Sphingoid Backbone Research Articles

Characterization of Unique Eukaryotic Sphingolipids with Temperature-Dependent Δ8-Unsaturation from the Picoalga Ostreococcus tauri.

Sphingolipids (SLs) are ubiquitous components of eukaryotic cell membranes and are found in some prokaryotic organisms and viruses. They are composed of a sphingoid backbone that may be acylated and glycosylated. Assembly of various sphingoid base, fatty acyl and glycosyl moieties results in highly diverse structures. The functional significance of variations in SL chemical diversity and abundance is still in the early stages of investigation. Among SL modifications, Δ8-desaturation of the sphingoid base occurs only in plants and fungi. In plants, SL Δ8-unsaturation is involved in cold hardiness. Our knowledge of the structure and functions of SLs in microalgae lags far behind that of animals, plants and fungi. Original SL structures have been reported from microalgae. However, functional studies are still missing. Ostreococcus tauri is a minimal microalga at the base of the green lineage and is therefore a key organism for understanding lipid evolution. In the present work, we achieved the detailed characterization of O. tauri SLs and unveiled unique glycosylceramides as sole complex SLs. The head groups are reminiscent of bacterial SLs, as they contain hexuronic acid residues and can be polyglycosylated. Ceramide backbones show a limited variety, and SL modification is restricted to Δ8-unsaturation. The Δ8-SL desaturase from O. tauri only produced E isomers. Expression of both Δ8-SL desaturase and Δ8-unsaturation of sphingolipids varied with temperature, with lower levels at 24°C than at 14°C. Overexpression of the Δ8-SL desaturase dramatically increases the level of Δ8 unsaturation at 24°C and is paralleled by a failure to increase cell size. Our work provides the first characterization of O. tauri SLs and functional evidence for the involvement of SL Δ8-unsaturation for temperature acclimation in microalgae, suggesting that this function is an ancestral feature in the green lineage.

Robust quantitation of gangliosides and sulfatides in human brain using UHPLC-MRM-MS: Method development and application in Alzheimer's disease

Gangliosides (GAs) and sulfatides (STs) are acidic glycosphingolipids that are particularly abundant in the nervous system and are closely related to aging and neurodegenerative disorders. To explore their roles in brain diseases, in-depth molecular profiling, including structural variations of sphingoid backbone, fatty acyl group, and sugar chain of GAs and STs was performed. A total of 210 GAs and 38 STs were characterized in the inferior frontal gyrus (IFG) of human brain, with 90 GAs discovered in brain tissues for the first time. Influential MS parameters for detecting GAs and STs in multiple reaction monitoring (MRM) mode were systematically examined and optimized to minimize in-source fragmentation, resulting in remarkable signal intensity enhancement for GAs and STs, especially for polysialylated species. To eliminate analytical variations, isotopic interference-free internal standards were prepared by simple and fast reduction reaction. The final established method facilitated the simultaneous quantitation of 184 GAs and 30 STs from 25 subtypes, which represents the highest number of GAs quantitated among all quantitation methods recorded in literature so far. The method was further validated and applied to reveal the aberrant change of GAs and STs in the IFG of 12 Alzheimer's disease (AD) patients. Four GAs exhibited high classification capacity for AD (AUC ≥0.80) and were thereby considered the most promising signatures for AD. These findings suggested the close correlation between GAs and the pathogenesis of AD, highlighting the achievements of our robust method for investigating the roles of GAs and STs in various physiological states and diseases.

Circulating Ceramides and Sphingomyelins and Risk of Mortality: The Cardiovascular Health Study.

Recent studies suggest that associations of ceramides (Cer) and sphingomyelins (SM) with health outcomes differ according to the fatty acid acylated to the sphingoid backbone. The purpose of this study was to assess associations of Cer and SM species with mortality. The study population included participants from the Cardiovascular Health Study (CHS), a community-based cohort of adults aged ≥65 years who were followed from 1992-2015 (n = 4612). Associations of plasma Cer and SM species carrying long-chain (i.e., 16:0) and very-long-chain (i.e., 20:0, 22:0, 24:0) saturated fatty acids with mortality were assessed using Cox proportional hazards models. During a median follow-up of 10.2 years, 4099 deaths occurred. High concentrations of Cer and SM carrying fatty acid 16:0 were each associated with an increased risk of mortality. Conversely, high concentrations of several ceramide and sphingomyelin species carrying longer fatty acids were each associated with a decreased risk of mortality. The hazard ratios for total mortality per 2-fold difference in each Cer and SM species were: 1.89 (95% CI), 1.65-2.17 for Cer-16, 0.79 (95% CI, 0.70-0.88) for Cer-22, 0.74 (95% CI, 0.65-0.84) for Cer-24, 2.51 (95% CI, 2.01-3.14) for SM-16, 0.68 (95% CI, 0.58-0.79) for SM-20, 0.57 (95% CI, 0.49-0.67) for SM-22, and 0.66 (0.57-0.75) for SM-24. We found no association of Cer-20 with risk of death. Associations of Cer and SM with the risk of death differ according to the length of their acylated saturated fatty acid. Future studies are needed to explore mechanisms underlying these relationships.

In-Depth Structural Characterization and Quantification of Cerebrosides and Glycosphingosines with Gas-Phase Ion Chemistry.

Cerebrosides (n-HexCer) and glycosphingosines (n-HexSph) constitute two sphingolipid subclasses. Both are comprised of a monosaccharide headgroup (glucose or galactose in mammalian cells) linked via either an α- or β-glycosidic linkage to the sphingoid backbone (n = α or β, depending upon the nature of the linkage to the anomeric carbon of the sugar). Cerebrosides have an additional amide-bonded fatty acyl chain linked to the sphingoid backbone. While differentiating the multiple isomers (i.e. glucose vs galactose, α- vs β-linkage) is difficult, it is crucial for understanding their specific biological roles in health and disease states. Shotgun tandem mass spectrometry has been a powerful tool in both lipidomics and glycomics analysis but is often limited in its ability to distinguish isomeric species. This work describes a new strategy combining shotgun tandem mass spectrometry with gas-phase ion chemistry to achieve both differentiation and quantification of isomeric cerebrosides and glycosphingosines. Briefly, deprotonated cerebrosides, [n-HexCer-H]-, or glycosphingosines, [n-HexSph-H]-, are reacted with terpyridine (Terpy) magnesium complex dications, [Mg(Terpy)2]2+, in the gas phase to produce a charge-inverted complex cation, [n-HexCer-H+MgTerpy]+ or [n-HexSph-H+MgTerpy]+. The collision-induced dissociation (CID) of the charge-inverted complex cations leads to significant spectral differences between the two groups of isomers, α-GalCer, β-GlcCer, and β-GalCer for cerebrosides and α-GlcSph, α-GalSph, β-GlcSph, and β-GalSph for glycosphingosines, which allows for isomer distinction. Moreover, we describe a quantification strategy with the normalized percent area extracted from selected diagnostic ions that quantify either three isomeric cerebroside or four isomeric glycosphingosine mixtures. The analytical performance was also evaluated in terms of accuracy, repeatability, and interday precision. Furthermore, CID of the product ions resulting from 443 Da loss from the charge-inverted complex cations ([n-HexCer-H+MgTerpy]+) has been performed and demonstrated for localization of the double-bond position on the amide-bonded monounsaturated fatty acyl chain in the cerebroside structure. The proposed strategy was successfully applied to the analysis of total cerebroside extracts from the porcine brain, providing in-depth structural information on cerebrosides from a biological mixture.

Synthesis of new ceramide analogues with allene in the sphingoid backbone

Ceramide is an important sphingolipid which can mediate a diverse range of biological activities. New ceramide analogues with an allene group in the sphingoid backbone were synthesized. The synthesis of the key intermediates involved the LiAlH4 reduction of oxazolidine intermediates via a directed reduction-elimination reaction. Hydrolysis of the oxazolidine, liberation of the amino group, and N-acylation provided new ceramide analogues. These new analogues may provide new resources to study their bioactivities and the structure-activity relationship.

Canonical and 1-Deoxy(methyl) Sphingoid Bases: Tackling the Effect of the Lipid Structure on Membrane Biophysical Properties.

Compared to the canonical sphingoid backbone of sphingolipids (SLs), atypical long-chain bases (LCBs) lack C1-OH (1-deoxy-LCBs) or C1-CH2OH (1-deoxymethyl-LCBs). In addition, when unsaturated, they present a cis-double bond instead of the canonical Δ4-5 trans-double bond. These atypical LCBs are directly correlated with the development and progression of hereditary sensory and autonomic neuropathy type 1 and diabetes type II through yet unknown mechanisms. Changes in membrane properties have been linked to thebiological actions of SLs. However, little is known about the influence of the LCB structure, particularly 1-deoxy(methyl)-LCB, on lipid-lipid interactions and their effect on membrane properties. To address this question, we used complementary fluorescence-based methodologies to study membrane model systems containing POPC and the different LCBs of interest. Our results show that 1-deoxymethyl-LCBs have the highest ability to reduce the fluidity of the membrane, while the intermolecular interactions of 1-deoxy-LCBs were found to be weaker, leading to the formation of less-ordered domains compared to their canonical counterparts-sphinganine and sphingosine. Furthermore, while the presence of a trans-double bond at the Δ4-5 position of the LCB increased the fluidity of the membrane compared to a saturated LCB, acis-double bond completely disrupted the ability of the LCB to segregate into ordered domains. In conclusion, even small changes on the structure of the LCB, as seen in 1-deoxy(methyl)-LCBs, strongly affects lipid-lipid interactions and membrane fluidity. These results provide evidence that altered balance between species with different LCBsaffect membrane properties and may contribute to the pathobiological role of these lipids.

Targeted analysis of ceramides and cerebrosides in yellow lupin seeds by reversed-phase liquid chromatography coupled to electrospray ionization and multistage mass spectrometry

Ceramides (Cer) and cerebrosides are important sphingolipids (SL) involved in many biological processes. Herein, the SL content of yellow lupin seeds (Lupinus luteus) was determined by liquid–liquid extraction, mild alkaline hydrolysis (1 h at 37 °C) and reversed-phase liquid chromatography with negative electrospray ionization coupled to either an orbital-trap (Fourier-transformed, FT) or linear ion-trap (LIT) mass spectrometry (RPLC-ESI/MS). The chemical identity of SL including the sugar residues and sphingoid backbone (SB) was inferred by collision-induced dissociation multiple-stage mass spectrometry (MSn, n = 2,3). Up to 52 Cer and 47 cerebrosides were successfully recognized and quantified in sample extracts of L. luteus seeds also counting isobaric species. As reported for other vegetables, a hydroxylated SB was observed whereby the N-acyl chains showed a high occurrence of very-long-chain moieties; phytosphingosine and 4-hydroxy-8-sphingenine were the predominant SB paired mainly with oleic acid (C18:1), hydroxylated behenic acid (C22:0;1) and hydroxylated lignoceric acid (C24:0;1).

Dynamics of sphingolipids and the serine palmitoyltransferase complex in rat oligodendrocytes during myelination

Myelin is a unique lipid-rich membrane structure that accelerates neurotransmission and supports neuronal function. Sphingolipids are critical myelin components. Yet sphingolipid content and synthesis have not been well characterized in oligodendrocytes, the myelin-producing cells of the CNS. Here, using quantitative real-time PCR, LC-MS/MS-based lipid analysis, and biochemical assays, we examined sphingolipid synthesis during the peak period of myelination in the postnatal rat brain. Importantly, we characterized sphingolipid production in isolated oligodendrocytes. We analyzed sphingolipid distribution and levels of critical enzymes and regulators in the sphingolipid biosynthetic pathway, with focus on the serine palmitoyltransferase (SPT) complex, the rate-limiting step in this pathway. During myelination, levels of the major SPT subunits increased and oligodendrocyte maturation was accompanied by extensive alterations in the composition of the SPT complex. These included changes in the relative levels of two alternative catalytic subunits, SPTLC2 and -3, in the relative levels of isoforms of the small subunits, ssSPTa and -b, and in the isoform distribution of the SPT regulators, the ORMDLs. Myelination progression was accompanied by distinct changes in both the nature of the sphingoid backbone and the N-acyl chains incorporated into sphingolipids. We conclude that the distribution of these changes among sphingolipid family members is indicative of a selective channeling of the ceramide backbone toward specific downstream metabolic pathways during myelination. Our findings provide insights into myelin production in oligodendrocytes and suggest how dysregulation of the biosynthesis of this highly specialized membrane could contribute to demyelinating diseases.

Ceramides: Nutrient Signals that Drive Hepatosteatosis.

Ceramides are minor components of the hepatic lipidome that have major effects on liver function. These products of lipid and protein metabolism accumulate when the energy needs of the hepatocyte have been met and its storage capacity is full, such that free fatty acids start to couple to the sphingoid backbone rather than the glycerol moiety that is the scaffold for glycerolipids (e.g., triglycerides) or the carnitine moiety that shunts them into mitochondria. As ceramides accrue, they initiate actions that protect cells from acute increases in detergent-like fatty acids; for example, they alter cellular substrate preference from glucose to lipids and they enhance triglyceride storage. When prolonged, these ceramide actions cause insulin resistance and hepatic steatosis, 2 of the underlying drivers of cardiometabolic diseases. Herein the author discusses the mechanisms linking ceramides to the development of insulin resistance, hepatosteatosis and resultant cardiometabolic disorders.

Large-scale lipidomics identifies associations between plasma sphingolipids and T2DM incidence.

Sphingolipids (SPs) are ubiquitous, structurally diverse molecules that include ceramides, sphingomyelins, and sphingosines. They are involved in various pathologies including obesity and type 2 diabetes mellitus (T2DM). Therefore, it is likely that perturbations in plasma concentrations of SPs are associated with disease. Identifying these associations may reveal useful biomarkers or provide insight into disease processes. We performed a lipidomics evaluation of molecularly-distinct SPs in the plasma of 2,302 ethnically-Chinese Singaporeans using electrospray ionization mass spectrometry coupled with liquid chromatography. SP profiles were compared to clinical and biochemical characteristics, and subjects were evaluated by follow-up visits for 11 years. We found that ceramides correlate positively but hexosylceramides correlate negatively with body mass index (BMI) and homeostatic model assessment of insulin resistance (HOMA-IR). Furthermore, SPs with a d16:1 sphingoid backbone correlate more positively with BMI and HOMA-IR, while d18:2 SPs correlate less positively, relative to canonical d18:1 SPs. We also found that higher concentrations of two distinct sphingomyelins were associated with a higher risk of T2DM (HR 1.45, 95% CI 1.18-1.78 for SM d16:1/C18:0; and HR 1.40, 95% CI 1.17-1.68 for SM d18:1/C18:0). We identified significant associations between SPs and obesity/T2DM characteristics, specifically, that of hexosylceramides, d16:1 SPs, and d18:2 SPs. This suggests that the balance of SP metabolism, rather than ceramide accumulation, is associated with the pathology of obesity. We further identified two specific SPs that may represent prognostic biomarkers for T2DM. Funding sources are listed in the Acknowledgements section.

Plasma Sphingolipids and Subclinical Atherosclerosis – Novel Associations Uncovered by a Large-scale Lipidomic Analysis (P18-129-19)

ObjectivesSphingolipids (SP) are a diverse class of heterogenous lipids that includes ceramides, sphingomyelins, and glycosphingolipids. Many SPs have diverse roles in cell functions including cell growth, inflammation and angiogenesis, and previous studies suggest that SPs may also be involved in the pathogenesis of cardiovascular diseases. We aimed to identify plasma SPs and subsequently evaluate associations between plasma SPs and subclinical atherosclerosis in a subset of the Multiethnic Cohort of Singapore with the goal of uncovering novel biomarkers predictive of heart disease. MethodsWe conducted a lipidomics evaluation of 103 molecularly-distinct SPs in the plasma of 559 Singaporeans aged 50 years and above using electrospray ionization mass spectrometry coupled with liquid chromatography. All participants did not have a history of diabetes, heart disease, stroke or cancer at baseline, and completed a detailed health screening that evaluated risk factors for cardiovascular disease including computed tomography scans of the abdomen and coronary arteries. Multivariable logistic regression models were used to estimate odds ratios (ORs) and 95% confidence intervals (CIs) for associations between SPs and subclinical atherosclerosis (defined as coronary artery calcium score ≥ 100). ResultsCeramides (Cer), particularly those with a d16:1 or d18:1 backbone, were directly associated with higher risk of subclinical atherosclerosis whereas a small number of monohexosylceramides (MHCer), dihexosylceramides (DHCer) and sphingomyelins (SM) with a d18:2 sphingoid backbone were inversely associated. However most associations were attenuated after adjusting for conventional cardiovascular risk factors, including blood lipid (low-density lipoprotein cholesterol, high-density lipoprotein cholesterol, triglycerides) concentrations and glycemic markers, suggesting that the associations may be mediated by these risk factors. ConclusionsHigh-throughput lipidomics may uncover novel sphingolipids predictive of heart disease. Characterization of these lipid species could provide insights into disease etiology. Funding SourcesThis work was supported by the National University Health System and the National Research Foundation Investigatorship grant. Supporting Tables, Images and/or Graphs▪▪

The role of dihydrosphingolipids in disease.

Dihydrosphingolipids refer to sphingolipids early in the biosynthetic pathway that do not contain a C4-trans-double bond in the sphingoid backbone: 3-ketosphinganine (3-ketoSph), dihydrosphingosine (dhSph), dihydrosphingosine-1-phosphate (dhS1P) and dihydroceramide (dhCer). Recent advances in research related to sphingolipid biochemistry have shed light on the importance of sphingolipids in terms of cellular signalling in health and disease. However, dihydrosphingolipids have received less attention and research is lacking especially in terms of their molecular mechanisms of action. This is despite studies implicating them in the pathophysiology of disease, for example dhCer in predicting type 2 diabetes in obese individuals, dhS1P in cardiovascular diseases and dhSph in hepato-renal toxicity. This review gives a comprehensive summary of research in the last 10-15years on the dihydrosphingolipids, 3-ketoSph, dhSph, dhS1P and dhCer, and their relevant roles in different diseases. It also highlights gaps in research that could be of future interest.

Novel phytoceramides containing fatty acids of diverse chain lengths are better than a single C18-ceramide N-stearoyl phytosphingosine to improve the physiological properties of human stratum corneum.

Ceramides in the human stratum corneum (SC) are a mixture of diverse N-acylated fatty acids (FAs) with different chain lengths. C24 is the major class of FAs of ceramides. However, there are also other classes of ceramides with diverse chain lengths of FAs, and these lengths generally range from C16 to C26. This study aimed to prepare several types of phytoceramide containing diverse chain lengths of N-acylated FAs and compare them with C18-ceramide N-stearoyl phytosphingosine (NP) in terms of their effects on the physiological properties of the SC. We chose natural oils, such as horse fat oil, shea butter, sunflower oil, and a mixture of macadamia nut, shea butter, moringa, and meadowfoam seed oil, as sources of FAs and phytosphingosine as a sphingoid backbone to synthesize diverse phytoceramides. Each phytoceramide exhibited a distinctive formation of the lamellar structure, and their FA profiles were similar to those of their respective natural oil. The skin barrier properties, as analyzed in human skin, clearly demonstrated that all the phytoceramides improved the recovery rate of the damaged SC and enhanced hydration better than C18-ceramide NP did. In conclusion, natural oil-derived phytoceramides could represent a novel class of ceramides for cosmetic applications in the development of an ideal skin barrier moisturizer.

Sphingomyelin - Cerebroside Exchange in Lipid Membrane Lateral Domain Segregation

Galactocerebrosides, commonly referred to as cerebrosides (Ceb), are a family of glycosphingolipids, which contain a saccharide head group (galactose, glucose) covalently linked to a double-tailed ceramide via a glycosidic linkage through the primary hydroxyl. They can be found in neuronal tissues in the central nervous system, in the epithelial cells of the small intestine and colon and in the granular layer of the skin epidermis. Ceb, like sphingomyelins (SM), are found to be highly saturated in natural sources and both share a sphingoid backbone but with a different headgroup, a phosphorylcholine (SM) or a mono/oligosaccharide (Ceb).

Possible roles of long-chain sphingomyelines and sphingomyelin synthase 2 in mouse macrophage inflammatory response

To evaluate the precise role of sphingomyelin synthase 2 (SMS2) in sphingomyelin (SM) metabolism and their anti-inflammatory properties, we analyzed species of major SM and ceramide (Cer) (18:1, 18:0 sphingoid backbone, C14 - C26 N-acyl part) in SMS2 knockout and wild-type mouse plasma and liver using HPLC-MS. SMS2 deficiency significantly decreased very long chain SM (SM (d18:1/22:0) and SM (d18:1/24:0 or d18:0/24:1)) and increased very long chain Cer (Cer (d18:1/24:0 or d18:0/24:1) and Cer (d18:1/24:1)), but not long chain SM (SM (d18:1/16:0), SM (d18:1/18:0 or d18:0/18:1) and SM (d18:1/18:1)) in plasma.To examine the effects of SM on inflammation, we studied the role of very long chain SM in macrophage activation. Addition of SM (d18:1/24:0) strongly upregulated several macrophage activation markers, SM (d18:1/6:0) and Cer (d18:1/24:0) however, did not.It was suggested that very long chain SM but not long chain SM were decreased in SMS2-deficient mice liver and plasma. And the exogenously added very long chain SM (d18:1/24:0) could activate macrophages directly, suggesting a novel role of plasma very long chain SM in modulating macrophage activation and resulting inflammation.

Sphingolipidomics: An Important Mechanistic Tool for Studying Fungal Pathogens.

Sphingolipids form of a unique and complex group of bioactive lipids in fungi. Structurally, sphingolipids of fungi are quite diverse with unique differences in the sphingoid backbone, amide linked fatty acyl chain and the polar head group. Two of the most studied and conserved sphingolipid classes in fungi are the glucosyl- or galactosyl-ceramides and the phosphorylinositol containing phytoceramides. Comprehensive structural characterization and quantification of these lipids is largely based on advanced analytical mass spectrometry based lipidomic methods. While separation of complex lipid mixtures is achieved through high performance liquid chromatography, the soft – electrospray ionization tandem mass spectrometry allows a high sensitivity and selectivity of detection. Herein, we present an overview of lipid extraction, chromatographic separation and mass spectrometry employed in qualitative and quantitative sphingolipidomics in fungi.

Approaches to polyunsaturated sphingolipids: new conformationally restrained analogs with minimal structural modifications

Conformationally restrained sphingolipids 1–4, arising from the introduction of a polyene or a polyenyne moiety as part of the sphingoid backbone, have been synthesized. While addition of polyene acetylides 6 and 7 to Garner's aldehyde afforded the corresponding Felkin-Anh anti-addition adducts, addition of alkenylzirconocenes showed no diastereoselectivity. This behaviour was also observed from protected serinal 14, which allowed the synthesis of the acid-sensitive pentaene sphingosine backbone present in ceramide 4. Despite the inherent limitations of the synthetic approaches here reported, their application to such highly conjugated polyene sphingoid analogs is unprecedented.

Ceramide synthases in biomedical research

Sphingolipid metabolism consists of multiple metabolic pathways that converge upon ceramide, one of the key molecules among sphingolipids (SLs). In mammals, ceramide synthesis occurs via N-acylation of sphingoid backbones, dihydrosphingosine (dhSo) or sphingosine (So). The reaction is catalyzed by ceramide synthases (CerS), a family of enzymes with six different isoforms, with each one showing specificity towards a restricted group of acyl-CoAs, thus producing ceramides (Cer) and dihydroceramides (dhCer) with different fatty acid chain lengths. A large body of evidence documents the role of both So and dhSo as bioactive molecules, as well as the involvement of dhCer and Cer in physiological and pathological processes. In particular, the fatty acid composition of Cer has different effects in cell biology and in the onset and progression of different diseases. Therefore, modulation of CerS activity represents an attractive target in biomedical research and in finding new treatment modalities. In this review, we discuss functional, structural and biochemical features of CerS and examine CerS inhibitors that are currently available.

Tuning of membrane electrostatic properties by single chain sphingolipids sphingosine and sphingosine-1-phosphate: The effect on bilayer dipole potential

Abstract An important question in membrane biological chemistry is whether bioactive signaling lipids act only as second messenger ligands or also through an effect on bilayer physical properties. Sphingosine (Sph) and sphingosine-1-phosphate (S1P) are single-chained charged sphingolipids that have antagonistic functions in the “sphingolipid rheostat” which determines cell fate. Sph and S1P respectively promote apoptosis and cell growth. In the present study, the effects of these bioactive lipids on the dipole potential of the lipid bilayer were evaluated. We have investigated the effect of both sphingolipids, incorporated separately or together, in large egg phosphadidylcholine (EPC) unilamellar vesicles on the fluorescence of di-8-ANEPPS, a probe of dipole potential. Both sphingolipids promote a decrease of the dipole potential which is more pronounced for Sph than for S1P. This can be explained by the polarization properties of both the single-chain sphingoid backbone and its surrounding water molecules, as well as by the opposite net charges of the two sphingolipids. When both sphingolipids are present together, as occurs physiologically, the dipole potential varies monotonously over a significant range as a function of the Sph/S1P ratio. This suggests that both sphingolipids are able to tune the dipole potential of cell membranes as a function of their ratio, thereby possibly influencing the binding of surface proteins and the activity of transmembrane proteins. This furthers the idea that the two sphingolipids might exert their biological activity not only as second messenger ligands but also through their effect on lipid membrane physicochemical properties.

The role of the trans double bond in skin barrier sphingolipids: permeability and infrared spectroscopic study of model ceramide and dihydroceramide membranes.

Dihydroceramides (dCer) are members of the sphingolipid family that lack the C4 trans double bond in their sphingoid backbone. In addition to being precursors of ceramides (Cer) and phytoceramides, dCer have also been found in the extracellular lipid membranes of the epidermal barrier, the stratum corneum. However, their role in barrier homeostasis is not known. We studied how the lack of the trans double bond in dCer compared to Cer influences the permeability, lipid chain order, and packing of multilamellar membranes composed of the major skin barrier lipids: (d)Cer, fatty acids, cholesterol, and cholesteryl sulfate. The permeability of the membranes with long-chain dCer was measured using various markers and was either comparable to or only slightly greater than (by up to 35%, not significant) that of the Cer membranes. The dCer were less sensitive to acyl chain shortening than Cer (the short dCer membranes were up to 6-fold less permeable that the corresponding short Cer membranes). Infrared spectroscopy showed that long dCer mixed less with fatty acids but formed more thermally stable ordered domains than Cer. The key parameter explaining the differences in permeability in the short dCer and Cer was the proportion of the orthorhombic phase. Our results suggest that the presence of the trans double bond in Cer is not crucial for the permeability of skin lipid membranes and that dCer may be underappreciated members of the stratum corneum lipid barrier that increase its heterogeneity.