Cardiovascular Implications of Sphingomyelin Presence in Biological Membranes

Abstract

Sphingomyelin (SM) is a type of sphingolipid found within plasma, cellular membranes and plasma lipoproteins. Here we highlight the basic biochemical features of SMs and their role in biological membranes. We further discuss evidence of the association between SM and cardiovascular diseases such as atherosclerosis, valvular disease, heart failure and diabetes mellitus.

Sphingolipids are one of the major categories of lipids and beyond their role as structural membrane components they have important functions as signalling molecules in a wide array of biological processes. They are composed of two key lipid building blocks – long-chain bases (usually sphingosine or 1,3-dihydroxy-2-amino-4-octadecene) and fatty acids – and use a glycerol-based backbone to which acyl chains are attached.[1,2]

Sphingomyelins (SMs) are among the most common sphingolipids in many mammalian cells and tissues, especially in the membranous myelin sheath that surrounds nerve cell axons.[3] SMs have significant structural and functional roles in the cell such as creating unique lateral structures (lipid rafts and ordered domains) in membranes, binding to and functional regulation of membrane-spanning proteins and involvement in cell signalling events.[3] They play an important role in the regulation of plasma membrane and cell cholesterol homeostasis.[3]

SM was first isolated from brain tissue by German chemist Johann LW Thudicum in the 1880s.[4] In 1927 the structure of SM was first reported by Pick and Bielschowsky; SM is an N-acyl-sphingosine-1-phosphorylcholine, consisting of a phosphocholine head group, a sphingosine and a fatty acid.[5] The most common long-chain base in SM is 1,3-dihydroxy-2-amino-4-octadecene (sphingosine or d18:1). The most common N-linked acyl chain of SMs in mammalian peripheral cells is palmitic acid (16:0), whereas stearic acid (18:0) is more common in neural tissue SMs. Long-chain fatty acids (e.g. 24:0 and 24:1) are also common constituents of the SMs found in most tissues.[3]

SMs are synthesised in the endoplasmic reticulum, where they can be found in low levels, and in the trans Golgi.[6] The first step in SM synthesis is the condensation of L-serine and palmitoyl-coenzyme A. This reaction is catalysed by serine palmitoyltransferase and yields 3-keto-dihydrosphingosine, which is reduced to dihydrosphingosine. Dihydrosphingosine undergoes N-acylation followed by desaturation to generate ceramide, which has a central role in sphingolipid metabolism.[3,7] These reactions occur on the cytosolic surface of the endoplasmic reticulum. Subsequently, ceramide is delivered to the Golgi apparatus where it is converted to SM or glucosylceramide. In most mammalian cell types the majority of ceramide is converted to SM in the lumen of the trans Golgi by an SM synthase (SMS) enzyme named SMS1, which catalyses the transfer of phosphocholine from phosphatidylcholine to ceramide, yielding a diacylglycerol side product.[3,7] A second enzyme, SMS2, resides at the plasma membrane and may convert locally produced ceramide to SM.[3,7] An alternative pathway of SM synthesis has been postulated in which ceramide is first converted to ethanolamine phosphorylceramide via transfer of the head group from phosphatidylethanolamine.[7] Ethanolamine phosphorylceramide is then converted to SM by stepwise methylation. Although this pathway has been demonstrated in isolated membrane fractions from rat brain and liver, its precise contribution to the de novo synthesis of SM remains to be established.[7]

Since major SMS activity is found in the luminal trans Golgi and the plasma membranes, it is not surprising that SM is enriched in membranes derived from trans Golgi and in the plasma membrane (including the endosomal recycling compartment). The asymmetric location of SMS activity (luminal trans Golgi and outer leaflet of plasma membranes) may explain why SMs are enriched at the plasma membrane with a greater concentration on the outer than the inner leaflet.[3,6,8] In humans, SMs comprise nearly 85 % of all sphingolipids and 10–20 mol % of the total plasma membrane lipids. SMs comprise 4–18 % of all sarcolemma membrane lipids with 93 % present in the outer leaflet.[8]

The Role of Sphingomyelin in Biological Membranes

The presence of SM in biological membranes is associated with various critical functions. SMs accumulate in the exoplasmic leaflet of the plasma membrane where their high packing density and affinity for sterols help create a rigid barrier to the extracellular environment.[7] Under this notion, the membranous myelin sheath that surrounds nerve cell axons is particularly rich in SMs, which may suggest a role as an insulator of nerve fibres.[9,10]

SM presence in the plasma membrane directly affects cholesterol homeostasis.[11,12] The addition of SM to cells has been shown to increase cholesterol biosynthesis and affect LDL binding to cell surface receptors and subsequent internalisation.[13] Together they form SM/sterol-rich domains that often are more ordered than the surrounding phase in biological membranes.[14] The growing body of evidence regarding their favourable interaction with sterols indicates that SM may be a key a regulator of cholesterol distribution within cellular membranes and cholesterol homeostasis in cells.[14] In addition, excess levels of SM in the red blood cell membrane (i.e. abetalipoproteinemia) cause increased lipid accumulation in the outer leaflet of the red blood cell plasma membrane, which results in abnormally shaped red cells called acanthocytes.[15]

The SM pool in the plasma membrane acts as a reservoir of lipid signalling molecules, the liberation of which is catalysed by acidic or neutral SMSs in response to a variety of biological stimuli. SM metabolites such as ceramide,[16] sphingosine[17,18] and sphingosine 1-phosphate[17,18] are emerging as critical regulators of cell proliferation, differentiation and apoptosis.[7] The ability of SM to function as a precursor of signalling molecules in a wide array of biological processes is extensively reviewed elsewhere.[19]

As SMs have a strong, inherent capacity to form microdomains, their production in the trans Golgi may affect the lateral organisation of other membrane molecules and are thus associated with the formation of lipid rafts, caveoles and other micro-membrane structures involved in various cellular processes (including endocytosis, intracellular trafficking, and signal transduction by membrane receptors, cell–cell communication and host–pathogen interactions).[7,10,20-23]

SM synthesis in the trans Golgi may create a local pool of diacylglycerol, which leads to protein kinase D recruitment and the formation of secretory vesicles.[7,10,24]

The proportion of SM within a membrane as well as SM structural configuration contribute to membrane thickness and recruitment of specific integral proteins, thereby modulating membrane structural integrity and function.[25,10] Furthermore, published evidence suggests that SM interacts with these integral membrane proteins, modulating their functional role.[10]

Interdigitation occurs when a long acyl chain in SM located in one leaflet penetrates through the bilayer into the opposite leaflet. Such interdigitation has been shown to affect the diffusion rates in opposing glycerophospholipid leaflets, suggesting interleaflet coupling with possible biological significance.[26,10]

Cardiovascular Implications Associated with Sphingomyelin

Atherosclerosis/Thrombosis

A study of human carotid plaques showed that levels of several sphingolipids (including SM) were increased in plaques associated with symptoms and correlated with inflammatory cytokines.[27] All sphingolipids were correlated with histological markers of plaque instability and also induced an inflammatory response in human coronary artery smooth muscle cells. SM and ceramide were also found to be positively correlated with lipid plaque burden.[27] A study of hypercholesterolaemic rabbits found that SMs were relatively abundant within atherosclerotic lesions compared with other lipids.[28] Furthermore, strikingly elevated intra-platelet levels of triglycerides and SM were found in patients with ST-elevation MI in comparison with matched controls.[29]

Several studies have assessed the association between circulating plasma SM levels and incidence of cardiovascular disease, with conflicting results. Fernandez et al. showed that SM (38:2) levels was the only lipid species of its class to be associated with increased risk of future cardiovascular disease.[30] In another study, patients with stable angina, unstable angina or acute MI showed higher plasma SM and ceramide levels as well as higher SMS activity compared with healthy individuals.[31] However, in the population-based Multi-Ethnic Study of Atherosclerosis, a high plasma SM level was not associated with an increased risk of incident coronary heart disease in adults free of clinical cardiovascular disease at baseline.[32]

Recently, erythrocytes have received considerable attention as significant players in accelerating plaque progression, with observations of erythrocytes being ‘driven’ to the plaque lipid core via intraplaque haemorrhages.[33] Assessing the sphingolipid content of erythrocyte membranes, Zhang et al. demonstrated that SM levels were elevated in patients with an acute coronary syndrome compared with patients with stable angina.[34] However, no differences were found between patients with stable coronary artery disease and healthy individuals regarding SM content in erythrocyte membranes. In the same study, a significant correlation between plasma SM levels, erythrocyte membrane SM content and lipoprotein alpha(a) levels were shown. These observations suggest a link between SM content of erythrocyte membrane and clinical instability in coronary artery disease, which may reflect another potential source of atheromatous plaque instability via intraplaque haemorrhages. Therefore, the SM content of erythrocyte membranes could serve as a marker for coronary artery disease activity independent of other risk factors and clinical features.[34]

Diabetes

As part of the Finnish Diabetic Nephropathy Study, 326 patients with type 1 diabetes were studied for presence of diabetic renal disease.[35] Serum levels of SM and large HDL particles were the strongest predictors after adjustment of established kidney injury biomarkers. In addition, SM levels assessed in serum was correlated significantly with 24 hour albumin excretion rate. In contrast, another study demonstrated that compared with healthy controls, patients with impaired fasting glucose levels or type 2 diabetes had significantly reduced serum concentrations of glycerophospholipids and SMs, even after adjusting for age, gender and BMI or even when separately analysed for those not receiving lipid-modifying medications.[36] The authors suggested that the decrease in the SM pool in diabetes could potentially contribute to increased oxidative stress and reduced insulin secretion, resulting in hyperglycaemia secondary to an inability to compensate for reduced insulin sensitivity.[36]

Lipoproteins

SM is the most abundant sphingolipid in lipoproteins. Very low density liopoprotein/LDL and HDL comprise approximately 63–75 % and 25–35 % of SMs, respectively.[37] SM is slightly more abundant (13 %) in large light HDL2 particles than in small dense HDL3 particles per millilitre of plasma.[38] The decrease in SM content in HDL particles associated with smaller and more dense HDL molecules was shown to correlate with the atheroprotective functionality of HDL, such as cholesterol efflux capacity, antioxidative activity toward LDL oxidation, antithrombotic activity in human platelets, anti-inflammatory activity and anti-apoptotic activity.[38] In another study, anti-apoptotic and anti-oxidative activities of small dense HDL particles were associated with depletion in SM within the same molecules.[39] This association between structural reformation of HDL molecules and their anti-atherogenic properties (mainly their capacity for cholesterol efflux) was explained by the negative impact that SM had on molecular surface fluidity and lecithin:cholesterol acyltransferase activity.

LDL present in atherosclerotic plaques has higher SM levels than plasma LDL; this is mainly due to de novo synthesis in the aorta.[37,40] Furthermore, early studies have shown that advanced aortic wall atherosclerosis is characterised by increased SM proportion.[41] Schissel et al. have shown that in rabbit aorta and human atherosclerotic lesions, arterial-wall SMS activity hydrolyses the SM of retained LDL leading to LDL aggregation, an important event regarding the initiation and progression of atherosclerosis.[37,42]

Valvular Disease

Recently, the presence of intra-leaflet haemorrhages and the associated increase in intra-leaflet lipid accumulation has been recognised as a novel mechanism for valve tissue degeneration. In a study by Lehti et al., higher SM:phosphatidylcholine ratio as well as higher proportions of lysophosphatidylcholine and unesterified cholesterol were found in lipid particles isolated from patients with stenotic aortic valves compared with non-stenotic counterparts.[43] Under the same notion, using a metabolomics approach in aortic tissue, Doppler et al. showed that levels of SM were significantly increased in patients with bicuspid aortic valve disease undergoing ascending thoracic aorta surgery and with tricuspid aortic valve associated aortic dissection compared with controls.[44]

Niemann–Pick disease is a rare lysosomal storage disease caused by deficient activity of acid SMS and the accumulation of SM within cells; these abnormalities occur especially in the monocyte–macrophage system. In a prospective, cross-sectional survey study, McGovern et al. demonstrated that most patients with Niemann–Pick disease had low HDL cholesterol, high total cholesterol, high triglyceride and high LDL levels compared with control subjects.[45] Two-dimensional echocardiograms performed during the study, showed abnormalities in the majority of patients with Niemann–Pick disease, most commonly mitral valve regurgitation. Furthermore, several published cases of patients with Niemman–Pick disease report severe mitral regurgitation as a clinical manifestation of the disease, thus suggesting an association between progressive accumulation of SM in lysosomes and degeneration of cardiac valves.[46]

Cardiomyopathies

Ceramides are produced by either de novo synthesis or hydrolysis of SM catalysed by acid and/or neutral SMS.[47] Accumulation of cardiac ceramides in the post-ischaemic heart is mediated by acid SMS and not by de novo sphingolipid synthesis.[48] Therefore, the presence of ceramides in the post-ischaemic myocardium is most likely due to SM degradation by SMS, thus suggesting a detrimental role of SM in a wide variety of cardiomyopathies, especially in ischaemic cardiomyopathy. However, a recent study in mice showed that inhibition of acid SMS does not result in improved heart function or survival after an induced MI despite reducing ischaemia-induced ceramide accumulation.[49] In a rodent model of lipotoxic cardiomyopathy, mice exhibited increased lipid uptake and oxidation, ceramide accumulation and a dilated cardiomyopathy, with decreased functional cardiomyocyte shortening.[49] In contrast, we must emphasise that within this cardiolipotoxic model, ceramides were de novo synthesised within the myocardium and that the mechanism of this cardiac dysfunction was not clear.

As part of the Alberta Heart Failure Etiology and Analysis Research Team (HEART) project, ambulatory patients with clinical diagnosis of heart failure (HF; with preserved or reduced ejection fraction) and age-matched non-HF controls were selected for metabolomic analysis.[50] Compared with non-HF control subjects, patients with HF with preserved ejection fraction had lower levels of phosphatidylcholines and SMs. Furthermore, in a recent study enrolling patients with either ischaemic or non-ischaemic cardiomyopathy, healthy controls and patients with pulmonary diseases, three specific metabolomic features belonging to the lipid classes of SMs, triglycerides and phosphatidylcholines together with N-terminal pro-B-type natriuretic peptide (NT-proBNP) distinguished patients with HF from healthy controls.[51] In addition, the diagnostic accuracy of this combination was significantly superior compared with the diagnostic accuracy of NT-proBNP alone.[51]

Conclusion

Dysregulation of sphingolipid synthesis and transport is associated with cardiovascular disease, diabetes and other metabolic disorders. However, further studies are needed to identify the molecular and pathophysiological pathways by which certain sphingolipids species are associated with different pathologies.