículas Extracelulares/metabolismo , Ratones , Glucolípidos/metabolismo , Gotas Lipídicas/metabolismo , Glicoproteínas/metabolismo , Lipidómica , Ratones Endogámicos C57BL , Masculino , Esfingomielinas/metabolismo , Ceramidas/metabolismo , Dieta , Hígado/metabolismo , Suplementos Dietéticos

RESUMEN

Ceramide (Cer) is synthesized de novo in the bilayer of the endoplasmic reticulum and transported to the cytosolic leaflet of the trans-Golgi apparatus for sphingomyelin (SM) synthesis. As the active site of SM synthase (SMS) is located on the luminal side of the Golgi membrane, Cer translocates to the lumen via transbilayer movement for SM synthesis. However, the mechanism of transbilayer movement is not fully understood. As the Cer-related translocases seem to localize near the SMS, the protein was identified using proximity-dependent biotin identification proteomics. Phospholipid scramblase 1 (PLSCR1), which is thought to act as a scramblase for phosphatidylserine and phosphatidylethanolamine, was identified as a protein proximal to the SMS isoforms SMS1 and SMS2. Although five isoforms of PLSCR have been reported in humans, only PLSCR1, PLSCR3, and PLSCR4 are expressed in HEK293T cells. Confocal microscopic analysis showed that PLSCR1 and PLSCR4 partially co-localized with p230, a trans-Golgi network marker, where SMS isoforms are localized. We established CRISPR/Cas9-mediated PLSCR1, PLSCR3, and PLSCR4 single-knockout cells and PLSCR1, 3, 4 triple knockout HEK293T cells. Liquid chromatography-tandem mass spectrometry revealed that the levels of species with distinct acyl chains in Cer and SM were not significantly different in single knockout cells or in the triple knockout cells compared to the wild-type cells. Our findings suggest that PLSCR1 is localized in the vicinity of SMS isoforms, however is not involved in the transbilayer movement of Cer for SM synthesis.

Asunto(s)

Proteínas de Transferencia de Fosfolípidos , Esfingomielinas , Transferasas (Grupos de Otros Fosfatos Sustitutos) , Humanos , Proteínas de Transferencia de Fosfolípidos/metabolismo , Proteínas de Transferencia de Fosfolípidos/genética , Transferasas (Grupos de Otros Fosfatos Sustitutos)/metabolismo , Transferasas (Grupos de Otros Fosfatos Sustitutos)/genética , Células HEK293 , Esfingomielinas/metabolismo , Esfingomielinas/biosíntesis , Proteínas de la Membrana/metabolismo , Proteínas de la Membrana/genética , Isoenzimas/metabolismo , Isoenzimas/genética , Aparato de Golgi/metabolismo , Aparato de Golgi/enzimología

RESUMEN

BACKGROUND: Traumatic brain injury (TBI) causes neuroinflammation and can lead to long-term neurological dysfunction, even in cases of mild TBI (mTBI). Despite the substantial burden of this disease, the management of TBI is precluded by an incomplete understanding of its cellular mechanisms. Sphingolipids (SPL) and their metabolites have emerged as key orchestrators of biological processes related to tissue injury, neuroinflammation, and inflammation resolution. No study so far has investigated comprehensive sphingolipid profile changes immediately following TBI in animal models or human cases. In this study, sphingolipid metabolite composition was examined during the acute phases in brain tissue and plasma of mice following mTBI. METHODS: Wildtype mice were exposed to air-blast-mediated mTBI, with blast exposure set at 50-psi on the left cranium and 0-psi designated as Sham. Sphingolipid profile was analyzed in brain tissue and plasma during the acute phases of 1, 3, and 7 days post-TBI via liquid-chromatography-mass spectrometry. Simultaneously, gene expression of sphingolipid metabolic markers within brain tissue was analyzed using quantitative reverse transcription-polymerase chain reaction. Significance (P-values) was determined by non-parametric t-test (Mann-Whitney test) and by Tukey's correction for multiple comparisons. RESULTS: In post-TBI brain tissue, there was a significant elevation of 1) acid sphingomyelinase (aSMase) at 1- and 3-days, 2) neutral sphingomyelinase (nSMase) at 7-days, 3) ceramide-1-phosphate levels at 1 day, and 4) monohexosylceramide (MHC) and sphingosine at 7-days. Among individual species, the study found an increase in C18:0 and a decrease in C24:1 ceramides (Cer) at 1 day; an increase in C20:0 MHC at 3 days; decrease in MHC C18:0 and increase in MHC C24:1, sphingomyelins (SM) C18:0, and C24:0 at 7 days. Moreover, many sphingolipid metabolic genes were elevated at 1 day, followed by a reduction at 3 days and an absence at 7-days post-TBI. In post-TBI plasma, there was 1) a significant reduction in Cer and MHC C22:0, and an increase in MHC C16:0 at 1 day; 2) a very significant increase in long-chain Cer C24:1 accompanied by significant decreases in Cer C24:0 and C22:0 in MHC and SM at 3 days; and 3) a significant increase of C22:0 in all classes of SPL (Cer, MHC and SM) as well as a decrease in Cer C24:1, MHC C24:1 and MHC C24:0 at 7 days. CONCLUSIONS: Alterations in sphingolipid metabolite composition, particularly sphingomyelinases and short-chain ceramides, may contribute to the induction and regulation of neuroinflammatory events in the early stages of TBI, suggesting potential targets for novel diagnostic, prognostic, and therapeutic strategies in the future.

Asunto(s)

Encéfalo , Ceramidas , Esfingolípidos , Esfingomielina Fosfodiesterasa , Esfingosina , Animales , Ratones , Esfingolípidos/sangre , Esfingolípidos/metabolismo , Encéfalo/metabolismo , Encéfalo/patología , Ceramidas/sangre , Ceramidas/metabolismo , Esfingomielina Fosfodiesterasa/metabolismo , Esfingomielina Fosfodiesterasa/sangre , Esfingomielina Fosfodiesterasa/genética , Esfingosina/análogos & derivados , Esfingosina/sangre , Esfingosina/metabolismo , Modelos Animales de Enfermedad , Masculino , Esfingomielinas/sangre , Esfingomielinas/metabolismo , Conmoción Encefálica/sangre , Conmoción Encefálica/metabolismo , Ratones Endogámicos C57BL , Lesiones Traumáticas del Encéfalo/metabolismo , Lesiones Traumáticas del Encéfalo/sangre , Lesiones Traumáticas del Encéfalo/patología , Lisofosfolípidos/sangre , Lisofosfolípidos/metabolismo

RESUMEN

Various lipid transfer proteins (LTPs) mediate the inter-organelle transport of lipids. By working at membrane contact zones between donor and acceptor organelles, LTPs achieve rapid and accurate inter-organelle transfer of lipids. This article will describe the emerging paradigm that the action of LTPs at organelle contact zones generates metabolic channeling events in lipid metabolism, mainly referring to how ceramide synthesized in the endoplasmic reticulum is preferentially metabolized to sphingomyelin in the distal Golgi region, how cholesterol and phospholipids receive specific metabolic reactions in mitochondria, and how the hijacking of host LTPs by intracellular pathogens may generate new channeling-like events. In addition, the article will discuss how the function of LTPs is regulated, exemplified by a few representative LTP systems, and will briefly touch on experiments that will be necessary to establish the paradigm that LTP-mediated inter-organelle transport of lipids is one of the mechanisms of compartmentalization-based metabolic channeling events.

Asunto(s)

Metabolismo de los Lípidos , Mitocondrias , Humanos , Animales , Mitocondrias/metabolismo , Transporte Biológico , Retículo Endoplásmico/metabolismo , Aparato de Golgi/metabolismo , Proteínas Portadoras/metabolismo , Orgánulos/metabolismo , Ceramidas/metabolismo , Colesterol/metabolismo , Esfingomielinas/metabolismo , Fosfolípidos/metabolismo

RESUMEN

There is a phenotype of obese individuals termed metabolically healthy obese that present a reduced cardiometabolic risk. This phenotype offers a valuable model for investigating the mechanisms connecting obesity and metabolic alterations such as Type 2 Diabetes Mellitus (T2DM). Previously, in an untargeted metabolomics analysis in a cohort of morbidly obese women, we observed a different lipid metabolite pattern between metabolically healthy morbid obese individuals and those with associated T2DM. To validate these findings, we have performed a complementary study of lipidomics. In this study, we assessed a liquid chromatography coupled to a mass spectrometer untargeted lipidomic analysis on serum samples from 209 women, 73 normal-weight women (control group) and 136 morbid obese women. From those, 65 metabolically healthy morbid obese and 71 with associated T2DM. In this work, we find elevated levels of ceramides, sphingomyelins, diacyl and triacylglycerols, fatty acids, and phosphoethanolamines in morbid obese vs normal weight. Conversely, decreased levels of acylcarnitines, bile acids, lyso-phosphatidylcholines, phosphatidylcholines (PC), phosphatidylinositols, and phosphoethanolamine PE (O-38:4) were noted. Furthermore, comparing morbid obese women with T2DM vs metabolically healthy MO, a distinct lipid profile emerged, featuring increased levels of metabolites: deoxycholic acid, diacylglycerol DG (36:2), triacylglycerols, phosphatidylcholines, phosphoethanolamines, phosphatidylinositols, and lyso-phosphatidylinositol LPI (16:0). To conclude, analysing both comparatives, we observed decreased levels of deoxycholic acid, PC (34:3), and PE (O-38:4) in morbid obese women vs normal-weight. Conversely, we found elevated levels of these lipids in morbid obese women with T2DM vs metabolically healthy MO. These profiles of metabolites could be explored for the research as potential markers of metabolic risk of T2DM in morbid obese women.

Asunto(s)

Diabetes Mellitus Tipo 2 , Lipidómica , Obesidad Mórbida , Humanos , Diabetes Mellitus Tipo 2/metabolismo , Diabetes Mellitus Tipo 2/sangre , Diabetes Mellitus Tipo 2/complicaciones , Femenino , Obesidad Mórbida/sangre , Obesidad Mórbida/metabolismo , Obesidad Mórbida/complicaciones , Lipidómica/métodos , Persona de Mediana Edad , Adulto , Lípidos/sangre , Metabolómica/métodos , Estudios de Casos y Controles , Triglicéridos/sangre , Esfingomielinas/sangre , Esfingomielinas/metabolismo , Ceramidas/sangre , Ceramidas/metabolismo , Metabolismo de los Lípidos

RESUMEN

The metabolites and microbiota in tongue coating display distinct characteristics in certain digestive disorders, yet their relationship with colorectal cancer (CRC) remains unexplored. Here, we employed liquid chromatography coupled with tandem mass spectrometry to analyze the lipid composition of tongue coating using a nontargeted approach in 30 individuals with colorectal adenomas (CRA), 32 with CRC, and 30 healthy controls (HC). We identified 21 tongue coating lipids that effectively distinguished CRC from HC (AUC = 0.89), and 9 lipids that differentiated CRC from CRA (AUC = 0.9). Furthermore, we observed significant alterations in the tongue coating lipid composition in the CRC group compared to HC/CRA groups. As the adenoma-cancer sequence progressed, there was an increase in long-chain unsaturated triglycerides (TG) levels and a decrease in phosphatidylethanolamine plasmalogen (PE-P) levels. Furthermore, we noted a positive correlation between N-acyl ornithine (NAOrn), sphingomyelin (SM), and ceramide phosphoethanolamine (PE-Cer), potentially produced by members of the Bacteroidetes phylum. The levels of inflammatory lipid metabolite 12-HETE showed a decreasing trend with colorectal tumor progression, indicating the potential involvement of tongue coating microbiota and tumor immune regulation in early CRC development. Our findings highlight the potential utility of tongue coating lipid analysis as a noninvasive tool for CRC diagnosis.

Asunto(s)

Neoplasias Colorrectales , Lipidómica , Fosfatidiletanolaminas , Espectrometría de Masas en Tándem , Lengua , Humanos , Neoplasias Colorrectales/metabolismo , Neoplasias Colorrectales/microbiología , Lipidómica/métodos , Masculino , Femenino , Lengua/microbiología , Lengua/metabolismo , Lengua/patología , Lengua/química , Persona de Mediana Edad , Espectrometría de Masas en Tándem/métodos , Fosfatidiletanolaminas/metabolismo , Fosfatidiletanolaminas/análisis , Anciano , Cromatografía Liquida , Lípidos/análisis , Lípidos/química , Triglicéridos/metabolismo , Triglicéridos/análisis , Adenoma/metabolismo , Adenoma/microbiología , Esfingomielinas/análisis , Esfingomielinas/metabolismo , Ácido 12-Hidroxi-5,8,10,14-Eicosatetraenoico/metabolismo , Ácido 12-Hidroxi-5,8,10,14-Eicosatetraenoico/química , Plasmalógenos/análisis , Plasmalógenos/metabolismo , Plasmalógenos/química , Estudios de Casos y Controles , Etanolaminas/metabolismo , Etanolaminas/análisis , Etanolaminas/química , Ceramidas/metabolismo , Ceramidas/análisis , Adulto

RESUMEN

Cell membrane stiffness is critical for cellular function, with cholesterol and sphingomyelin as pivot contributors. Current methods for measuring membrane stiffness are often invasive, ex situ, and slow in process, prompting the need for innovative techniques. Here, we present a fluorescence resonance energy transfer (FRET)-based protein sensor designed to address these challenges. The sensor consists of two fluorescent units targeting sphingomyelin and cholesterol, connected by a linker that responds to the proximity of these lipids. In rigid membranes, cholesterol and sphingomyelin are in close proximity, leading to an increased FRET signal. We utilized this sensor in combination with confocal microscopy to explore changes in plasma membrane stiffness under various conditions, including differences in osmotic pressure, the presence of reactive oxygen species (ROS) and variations in substrate stiffness. Furthermore, we explored the impact of SARS-CoV-2 on membrane stiffness and the distribution of ACE2 after attachment to the cell membrane. This tool offers substantial potential for future investigations in the field of mechanobiology.

Asunto(s)

Membrana Celular , Colesterol , Transferencia Resonante de Energía de Fluorescencia , SARS-CoV-2 , Esfingomielinas , Transferencia Resonante de Energía de Fluorescencia/métodos , Humanos , Membrana Celular/metabolismo , Membrana Celular/química , Esfingomielinas/análisis , Esfingomielinas/metabolismo , Colesterol/análisis , Colesterol/metabolismo , Microscopía Confocal/métodos , Especies Reactivas de Oxígeno/metabolismo , Especies Reactivas de Oxígeno/análisis , COVID-19/virología , Enzima Convertidora de Angiotensina 2/metabolismo , Técnicas Biosensibles/métodos

RESUMEN

Adipogenesis is one of the major mechanisms for adipose tissue expansion, during which spindle-shaped mesenchymal stem cells commit to the fate of adipocyte precursors and differentiate into round-shaped fat-laden adipocytes. Here, we investigated the lipidomic profile dynamics of ex vivo-differentiated brown and white adipocytes derived from the stromal vascular fractions of interscapular brown (iBAT) and inguinal white adipose tissues. We showed that sphingomyelin was specifically enriched in terminally differentiated brown adipocytes, but not white adipocytes. In line with this, freshly isolated adipocytes of iBAT showed higher sphingomyelin content than those of inguinal white adipose tissue. Upon cold exposure, sphingomyelin abundance in iBAT gradually decreased in parallel with reduced sphingomyelin synthase 1 protein levels. Cold-exposed animals treated with an inhibitor of sphingomyelin hydrolases failed to maintain core body temperature and showed reduced oxygen consumption and iBAT UCP1 levels. Conversely, blockade of sphingomyelin synthetic enzymes resulted in enhanced nonshivering thermogenesis, reflected by elevated body temperature and UCP1 levels. Taken together, our results uncovered a relation between sphingomyelin abundance and fine-tuning of UCP1-mediated nonshivering thermogenesis.

Asunto(s)

Esfingomielinas , Termogénesis , Proteína Desacopladora 1 , Animales , Proteína Desacopladora 1/metabolismo , Proteína Desacopladora 1/genética , Esfingomielinas/metabolismo , Ratones , Masculino , Tejido Adiposo Blanco/metabolismo , Tejido Adiposo Pardo/metabolismo , Ratones Endogámicos C57BL

RESUMEN

Cholesterol-rich lipid rafts are found to facilitate membrane fusion, central to processes like viral entry, fertilization, and neurotransmitter release. While the fusion process involves local, transient membrane dehydration, the impact of reduced hydration on cholesterol's structural organization in biological membranes remains unclear. Here, we employ confocal fluorescence microscopy and atomistic molecular dynamics simulations to investigate cholesterol behavior in phase-separated lipid bilayers under controlled hydration. We unveiled that dehydration prompts cholesterol release from raft-like domains into the surrounding fluid phase. Unsaturated phospholipids undergo more significant dehydration-induced structural changes and lose more hydrogen bonds with water than sphingomyelin. The results suggest that cholesterol redistribution is driven by the equalization of biophysical properties between phases and the need to satisfy lipid hydrogen bonds. This underscores the role of cholesterol-phospholipid-water interplay in governing cholesterol affinity for a specific lipid type, providing a new perspective on the regulatory role of cell membrane heterogeneity during membrane fusion.

Asunto(s)

Colesterol , Membrana Dobles de Lípidos , Simulación de Dinámica Molecular , Agua , Colesterol/química , Colesterol/metabolismo , Membrana Dobles de Lípidos/química , Membrana Dobles de Lípidos/metabolismo , Agua/química , Agua/metabolismo , Microdominios de Membrana/química , Microdominios de Membrana/metabolismo , Enlace de Hidrógeno , Esfingomielinas/química , Esfingomielinas/metabolismo , Fusión de Membrana , Fosfolípidos/química , Fosfolípidos/metabolismo

RESUMEN

Sphingomyelin (SM) has key roles in modulating mammalian membrane properties and serves as an important pool for bioactive molecules. SM biosynthesis is mediated by the sphingomyelin synthase (SMS) family, comprising SMS1, SMS2 and SMS-related (SMSr) members. Although SMS1 and SMS2 exhibit SMS activity, SMSr possesses ceramide phosphoethanolamine synthase activity. Here we determined the cryo-electron microscopic structures of human SMSr in complexes with ceramide, diacylglycerol/phosphoethanolamine and ceramide/phosphoethanolamine (CPE). The structures revealed a hexameric arrangement with a reaction chamber located between the transmembrane helices. Within this structure, a catalytic pentad E-H/D-H-D was identified, situated at the interface between the lipophilic and hydrophilic segments of the reaction chamber. Additionally, the study unveiled the two-step synthesis process catalyzed by SMSr, involving PE-PLC (phosphatidylethanolamine-phospholipase C) hydrolysis and the subsequent transfer of the phosphoethanolamine moiety to ceramide. This research provides insights into the catalytic mechanism of SMSr and expands our understanding of sphingolipid metabolism.

Asunto(s)

Microscopía por Crioelectrón , Modelos Moleculares , Esfingomielinas , Transferasas (Grupos de Otros Fosfatos Sustitutos) , Humanos , Transferasas (Grupos de Otros Fosfatos Sustitutos)/metabolismo , Transferasas (Grupos de Otros Fosfatos Sustitutos)/química , Esfingomielinas/metabolismo , Esfingomielinas/química , Esfingomielinas/biosíntesis , Ceramidas/metabolismo , Ceramidas/química , Etanolaminas/metabolismo , Etanolaminas/química , Fosfatidiletanolaminas/metabolismo , Fosfatidiletanolaminas/química , Diglicéridos/metabolismo , Diglicéridos/química , Proteínas del Tejido Nervioso/metabolismo , Proteínas del Tejido Nervioso/química , Proteínas de la Membrana

RESUMEN

Hormone treatments are frequently associated with cardiovascular diseases and cancers in women. Additionally, the detrimental effects of their presence as contaminants in water remain a concern. The transport of hormones through cell membranes is essential for their biological action, but investigating cell permeability is challenging owing to the experimental difficulty in dealing with whole cells. In this paper, we study the interaction of the synthetic hormone 17α-ethynylestradiol (EE2) with membrane models containing the key raft components sphingomyelin (SM) and cholesterol (Chol). The models consisted of Langmuir monolayers and giant unilamellar vesicles (GUVs) that represent bilayers. EE2 induced expansion of SM monolayers upon interacting with the non-hydrated amide group of SM head, but it had practically no effect on SM GUVs because these group are not available for interaction in bilayers. In contrast, EE2 interacted with hydrated phosphate group (PO2-) and amide group of SM/Chol mixture monolayer, which could explain the loss in phase contrast of liquid-ordered GUVs suggesting pore formation. A comparison with reported EE2 effects on GUVs in the fluid phase, for which no loss in phase contrast was observed, indicates that the liquid-ordered phase consisting of lipid rafts is relevant to be associated with the changes on cell permeability caused by the hormones.

Asunto(s)

Esfingomielinas , Liposomas Unilamelares , Femenino , Humanos , Esfingomielinas/metabolismo , Hormonas , Colesterol , Microdominios de Membrana/metabolismo , Amidas