Glia are of vital importance for all complex nervous system. One of the many functions of glia is to insulate and provide trophic and metabolic support to axons. Here, using glial-specific RNAi ...knockdown in Drosophila, we silenced 6930 conserved genes in adult flies to identify essential genes and pathways. Among our screening hits, metabolic processes were highly represented, and genes involved in carbohydrate and lipid metabolic pathways appeared to be essential in glia. One critical pathway identified was de novo ceramide synthesis. Glial knockdown of lace, a subunit of the serine palmitoyltransferase associated with hereditary sensory and autonomic neuropathies in humans, resulted in ensheathment defects of peripheral nerves in Drosophila. A genetic dissection study combined with shotgun high-resolution mass spectrometry of lipids showed that levels of ceramide phosphoethanolamine are crucial for axonal ensheathment by glia. A detailed morphological and functional analysis demonstrated that the depletion of ceramide phosphoethanolamine resulted in axonal defasciculation, slowed spike propagation, and failure of wrapping glia to enwrap peripheral axons. Supplementing sphingosine into the diet rescued the neuropathy in flies. Thus, our RNAi study in Drosophila identifies a key role of ceramide phosphoethanolamine in wrapping of axons by glia.
•The lack of TLR 4 increases the neutral sphingomyelinase expression and activity.•Neutral sphingomyelinase markedly delocalizes in the absence of TLR4.•TLR4 deficiency also affects both heavy ...neurofilaments and glial fibrillary protein.
Both sphingomyelinase and Toll-Like Receptor 4 (TLR4) are implicated in neurodegenerative diseases. However, the relationship between the two molecules remains unclear. In this study, using WT and TLR4-deficient mice, treated or not with 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), we aimed to investigate the relation between TLR4 and neutral sphingomyelinase (nSMase) in the midbrain. We found that the lack of TLR4 caused increase in nSMase protein expression and enzyme activity in the midbrain, as well as a marked delocalization from the cell membranes. This provoked a decrease in sphingomyelin (SM) species and an increase in ceramide levels. We found that exposure of TLR4-deficient mice to MPTP reduces unsaturated SM species by increasing saturated/unsaturated SM ratio. Saturated fatty acid make SM more rigid and could contribute to reducing neural plasticity. In this study we showed that the absence of TLR4 also induced reduction of both heavy neurofilaments and glial fibrillary acidic protein (GFAP) and mice exhibited higher sensitivity to MPTP administration. We speculated about the possible association between nSMase-TLR4 complex and MPTP midbrain damage. Taken together, our findings provide for the first time indications about the role of TLR4 in change of SM metabolism in MPTP neurotoxicity.
Lysosomes are degradative organelles that facilitate the removal and recycling of potentially cytotoxic materials and mediate a variety of other cellular processes, such as nutrient sensing, ...intracellular signaling, and lipid metabolism. Due to these central roles, lysosome dysfunction can lead to deleterious outcomes, including the accumulation of cytotoxic material, inflammation, and cell death. We previously reported that cationic amphiphilic drugs, such as imipramine, alter pH and lipid metabolism within macrophage lysosomes. Therefore, the ability for imipramine to induce changes to the lipid content of isolated macrophage lysosomes was investigated, focusing on sphingomyelin, cholesterol, and glycerophospholipid metabolism as these lipid classes have important roles in inflammation and disease. The lysosomes were isolated from control and imipramine-treated macrophages using density gradient ultracentrifugation, and mass spectrometry was used to measure the changes in their lipid composition. An unsupervised hierarchical cluster analysis revealed a clear differentiation between the imipramine-treated and control lysosomes. There was a significant overall increase in the abundance of specific lipids mostly composed of cholesterol esters, sphingomyelins, and phosphatidylcholines, while lysophosphatidylcholines and ceramides were overall decreased. These results support the conclusion that imipramine's ability to change the lysosomal pH inhibits multiple pH-sensitive enzymes in macrophage lysosomes.
Alcohol disturbs hepatic lipid synthesis and transport, but the role of lipid dysfunction in alcohol-related liver disease (ALD) is unclear. In this biopsy-controlled, prospective, observational ...study, we characterized the liver and plasma lipidomes in patients with early ALD.
We performed mass spectrometry–based lipidomics of paired liver and plasma samples from 315 patients with ALD and of plasma from 51 matched healthy controls. We associated lipid levels with histologic fibrosis, inflammation, and steatosis with correction for multiple testing and adjustment for confounders. We further investigated sphingolipid regulation by means of quantitative real-time polymerase chain reaction sequencing of microRNA, prediction of liver-related events, and tested causality with Mendelian randomization.
We detected 198 lipids in the liver and 236 lipids in the circulation from 18 lipid classes. Most sphingolipids (sphingomyelins and ceramides) and phosphocholines were co–down-regulated in both liver and plasma, where lower abundance correlated with higher fibrosis stage. Sphingomyelins showed the most pronounced negative correlation to fibrosis, mirrored by negative correlations in both liver and plasma with hepatic inflammation. Reduced sphingomyelins predicted future liver-related events. This seemed to be characteristic of “pure ALD,” as sphingomyelin levels were higher in patients with concomitant metabolic syndrome and ALD/nonalcoholic fatty liver disease overlap. Mendelian randomization in FinnGen and UK Biobanks indicated ALD as the cause of low sphingomyelins, and alcohol use disorder did not correlate with genetic susceptibility to low sphingomyelin levels.
Alcohol-related liver fibrosis is characterized by selective and progressive lipid depletion in liver and blood, particularly sphingomyelins, which also associates with progression to liver-related events.
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The abnormal composition of 18 types of lipid classes in the liver and blood in people with early alcohol-related liver disease is specific to the type of liver damage and has implications on the risk of developing symptomatic liver disease.
Antibodies to lipids are part of the first line of defense against microorganisms and regulate the pro/anti-inflammatory balance. Viruses modulate cellular lipid metabolism to enhance their ...replication, and some of these metabolites are proinflammatory. We hypothesized that antibodies to lipids would play a main role of in the defense against SARS-CoV-2 and thus, they would also avoid the hyperinflammation, a main problem in severe condition patients.
Serum samples from COVID-19 patients with mild and severe course, and control group were included. IgG and IgM to different glycerophospholipids and sphingolipids were analyzed using a high-sensitive ELISA developed in our laboratory. A lipidomic approach for studying lipid metabolism was performed using ultra-high performance liquid chromatography coupled to electrospray ionization and quadrupole time-of-flight mass spectrometry (UHPLC-ESI-QTOF-MS).
Mild and severe COVID-19 patients had higher levels of IgM to glycerophosphocholines than control group. Mild COVID-19 patients showed higher levels of IgM to glycerophosphoinositol, glycerophosphoserine and sulfatides than control group and mild cases. 82.5% of mild COVID-19 patients showed IgM to glycerophosphoinositol or glycerophosphocholines plus sulfatides or glycerophosphoserines. Only 35% of severe cases and 27.5% of control group were positive for IgM to these lipids. Lipidomic analysis identify a total of 196 lipids, including 172 glycerophospholipids and 24 sphingomyelins. Increased levels of lipid subclasses belonging to lysoglycerophospholipids, ether and/or vinyl-ether-linked glycerophospholipids, and sphingomyelins were observed in severe COVID-19 patients, when compared with those of mild cases and control group.
Antibodies to lipids are essential for defense against SARS-CoV-2. Patients with low levels of anti-lipid antibodies have an elevated inflammatory response mediated by lysoglycerophospholipids. These findings provide novel prognostic biomarkers and therapeutic targets.
A mixture of sphingomyelin (SM) and cholesterol (Chol) exhibits a characteristic lipid raft domain of the cell membranes that provides a platform to which various signal molecules as well as virus ...and bacterial proteins are recruited. Several proteins capable of specifically binding either SM or Chol have been reported. However, proteins that selectively bind to SM/Chol mixtures are less well characterized. In our screening for proteins specifically binding to SM/Chol liposomes, we identified a novel ortholog of Pleurotus ostreatus, pleurotolysin (Ply)A, from the extract of edible mushroom Pleurotus eryngii, named PlyA2. Enhanced green fluorescent protein (EGFP)-conjugated PlyA2 bound to SM/Chol but not to phosphatidylcholine/Chol liposomes. Cell surface labeling of PlyA2-EGFP was abolished after sphingomyelinase as well as methyl-β-cyclodextrin treatment, removing SM and Chol, respectively, indicating that PlyA2-EGFP specifically binds cell surface SM/Chol rafts. Tryptophan to alanine point mutation of PlyA2 revealed the importance of C-terminal tryptophan residues for SM/Chol binding. Our results indicate that PlyA2-EGFP is a novel protein probe to label SM/Chol lipid domains both in cell and model membranes.
Phospholipids have been used widely in the food industry as emulsifiers, but it is their biological and nutritional function that has been the focus over recent years. The recognition of the ...importance of phospholipids and sphingomyelin for infant development has led to an increase in the number of infant formulas claiming to contain these complex lipid components. Therefore, the ability to measure these lipids in infant formulas and dairy-derived complex lipid ingredients for fortification purposes is important. A high-performance liquid chromatography–tandem mass spectrometry method that quantifies phosphatidylcholine, phosphatidylethanolamine, phosphatidylinositol, phosphatidylserine, and sphingomyelin found in infant formulas and dairy-derived complex lipid ingredients is described. The method uses external standards of similar fatty acid profile for calibration. The recovery of phospholipids ranged from 92% to 102% with a method repeatability coefficient of variation of 6–10%. In addition to the specificity and selectivity of the method, details of the molecular species in the individual phospholipid classes are available using this method.
Despite the high prevalence and devastating impact of psychiatric disorders, little is known about their etiopathology. In this review, we provide an overview on the participation of sphingolipids ...and enzymes responsible for their metabolism in mechanisms underlying psychiatric disorders. We focus on the pathway from sphingomyelin to proapoptotic ceramide and the subsequent metabolism of ceramide to sphingosine, which is in turn phosphorylated to yield anti-apoptotic sphingosine-1-phosphate (S1P).The sphingomyelinase/ceramide system has been linked to effects of reactive oxygen species and proinflammatory cytokines in the central nervous system as well as to synaptic transmission. Compared to ubiquitously expressed acid sphingomyelinase, acid and neutral ceramidase and neutral sphingomyelinase are highly active in brain regions. Depressed patients show elevated plasma ceramide levels and increased activities of acid sphingomyelinase which is functionally inhibited by many anti-depressive drugs. Exposure to alcohol is associated with an activation of acid and neutral sphingomyelinase observed in cell culture, mouse models and in alcohol-dependent patients and with increased concentrations of ceramide in various organs.Levels of sphingomyelin and ceramide are altered in erythrocytes and post-mortem brain tissues of schizophrenic patients in addition to changes in expression patterns for serine palmitoyltransferase and acid ceramidase leading to impaired myelination. After induction of anxiety-like behavior in animal models, higher serum levels of S1P were reported to lead to neurodegeneration. Correspondingly, S1P infusion appeared to increase anxiety-like behavior. Significantly upregulated levels of the endogenous ceramide catabolite N,N-dimethylsphingosine were observed in rat models of allodynia. Conversely, rats injected intrathecally with N,N-dimethylsphingosine developed mechanical allodynia. Moreover, S1P has been implicated in spinal nociceptive processing.The increasing interest in lipidomics and improved analytical methods led to growing insight into the connection between psychiatric and neurological disorders and sphingolipid metabolism and may once provide new targets and strategies for therapeutic intervention.
Abstract
Background
Experimental evidence supports a role of lipid dysregulation in ovarian cancer progression. We estimated associations with ovarian cancer risk for circulating levels of four lipid ...groups, previously hypothesized to be associated with ovarian cancer, measured 3–23 years before diagnosis.
Methods
Analyses were conducted among cases (N = 252) and matched controls (N = 252) from the Nurses’ Health Studies. We used logistic regression adjusting for risk factors to investigate associations of lysophosphatidylcholines (LPCs), phosphatidylcholines (PCs), ceramides (CERs), and sphingomyelins (SMs) with ovarian cancer risk overall and by histotype. A modified Bonferroni approach (0.05/4 = 0.0125, four lipid groups) and the permutation-based Westfall and Young approach were used to account for testing multiple correlated hypotheses. Odds ratios (ORs; 10th–90th percentile), and 95% confidence intervals of ovarian cancer risk were estimated. All statistical tests were two-sided.
Results
SM sum was statistically significantly associated with ovarian cancer risk (OR = 1.97, 95% CI = 1.16 to 3.32; P = .01/permutation-adjusted P = .20). C16:0 SM, C18:0 SM, and C16:0 CERs were suggestively associated with risk (OR = 1.95–2.10; P = .004–.01; permutation-adjusted P = .08–.21). SM sum, C16:0 SM, and C16:0 CER had stronger odds ratios among postmenopausal women (OR = 2.16–3.22). Odds ratios were similar for serous/poorly differentiated and endometrioid/clear cell tumors, although C18:1 LPC and LPC to PC ratio were suggestively inversely associated, whereas C18:0 SM was suggestively positively associated with risk of endometrioid/clear cell tumors. No individual metabolites were associated with risk when using the permutation-based approach.
Conclusions
Elevated levels of circulating SMs 3–23 years before diagnosis were associated with increased risk of ovarian cancer, regardless of histotype, with stronger associations among postmenopausal women. Further studies are required to validate and understand the role of lipid dysregulation in ovarian carcinogenesis.
Under certain conditions, biological membranes exhibit resistance to solubilization, even at high detergent concentration. These insoluble fragments are enriched in sphingolipids, cholesterol, and ...certain proteins having a preference for more organized environments. Here we investigated the effect of detergent Triton X-100 (TX-100) on raft-like lipid mixtures composed of POPC (palmitoyl oleoyl phosphatidylcholine, an unsaturated lipid), SM (sphingomyelin, a saturated lipid), and cholesterol, focusing on the detergent-induced phase separation at subsolubilizing concentration and the extent of solubilization at higher concentration. Giant unilamellar vesicles (GUVs) of POPC/SM/chol containing a fluorescent probe known to prefer the liquid-disordered phase were prepared and observed with fluorescence microscopy. A phase diagram constructed in the presence and absence of 0.1 mM TX-100 showed that the detergent induces macroscopic liquid-ordered/liquid-disordered (Lo/Ld) phase separation over a wide range of membrane composition, indicating that TX-100 has the ability to rearrange the lateral heterogeneity of the lipid mixture. The extent of solubilization of the POPC/SM/chol GUVs was quantified by measuring the vesicle size before and after the injection of a high concentration of TX-100. In parallel, the solubilization extent of large unilamellar vesicles (LUVs) was assessed by turbidity measurements. The extent of solubilization decreases significantly as the fractions of SM and cholesterol in the mixture increase. The origin of the detergent resistance is the low partitioning of TX-100 in cholesterol-rich membranes, especially in SM-containing ones, as evidenced by isothermal titration calorimetry experiments on LUVs. Our results provide a guide to future research on the effects of TX-100 on raft-like lipid mixtures.
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