Sphingolipids such as sphingosine-1-phosphate (S1P), ceramide, or sphingomyelin are essential constituents of plasma membranes and regulate many (patho)physiological cellular responses inducing apoptosis and cell survival, vascular permeability, mast cell activation, and airway smooth muscle functions. The complexity of sphingolipid biology is generated by a great variety of compounds, diverse receptors, and often antagonistic functions of different sphingolipids. For instance, apoptosis is promoted by ceramide and prevented by S1P, and pulmonary vascular permeability is increased by S1P2/3 receptors and by ceramide, whereas S1P1 receptors stabilize barrier integrity. Several enzymes of the sphingolipid metabolism respond to external stimuli such as sphingomyelinase isoenzymes that are activated by many stress stimuli and the sphingosine kinase isoenzymes that are activated by allergens. The past years have provided increasing evidence that these processes contribute to pulmonary disorders including asthma, chronic obstructive pulmonary disease, acute lung injury, and cystic fibrosis. Sphingolipid metabolism offers several novel therapeutic targets for the treatment of lung diseases such as emphysema, asthma, cystic fibrosis, respiratory tract infection, sepsis, and acute lung injury.
Brief Overview of Sphingolipid Biochemistry
Tools to Study Sphingolipids
Apoptosis and Proliferation
Sphingolipids in Pulmonary Disease
Niemann-Pick Disease (Acid Sphingomyelinase Deficiency)
Sphingolipids in Acute Lung Injury
Sphingolipids and Pulmonary Emphysema
Lung Ceramide and Cystic Fibrosis
Viral Infections of the Respiratory Tract, Ceramide, and Rafts
The sphingolipids have certainly lived up to their name. Derived from the Greek word sphinx, meaning enigmatic, many of their functions remain puzzling. Initially recognized primarily as structure-bearing elements of biological membranes, sphingolipids are now known to regulate key physiological processes such as apoptosis, innate and acquired immunity, vascular permeability, smooth muscle tone, and to contribute to various pathological conditions such as pulmonary edema, emphysema, cystic fibrosis, or pneumonia.
The central role of lipids in many biological processes has not been appreciated until recently, perhaps because the focus of biological research was on the genome and the proteome. A typical mammalian cell contains 109 lipid molecules, which show an enormous diversity. For instance, lipids differ in their headgroups and the length of the acyl chains, and these differences result in different structural effects in the membrane bilayer and/or functions in cell signaling. Thus, even within a certain lipid class, the functions of similar molecules might be very different. This illustrates the immense diversity of lipids, and one of the future challenges will be the systemic analysis of the function of different lipids. The purpose of this perspective is to review the emerging significance of sphingolipids, in particular ceramide and sphingosine-1-phosphate (S1P), for the (patho)physiology of the lung.
Sphingolipids are amphiphatic molecules formally derived from sphingosine. Its phosphorylation leads to S1P and its acylation to ceramide and, after coupling with phosphocholine, to sphingomyelin (Figure 1). The reverse pathway (i.e., the production of ceramide from sphingomyelin by sphingomyelin phosphodiesterases [gene symbol: SMPD], also known as sphingomyelinase isoenzymes) is of particular pathophysiological relevance. Hundreds of sphingolipid species are known (1, 2), but two of the best-studied sphingolipids are S1P and ceramide. S1P mediates cell survival and proliferation, whereas ceramide promotes apoptosis, differentiation, and cell-cycle arrest. These findings have led to the concept of the sphingolipid rheostat (3) as a critical regulator of cell fate, particularly in response to all kinds of stress (4).
Increased ceramide production occurs in many situations associated with cellular stress, such as ischemia, radiation, oxidative stress, chemotherapeutic agents, or exposure to oxidized low-density lipoproteins, and is also induced by proinflammatory mediators such as tumor necrosis factor (TNF), IL 1, and platelet-activating factor (PAF) (5–15). Ceramide may exert its actions via an alteration of the physical membrane properties (see 2.5. Membrane Microdomains) and/or by specific interaction with intracellular targets such as protein phosphatases, phospholipase A2 (PLA2), cathepsin D, or ion channels (see 2.5. Membrane Microdomains). Ceramide can be generated de novo, from sphingosine or by sphingomyelinase activity. The de novo synthesis, commencing with the condensation of serine with palmitoyl CoA, is catalyzed in the cytosplasmatic leaflet of the endoplasmatic reticulum. The hydrophobic ceramide is then transported by the ceramide-binding protein CERT to the Golgi apparatus (16). The synthesis of ceramide from sphingosine involves hydrolysis of glycosphingolipids and a retrograde activity of the enzyme ceramidase (17). The release of ceramide from sphingomyelin is catalyzed by several sphingomyelinase isoenzymes: the lysosomal acid sphingomyelinase (A-SMase), the secretory A-SMase, the Mg2+ dependent neutral sphingomyelinase isoenzymes (N-SMase), and the alkaline SMase (18).
The lysosomal A-SMase mediates sphingolipid turnover and degradation. The enzyme interacts with lipid membranes via its sphingolipid activator domain and also binds to the mannose-6-phosphate receptor. Both lysosomal and secretory A-SMases are derived from the same gene (SMPD1) and are Zn2+ dependent enzymes. However, whereas the lysosomal form is already tightly bound to the cation, the secreted form is stimulated by zinc (19). It appears likely that the extracellular A-SMase acts on plasmalemmal sphingomyelin because sphingomyelin is almost exclusively localized to the outer membrane leaflet (20). Endothelial cells are a wealthy source of A-SMase (21), A-SMase is present in most body fluids (22), and serum concentrations are raised in a number of disorders, many of them inflammatory (see Table 1).
|Chronic heart failure||Serum||Human||2007||(223)|
|Depression||Peripheral blood monocytes||Human||2005||(224)|
|Ceramide (intratracheal)||Bronchoalveolar lavage||Mouse||2005||(198)|
|LPS inhalation||Lung tissue||Mouse||2008||(142)|
|Repeated lung lavage||Lung tissue, serum||Piglet||2008||(142,143)|
|Perfusate of isolated rat lung||Rat|
|TNF, IL 1 (intravenous)||Serum||Human||2000||(140)|
The ubiquitously expressed N-SMase1 (SMPD2) is Mg2+-dependent, localizes to the endoplasmic reticulum (ER), and is reversibly inhibited by oxidized glutathione, but its role remains poorly defined; it was suggested to act mainly as a lysoPAF phospholipase C (PLC). N-SMase2 (SMPD3) is expressed in the Golgi, but probably also in the plasma membrane and appears to be the major N-SMase isoenzyme that becomes activated by oxidative stress (18, 23, 24). Recently, N-SMase3 (SMPD4) was cloned that is highly expressed in the heart, but also in the lungs and other organs (25).
Ceramide may further be converted by ceramide kinase (Cerk) to ceramide-1-phosphate that appears to have its own specific actions, such as activation of cytosolic phospholipase A2 (cPLA2) and regulation of apoptosis (26, 27). A recent study demonstrated a critical role of ceramide-1-phosphate in neutrophil homeostasis and showed strikingly reduced neutrophil numbers in the blood and spleen of Cerk−/− animals (28).
Sphingosine cannot be synthesized de novo, but it is derived from N-deacetylation of ceramide by ceramidase or by hydrolysis from plasma membrane sphingolipids. While the initially suspected function of S1P as an intracellular messenger is still under debate (4), the discovery that the once orphaned G protein–coupled edg (endothelial developmental gene) receptors represent S1P receptors (S1P1–5) has greatly stimulated research in this area. S1P is now recognized as a ubiquitous regulator of cell proliferation and survival, angiogenesis and endothelial barrier functions, cell mobility and chemotaxis, cytoskeletal organization, cellular calcium homeostasis, cell-to-cell contacts, and adhesion (4, 29, 30). There are several examples showing that the different S1P receptor subtypes may act antagonistically, such as in chemotaxis (stimulation by S1P1 and S1P3, inhibition by S1P2) (31) or in endothelial barrier functions (see below). Another interesting antagonism seems to exist between S1P and ceramide, with ceramide being proapoptotic and S1P being antiapoptotic (4). In view of the consequences for cell growth and differentiation, and in consideration of the highly dynamic S1P and ceramide levels, this antagonism has been termed the sphingolipid “rheostat” (4). S1P4 and S1P5 are enriched in lymphoid tissues, and in brain and spleen tissue, respectively (30); these receptors have not been implicated in pulmonary (patho)physiology. It should be noted that the S1P receptors show a high homology to the receptors for lysophosphatidic acid, LPA1/2/3, (32) and some receptors may even recognize both S1P and lysophosphatidic acid (33).
S1P is synthesized by two sphingosine kinase isoenzymes (SphK1, SphK2) and degraded by S1P phosphatases, lipid phosphate phosphatases, or S1P lyase (4). Platelets lack S1P lyase, implicating these cells as sources of the relatively high (0.5 μM) serum S1P levels (34), where it is bound to albumin and high-density lipoproteins (35). However, other cells such as erythrocytes, neutrophils, mast cells, mononuclear cells, and endothelial cells may also secrete S1P (36–38). In any case, control of extracellular S1P seems to be complex and apparently also involves export of SphK-1a to produce S1P outside the cells (39), degradation by lipid phosphate phosphatases (which are integral membrane proteins facing the extracellular side), and reuptake (40). SphK activity is stimulated by a variety of stimuli (41) including muscarinic M2 receptor agonists (leading to airway constriction ), activation of IgE receptors (43, 44), histamine, several growth factors, and various cytokines (29).
The study of sphingolipids is fraught with technical problems that need to be taken into account when following progress in this area. One particular problem is the significant hydrophobicity, particularly of ceramide, that makes it difficult to identify specific intracellular binding partners. Another shortcoming is the lack of quick and efficient ways to determine sphingolipid concentrations in biological samples. Although several ceramide antibodies are available, some need to be used with caution (45).
The nearly complete insolubility of long chain ceramides (>C16) in aqueous solutions also poses problems for studying the role of exogenously added ceramide. Thus, many studies using exogenous ceramide have used C2- or C6-ceramide, although their biophysical properties differ from those of long chain ceramides (46). In addition, it seems possible that specific ceramide species have specific roles, such as C16- and C24-ceramide in different stages of apoptosis (47). Therefore, the results from experiments with short chain ceramides need to be interpreted cautiously. The use of extracellular A-SMase is another approach to generate ceramide, at least in cell culture, although it is difficult to mimic transient changes of ceramide, to increase intracellular ceramide, and to control the amount of ceramide with this treatment.
Ceramide synthesis can be inhibited at several levels: inhibitors of ceramide synthase, A-SMase and N-SMase are available (Table 2). Ceramide synthase is inhibited by fumonisin B1, a fungal toxin that is carcinogenic, neurotoxic, and causes pulmonary edema (48). Imipramine, D609, and NB6 are all very effective inhibitors of A-SMase–dependent ceramide release (18). However, the xanthogenate D609 and the tricyclic antidepressant imipramine do not inhibit A-SMase directly. Imipramine stimulates proteolysis of A-SMase (49) but also down regulates acid ceramidase (50). D609 interferes with the activation of A-SMase, although its mode of action remains somewhat controversial, and it may also block sphingomyelin synthase (9, 51,52). N-SMase is inhibited by several agents (Table 2), but the specificity of these inhibitors requires further characterization.
|Fumonisin B1||Ceramide synthase||I||Inhibition of protein biosynthesis at higher concentrations||(48, 227)|
|D609, imipramine, amitriptyline, NB6 carnithine, SR33557||A-SMase||I||See text; imipramine and amitryptiline are also known as tricyclic antidepressants; SR33557 is used as a chemosensitizer||(18)|
|Scyophostatin, SMA7, 3 O-methylsphingomyelin, GW4869, C11AG, manumycin A||N-SMase||I||See text||(18,70,228–230)|
|2-acetyl-4-tetrahydroxybutylimdazole||S1P lyase||I||Used as a food colorant||(231)|
|N,N-dimethylsphingosine, L-threo-dihydrosphingosine, SKI 2||Sphingosine kinase||I||See text||(41,53,54)|
|N-oleoylethanolamine, B13||Ceramidase||I||See text||(56)|
|FTY720 (agonist)||S1P1, S1P3, S1P4, S1P5||A||Needs activation by sphingosine kinase; see text||(57)|
|Dihydro S1P, SEW2871 (agonist)||S1P||A||See text||(64)|
| JTE 013||S1P2||I||—||(66)|
Sphingosine kinase isoenzymes are inhibited by sphingosine analogs that appear to be fairly nonspecific and should not be accepted as single evidence for an involvement of SphK (41, 53). Another recently described SphK inhibitor is SKI-2 (54), although—at least with respect to neutrophils—there is an unresolved discrepancy between results with this inhibitor and SphK-deficient mice (53, 55). Ceramide analogs such as B13 and N-oleoylethanolamine are ceramidase inhibitors and can increase apoptosis at least under some circumstances (56).
The action of S1P is frequently blocked by interfering with S1P1–5 receptors. Much attention has been given to FTY720 (fingolimod), a prodrug that needs to be activated (phosphorylated) by SphK2, before it acts on all S1P receptors except S1P2 (57). It is currently under development as an immunosuppressive agent for ailments such as renal graft rejection and multiple sclerosis, and in experimental animal models FTY720 has provided protection in models of acute lung injury (ALI) (58) and asthma (59, 60). Recent studies, however, have shown that unphosphorylated FTY720 possesses S1P-receptor independent effects in regulating endothelial cell permeability (61), in interacting with the evolutionary related cannabinoid receptors (62), and in inhibiting cPLA2 with a broad effect on eicosanoid synthesis (e.g., in the mast cells' response to antigen ). Compared with FTY720, SEW2871 appears to be relatively specific for S1P1 receptors with less S1P3-mediated cardiovascular side effects (64). So far, only two S1P-receptor antagonists are known (i.e., W146 blocks S1P1 receptors  and JTE 013 S1P2 receptors ).
Sphingolipids are attractive targets to treat many disorders including acute and chronic lung diseases, as discussed below. In particular, the specific targeting of S1P receptors and the inhibition of ceramide during stress responses appears to be attractive areas for pharmacological developments. However, wide clinical use will require specific drugs with fewer side effects than drugs currently available.
Several knockout mice have been generated, and, except for A-SMase–null mice (Smpd1−/−), they do not show developmental alterations in the lungs. Smpd1 −/− mice are fertile, but suffer from progressive lysosomal storage in mononuclear cells and in visceral organs, elevated blood cholesterol levels, increased sphingomyelin levels in liver and brain, the neurovisceral form of Niemann-Pick disease (see below), and death by 8 months of age (67, 68). In addition, these mice show elevated levels of MIP1α, growth hormone, and total iron (69). N-SMase1–deficient mice (Smpd2−/−) have no defects in sphingomyelin storage and a normal (>2 yr) life span (70). N-SMase2 deficiency (Smpd3−/−) results in severe retardation of late embryonic and postnatal growth and bone fragility, indicating a pivotal function of Smpd3 in the control of the hypothalamus–pituitary growth axis and bone formation (71).
S1P1-null mice are embryonic lethal because of incomplete vascular maturation (72), whereas S1P2 and S1P3 knockout mice are viable and fertile; S1P2/S1P3 double knockout mice show perinatal lethality (73, 74). S1P2−/− mice develop seizures during 3 to 7 weeks of age as a result of hyperexcitable neurons (75) and are deaf because of vascular disturbances in the inner ear (76). Sphk1-null mice are viable, fertile, and without any obvious abnormalities, although their serum S1P levels are only about 50% (77). SphK2-null mice are viable, but subfertile; their serum S1P concentrations are only 25% of wild type mice (78). Neutrophil functions in SphK1- and SphK2-deficient mice are normal (53). Double knockout SphK1/ SphK2 mice lack S1P and show severe defects in angiogenesis and neurogenesis (79).
Mutations in the acid ceramidase gene (Asah1) result in Farber Lipogranulomatosis, a fatal human genetic disorder, and Asah1-null embryos do not survive beyond the 2-cell stage because proapoptotic ceramide cannot be removed (80). No specific pulmonary phenotype has been described in patients with Farbers disease (56). Serine palmitoyltransferase deficiency leads to hereditary neuropathy (81).
Many growth factors induce S1P, whereas many agents that cause apoptosis invoke ceramide by several different mechanisms, as proapoptotic ceramide may be derived from A-SMase, N-SMase or even de novo synthesis (82, 83). Because ceramide is activated by so many different stimuli that do not cause apoptosis, it is important to question how cells can avoid apoptosis in the presence of increased ceramide levels. Some cells produce both S1P and ceramide, to the effect that S1P prevents ceramide-induced apoptosis that would occur otherwise (84). Other mechanisms that protect cells from ceramide-induced apoptosis are the strong expression of acid ceramidase in alveolar macrophages (85) or the inhibition of caspase 3 by interaction with A-SMase in airway epithelial cells (86).
Ceramide appears to cause apoptosis by several mechanisms such as the recruitment of death receptors to lipid rafts (87) as well as by the direct activation of protein phosphatase 2a and cathepsin (82, 88). The proapoptotic properties of ceramide are well illustrated by the fact that radiation increases pulmonary ceramide levels and that pulmonary endothelial cells in Smpd1−/− mice are resistant to radiation-induced apoptosis as shown by TdT-mediated dUTP biotin nick end labeling (TUNEL) assay (15). TUNEL is a frequently-used apoptosis assay, although it is not entirely specific for apoptosis (89). With reference to its apoptotic properties, ceramide has been called the “tumor-suppressor lipid” (82). Of note, sphingosine also has proapoptotic properties that appear to be independent of ceramide (82).
The antiapoptotic actions of S1P are currently explained by its activation of nuclear factor (NF)-κB, Akt kinase and the intracellular inhibition of ceramide synthesizing enzymes (82, 90). For cancer cell lines (e.g., HEK293 and A549) S1P is an important survival factor and inhibition of S1P synthesis or action sensitizes these cells for some chemotherapeutics (91, 92). SphK1 is overexpressed in human tumor lung tissue (93). Thus, inhibition of sphingosine metabolism has been suggested as a means to sensitize resistant tumors. Interestingly, only S1P derived from SphK1 appears to be antiapoptotic, whereas overexpression of SphK2 suppresses growth and favors apoptosis (92). S1P is also mitogenic for transformed cell lines (92) and for human lung fibroblasts (94).
Unlike those suggested by the classical fluid mosaic model (95), cell membranes show a high degree of organization to which sphingolipids make an important contribution. Sphingolipids have a higher melting temperature than most other membrane lipids and thus increase membrane rigidity. Besides sphingolipids, cellular membranes are predominantly composed of cholesterol and (glycero)phospholipids. Sphingolipid headgroups interact with each other and the hydroxyl group in the cholesterol molecule via hydrophilic interactions, whereas the alkyl chains and the sterol ring system of cholesterol exhibit hydrophobic van der Waals interactions (96). The tight interactions result in a separation of sphingolipids and cholesterol from other phospholipids in the cell membrane and the formation of very small distinct domains in the cell membrane enriched in sphingolipids and cholesterol (96, 97), which are termed “rafts.” Cholesterol molecules seem to fill the voids between the bulky sphingolipids and thus promote the structure of these domains (98). The formation of these small membrane domains was concluded from biophysical experiments using artificial membranes and the detergent-resistant extraction of these domains in biochemical studies (96, 97, 99).
The generation of ceramide within lipid rafts was shown to result in the formation of larger membrane domains (100–102). In general, rafts serve as anchoring points of the cytoskeleton, as gathering places for receptors, as starting points of signaling cascades, and as locations for the uptake of microorganisms; although it should be pointed out that many protein–protein or protein–lipid interactions and receptor-mediated signaling processes are independent of rafts. In addition, many different rafts with varying lipid compositions exist, which may result in different modes of action. Although the exact functions of many rafts still require definition, it may be safe to state that ceramide-enriched membrane rafts seem to be predominantly involved in cellular stress responses. Caveolae are one particular form of lipid raft. In the lungs, caveolae are particularly abundant in endothelial cells (10,000–30,000 caveolae/cell, or ∼15% of the total cell volume), alveolar type II cells, fibroblasts, and smooth muscle (103, 104).
Hydrolysis of sphingomyelin by sphingomyelinases results in the formation of ceramide, which dramatically alters the biophysical properties of biological membranes. Ceramide molecules interact with each other resulting in the formation of tightly packed small ceramide-enriched membrane domains (97, 105). These microdomains have the tendency to spontaneously fuse to larger domains called ceramide-enriched membrane platforms (101, 106, 107). Ceramide-enriched membrane domains are formed after stimulation of several receptors and application of stress stimuli, for instance, after stimulation via CD95 (107), CD40 (108), DR5 (109), FcγRII (110), CD14 (111), also after infection with Pseudomonas aeruginosa (112), Staphylococcus aureus (113), Neisseria gonorrhoeae (114), rhinovirus (115), after application of stress stimuli such as γ-irradiation (15), UV light (116–118), cisplatin (12), Cu2+ treatment (119), and, they are formed as well, in some conditions of developmental death (120, 121).
Ceramide-enriched membrane domains may have several functions. First, the composition and fluidity of ceramide-enriched membrane domains differ from the surrounding areas in biological membranes, which may permit a trapping and aggregation of receptors and/or cellular-signaling molecules within these membrane domains. Clustering of activated receptors in ceramide-enriched membrane domains was demonstrated for CD95, DR5, or CD40 (107–109), but many more receptors may use the mechanism of clustering to reach a very high density in circumscribed areas of the cell membrane. The reorganization of a receptor within ceramide-enriched membrane domains may also result in a preferential interaction with downstream signaling molecules, whereas inhibitory molecules might be excluded from these domains, although this concept still requires experimental proof. One of the future challenges will be to understand how molecules cluster in ceramide-enriched domains and how specific signaling pathways are initiated and/or amplified by the formation of ceramide-enriched membrane domains. Biophysical and electron microscopy studies will be required to characterize the spatial structure of these domains.
Second, ceramide was shown to directly regulate several molecules including cathepsin D (122), phospholipase A2 (123), kinase suppressor of Ras (identical to ceramide-activated protein kinase) (124), ceramide-activated protein serine-threonine phosphatases (125), protein kinase C isoforms (126) and c-Raf-1 (127). The details and the specificity of the interaction of ceramide with these molecules are presently unknown except for cathepsin D that binds ceramide via a short domain in the cathepsin D molecule.
Third, ceramide has been shown to regulate several ion channels including the Kv1.3 channel and calcium release-activated calcium channels (128, 129). Further, ceramide molecules were shown to trigger the formation of pores, at least in the outer mitochondrial membrane (130). These channels might be important for the induction of apoptosis, although it is unknown whether ceramide pores are also formed in vivo. The regulation of ion channels by ceramide is a poorly investigated field, although its potential for many physiological and pathophysiological processes appears immense.
In summary, ceramide-enriched membrane domains serve the temporal and spatial organization of signaling molecules to regulate multiple cell functions.
The crucial role of sphingolipids for normal cell functions and the pathogenesis of several pulmonary diseases is becoming increasingly recognized (Table 3). The enzyme most frequently associated with human ailments is A-SMase, elevated levels of which are found in various diseases (Table 1) and may contribute to their pathogenesis (Table 4). N-SMase has so far only been associated with emphysema, whereas sphingosine kinase isoenzymes have been implicated with immune-cell regulation.
|Inflammation and infection|
|Emphysema||Ceramide||—||A-SMase, ceramide synthase, serine palmitoyl transferase||(198)|
|Mast cell activation||S1P||S1P1||SphK1||(44)|
|Dendritic cell trafficking||S1P||S1P1||—||(232)|
|Dendritic cell activation||S1P3||—||(137)|
|Pathogen uptake||Ceramide-rich platforms||—||A-SMase||(112)|
|Airway smooth muscle|
|Airway contraction||S1P||S1P2, S1P3||SphK||(42,183)|
|▴ Apoptosis||Ceramide||—||A-SMase, N-SMase, de novo pathway||(82,83)|
|—||A-SMase knockout||Niemann Pick disease||2003||(234)|
|PAF||A-SMase knockout, imipramine, D609||▾Pulmonary edema||2004||(7)|
|Endotoxemia||A-SMase knockout,||▾Endothelial cell apoptosis in lung||1998||(142,148)|
|Endotoxemia||Imipramine, D609||▾Pulmonary edema||2004||(7)|
|Endotoxin inhalation||A-SMase knockout, imipramine||Cell counts in BAL||2008||(143)|
|Acid instillation||D609||▾Pulmonary edema||2004||(7)|
|Repeated lavage||Imipramine||▾Pulmonary edema||2008||(143)|
|Cystic fibrosis||Amitryptyline, A-SMase heterozygosity||▴Lung functions||2008||(208)|
|P. aeruginosa infection||A-SMase knockout||▴Bacterial growth||2003||(113)|
The rare Niemann-Pick disease (NPD) type A and B (recently reviewed in ), is the consequence of aberrant and at least partly inactive A-SMase variants. Type A NPD, which is lethal within the first 3 years of life, is characterized by neurological deficits, enlarged livers and spleens, and often by recurrent pulmonary infections. Patients with Type B NPD frequently show enlarged livers and spleens, proatherogenic alterations, and interstitial lung disease (132, 133), but otherwise the symptoms may vary. This variation is explained by over 100 known mutations of the SMPD1 gene (133, 134). In recent studies more than 90% of all patients with Type B NPD showed interstitial lung disease and more than 70% had abnormal lung function tests, although these clinical findings did not correlate well (132, 133, 135). In addition, foamy alveolar macrophages (Niemann-Pick cells) are frequently observed in patients with NPD (135) as they are in aged A-SMase–deficient animals (69). These findings show that in NPD pulmonary involvement is common. Currently, no specific therapy is available, but after a promising experimental study (136), a clinical trial (NCT00410566) examining A-SMase replacement therapy has been initiated.
Most studies in this area have focused on the role of A-SMase and S1P receptors on endothelial cells. These studies are reviewed below. More recently, further cell types have begun to be studied in models of ALI. Such studies showed that S1P3 receptors play a particularly noteworthy role in promoting pulmonary inflammation in that they stimulate the late phase (18 h) of sepsis by propagating the release of IL-1β and tissue factor from dendritic cells (137). Thus, sphingolipids appear to regulate ALI at different levels, but this needs further exploration.
The accumulation of experimental and clinical evidence indicates the critical role of the secretory A-SMase in the pathogenesis of ALI and sepsis (Tables 1 and 4). A-SMase may be released from endothelial cells but also from other cells such as activated platelets (138). Mediators of sepsis and acute respiratory distress syndrome (ARDS), such as PAF (7) and TNF (139), elevate A-SMase activity in vitro (see 2.1. Ceramide) as well as in vivo in serum or bronchoalveolar lavage (BAL) samples. In addition, IL-1 was suggested to mediate the LPS-induced increase of circulating A-SMase activity (140). A-SMase activity is increased in animal models of sepsis and ALI such as in the serum during endotoxemia (140, 141) and in the lungs after LPS inhalation (142) or following repeated lung lavage (142, 143). Most importantly, circulating A-SMase activity is increased in patients with sepsis (141), and elevated systemic A-SMase levels as well as ceramide levels in plasma or monocytes correlate with mortality (141, 144, 145).
The relevance of these intriguing findings has been probed in several experimental settings using A-SMase–null mice and pharmacological inhibitors. In the LPS model, pulmonary edema formation is attenuated by D609 (7), pulmonary inflammation by imipramine (142), and mortality by D609 (146), NB6 (141) and in A-SMase–null mice (147). In acid-induced ALI, D609 treatment attenuates pulmonary edema and improves oxygenation (7). And finally, imipramine ameliorates edema formation and advances oxygenation in ALI induced by repeated lung lavage when given together with surfactant; this beneficial effect of imipramine was remarkably long-lived and lasted for 24 hours (142).
There are several possible explanations for these findings, although the mechanism is far from clear. Early work has shown that D609 prevents TNF-induced expression of adhesion molecules in the pulmonary vasculature and leukocyte infiltration (146). Because leukocytes are known to be critical in the pathogenesis of ALI, this could explain some of the findings, although the link between A-SMase and leukocyte activation remains elusive. Furthermore, A-SMase–derived ceramide can increase vascular permeability and promote edema formation (see 3.2.2. Vascular Permeability) (7). Taken together, these findings suggest that A-SMase may be a useful therapeutic target in the treatment of ALI and sepsis.
Ceramide was also suggested to injure pulmonary endothelial and hepatic cells during sepsis. Inhibition of ceramide formation by genetic A-SMase ablation or by NB6 attenuated LPS-induced apoptosis (TUNEL-positive cells) in lung and liver and also mortality (141, 147). The hypothesis that the correlation between apoptosis and mortality under these conditions hints at a causal relationship is very intriguing but certainly requires further evidence, such as additional markers of apoptosis and additional experiments, to further define the respective roles of apoptosis for cell type specific death, tissue repair, and failed apoptotic cell removal (efferocytosis) (148) in these models. Clearly, depending on the cell type, apoptosis of lung cells may have various implications other than ALI (for review see References 149–151) such as emphysema (see 3.5. Sphingolipids and Pulmonary Emphysema), fibrosis (152) or prolonged survival of neutrophils (153). In many of these instances, the contribution of sphingolipids to the apoptotic process is largely unknown.
Formation of pulmonary edema due to the loss of endothelial and epithelial barrier functions is the hallmark of ALI. Endothelial and epithelial permeability are regulated by sphingolipids in numerous ways: barrier disruptive are ceramide, endothelial S1P2 and S1P3 receptors and epithelial S1P3 receptors; barrier stabilizing are endothelial S1P1 receptors (Table 3). These receptors couple to Rac-GTPase or to Rho-kinase that act in an antagonistic fashion to increase or decrease barrier functions, respectively (154, 155).
It has long been known that S1P stabilizes endothelial adherens junctions (156). This barrier stabilizing function appears to be mainly mediated by S1P1 receptors and involves activation of Rac GTPase dependent cytoskeletal reorganization and focal adhesion assembly (154). Initially, in the lungs this was demonstrated by the ability of S1P1 receptor agonists to stabilize endothelial barrier functions (157). Subsequently, S1P was shown to protect against LPS- and ischemia/reperfusion-induced pulmonary edema and ALI (58, 158, 159). More recently, using the novel S1P1-receptor antagonist W146, it was demonstrated that blockade of S1P1 receptors causes pulmonary edema, suggesting that persistent activation of S1P1 receptors is required for maintaining the barrier function of the normal pulmonary endothelium (63).
These actions are opposed by activation of S1P2 receptors, causing Rho-kinase–dependent inhibition of Rac and disruption of adherence junctions, a process that contributes to pulmonary edema induced by H2O2 in rats (160). Preferential Rho kinase activation was also demonstrated for S1P3 receptors that mediate increased epithelial permeability in mice (161) and endothelial permeability in human pulmonary endothelial cells (162, 163). An important novel aspect in endothelial S1P receptor signaling is that these receptors are subject to transactivation. S1P1 receptors are transactivated by endothelial protein C receptors, by protease activated receptor-1 receptors (ligand: low thrombin concentrations) and CD44s (ligand: high molecular weight hyaluronan) leading to barrier function enhancement (162, 164, 165); S1P3 receptors are transactivated by CD44v10 (ligand: low molecular weight hyaluronan) and μ-opoid receptors (ligand: morphine) leading to increased endothelial permeability (164, 163). All these findings point to a highly important role of S1P receptors in the control of endothelial and epithelial barrier functions.
Not only S1P, but also ceramide is involved in the regulation of vascular and epithelial permeability. Both the instillation of ceramide in rat airways (139) and perfusion of C2 ceramide in isolated rat lungs cause edema (7). In addition, ceramide increases vascular permeability in human pulmonary microvascular endothelial cells in culture (166). Anticeramide specific antibodies blocked PAF induced edema in isolated rat lungs and in vivo (7). PAF is a mediator of ALI and causes pulmonary edema within 5 to 15 minutes (167). In isolated lungs, but also in macrophages and erythrocytes, PAF rapidly stimulates ceramide formation by the activation of the A-SMase (6, 7, 168). Inhibition of the A-SMase partly reduces PAF-induced edema formation in the lungs (7, 167). The other part of PAF-induced pulmonary edema is mediated by PGE2 (167, 169). The A-SMase– ceramide pathway also offers a novel approach to explain the antiedematogenic properties of steroids, insofar as steroids inhibit both A-SMase activity (170) and PAF-triggered ceramide release (7). Of note, endothelial cell apoptosis appears not to be involved in the ceramide-mediated increase in vascular permeability (166, 167). The molecular mechanisms of how ceramide mediates pulmonary edema are unknown. However, the fact that L108 (a blocker of phosphatidyl inositol–specific PLC and thus IP3-dependent calcium release) prevents both PAF-induced edema formation (171) and ceramide-induced hyperpermeability in endothelial cell monolayers (166) indicates that ceramide may increase intracellular calcium, although this is speculative.
In summary, pulmonary edema caused by increased vascular permeability as it occurs in ARDS might be attenuated by the activation of S1P1 receptors or by blocking S1P2 receptors, S1P3 receptors, or A-SMase. Interesting open questions for future studies relate to the interaction between A-SMase and S1P receptors and the relative importance of the different S1P receptors in the control of barrier integrity under pathophysiological conditions.
Allergic asthma is a chronic inflammatory disease characterized by a predominance of Th2 lymphocytes, recurrent activation of mast cells by activation of FcεR1 receptors, recurrent airway smooth muscle contraction, airway hyperresponsiveness, and airway remodeling, all of which appear to be regulated by sphingolipids (172), in particular by S1P, which is increased in asthmatic airways (173).
S1P regulates trafficking of lymphocytes. It stimulates immature dendritic cells (DC) (174) but prevents the egress of lymphocytes from secondary lymph nodes and thymus (57, 175). S1P also favors Th2 lymphocyte–dominated immunity (174).
Upon crosslinking, FcεR1 receptors move into lipid rafts (176). Activation of the FcεRI receptor activates SphK2 that, in turn, produces S1P from sphingosine. Both the decrease in sphingosine and the elevation in S1P are thought to contribute to the critical increase in intracellular Ca2+, cytokine production, and degranulation (177). S1P may act intracellularly, but is also released (by ABC transporters) and activates S1P1 and S1P2 receptors on mast cells in an autocrine and paracrine fashion (178). Activation of S1P1 receptors induces mast cell chemotaxis, activation of S1P2 receptors degranulation, and at higher S1P concentrations, also inhibition of chemotaxis. In addition, mast cell responsiveness is up-regulated by extracellular S1P produced by SphK1 in nonmast cells (177). In addition, S1P (derived from SphK) appears to contribute to eosinophilic inflammation in the murine ovalbumin model of asthma (179).
A comprehensive model proposes that low antigen concentrations are signaled via S1P1 receptors to mediate mast cell chemotaxis, whereas the mast cell response to higher antigen concentrations predominantly involves S1P2 receptors, thus terminating chemotaxis and stimulating degranulation (178, 180). Mast cell activation by S1P is antagonized by ceramide and sphingosine, providing another example of the sphingolipid “rheostat” (178). For example, ceramide inhibits LPS-induced production of IL 5, IL 10, and IL 13 from mast cells (181).
Finally, airway smooth muscle cells, like many other types of smooth muscle (182), also appear to be regulated by S1P. S1P contracts human airway smooth muscle cells by activating S1P2 or S1P3 receptors (183) and is involved in muscarinic M2 receptor–mediated bronchoconstriction (42). S1P-also causes bronchial hyperresponsiveness in mice, and ovalbumin-sensitized mouse lung is hyperresponsive to S1P (184, 185). Conversely, inhibition of SphK attenuates airway hyperresponsiveness in a murine asthma model (179). In addition, proliferation of airway smooth muscle, which is part of the asthmatic airway remodeling, is stimulated by S1P. S1P increases human airway smooth muscle cell proliferation and potentiates growth stimulation by thrombin and or epidermal growth factor (173, 186).
In the murine asthma model, FTY720 inhibited airway inflammation, bronchial hyperresponsiveness, and goblet cell hyperplasia (59). However, given the recent finding that FTY720 also inhibits cPLA2 (63), the relative role of S1P receptors in this process is now less clear than originally thought. Another study, based on the protective effects of fumonisin B1 in a guinea pig asthma model, also proposed, for the first time, a role for (increased) ceramide-mediated oxidative stress in the pathogenesis of asthma (187).
Because many of these pathways have been worked out in mice, it should be emphasized that human mast cells may behave differently (188, 189). For instance, whereas activation of FcεR1 receptors in mice activates predominantly SphK2 (177), in human mast cells, SphK1 may be at least equally important (190). This also highlights the need of specific inhibitors of the involved receptor and enzyme systems for future studies and treatments.
Ceramide-enriched membrane domains and rafts are critically involved in the infection of epithelial cells and the lung with several pathogens. One well-studied example is P. aeruginosa infection (112). P. aeruginosa induces, via unknown mechanisms, an activation of the A-SMase in epithelial cells and a translocation of intracellular localized A-SMase onto the extracellular leaflet of the cell membrane resulting in the formation of ceramide and ceramide-enriched membrane platforms in the outer leaflet (112). Genetic deficiency of the A-SMase or disruption of rafts by interference with cellular cholesterol not only prevented the formation of ceramide-enriched membrane platforms upon cellular infection with P. aeruginosa but also affected salient steps of the infection process such as P. aeruginosa internalization and induction of death in infected cells, whereas, at the same time, an uncontrolled cytokine release was observed.
Although the exact molecular mechanisms that are initiated in ceramide-enriched membrane domains during infection are unknown, it is possible that receptor clustering within these domains is critically involved in the effects induced by pulmonary P. aeruginosa infections. Thus, it was shown that CD95 receptor and cystic fibrosis transmembrane conductance regulator (CFTR) molecules cluster within ceramide-enriched membrane domains upon cellular infection with P. aeruginosa (112, 191). CD95 was previously shown to mediate apoptosis of cells infected with P. aeruginosa. Furthermore, clustering of the receptor in ceramide-enriched membrane platforms has been demonstrated to be required for the induction of cell death by CD95. Studies by G. Pier and colleagues (192, 193) demonstrated that CFTR functions as a receptor for P. aeruginosa, mediating internalization of the bacteria. P. aeruginosa lipopolysaccharide molecules bind to a short amino acid sequence in CFTR (AA 103–117) and deficiency of CFTR, or blockade of this binding motif, prevents P. aeruginosa internalization. Thus, the reorganization of cellular receptors such as CD95 and CFTR within P. aeruginosa–triggered ceramide-enriched membrane domains seems to be central for an adequate immune response of respiratory epithelial cells to P. aeruginosa. However, it is also possible that ceramide-enriched membrane domains have additional, yet undefined functions, during the infection of epithelial cells with P. aeruginosa. The mechanisms that result in an uncontrolled release of cytokines in the lung of Smpd1-deficient mice infected with P. aeruginosa are presently unknown. In vivo studies demonstrate that Smpd1-deficient mice, which are unable to form ceramide upon pulmonary infection with P. aeruginosa, are highly susceptible to an acute pulmonary P. aeruginosa infection, and a high percentage of the mice succumbed by a generalized sepsis within a few days after nasal infection with the bacteria, whereas wild-type mice were resistant (112).
It is unknown whether ceramide-enriched membrane domains mediate a negative effect on cytokine release or whether other effects of ceramide are required to balance cytokine release upon infection. The central role of membrane rafts for the response of epithelial cells to P. aeruginosa infections is also supported by studies on respiratory tract–derived epithelial cells (112, 194). These studies revealed the formation of large GM1-enriched membrane domains from small rafts, that colocalize with the previously described ceramide-enriched membrane domains (112). CFTR translocated into the raft fraction after infection, which was prevented by destruction of rafts by interference with the cholesterol metabolism (194). Most important, destruction of rafts prevented internalization of P. aeruginosa into epithelial cells, NF-κB nuclear translocation, and cellular apoptosis in vitro and in vivo in respiratory epithelial cells after pulmonary infection (194), supporting the notion that distinct membrane domains are critically involved in an adequate response of host cells to P. aeruginosa.
The notion that ceramide-enriched membrane domains are importantly involved in the infection with pathogens is consistent with previous findings on the infection of epithelial and endothelial cells with Neisseriae gonorrheae and Staphylococcus aureus (113, 114). These studies demonstrated that activation of the A-SMase and release of ceramide upon infection of epithelial cells with N. gonorrheae are required for internalization of the bacteria. Furthermore, expression of the A-SMase was a prerequisite for induction of death in endothelial cells infected with S. aureus.
S1P has also been implicated in infection. S1P is decreased in patients with pulmonary tuberculosis (195), possibly because mycobacteria inhibit macrophage SphK1 (196). This may be relevant for the disease because S1P stimulates the antimycobacterial activity of human macrophages and reduces mycobacterial growth, granuloma formation, and necrosis in the lungs of Mycobacterium tuberculosis–infected mice (197).
The role of rafts and ceramide-enriched membrane domains for pulmonary infections has been mainly investigated in the context of cystic fibrosis and P. aeruginosa infections. However, given the clinical significance of pneumonia, it will be important to define whether distinct membrane domains are also involved in pulmonary infections with other bacterial pathogens such as pneumococci or mycobacteria and even viral pathogens, for instance, influenza. Such studies may also help to explain the pulmonary infections that occur in approximately one-half of all patients with Niemann-Pick diseases, type B (133). Furthermore, it needs to be defined whether rafts can be targeted to prevent infections without altering normal lung physiology beyond a pathophysiological level.
Very elegant studies by I. Petrarche and colleages identified ceramide as a central mediator in the development of emphysema (198). These authors used blockade of vascular endothelial growth factor receptors as a model to investigate mechanisms of lung emphysema that typically are present in human chronic obstructive pulmonary disease (COPD) and emphysema caused by cigarette smoke. Vascular endothelial growth factor blockade resulted in the formation of ceramide predominantly in alveolar cells, which was mediated by an increased activity of the ceramide synthase pathway and a feed-forward activation of the A-SMase (198). A similar mechanism with two waves of ceramide has also been suggested for radiation-induced ceramide production (199). Increased ceramide was critical to trigger alveolar cell death, oxidative stress (200), and emphysema, events that were prevented by the inhibition of the ceramide synthase pathway in vivo using fumonisin B1 or myriocin. Intrapulmonary application of ceramide mimicked the effects of endogenous ceramide and also resulted in activation of caspase 3 in the lung, alveolar cell death, and emphysema. Most interestingly, cigarette smoke induced the production of ceramide in endothelial cells as well as the release of the proinflammatory mediator, TNF. Emphysema patients had increased ceramide levels that were localized to alveolar septal cells and macrophages. The authors concluded that pulmonary emphysema develops as a consequence of ceramide-induced oxidative stress and apoptosis of alveolar endothelial and/or epithelial cells with subsequent digestion of interstitial tissue by matrix metalloproteases (198, 200). In some sense, this mechanism is opposed to the classical view that one function of apoptosis is to diminish inflammation. In the emphysema model it would be the massive apoptosis stimulating an inflammatory response that finally leads to emphysema. Interestingly, also in this case, S1P antagonizes ceramide (198).
Additional studies extended these observations to the effects of ceramide on endothelial cells in the lung. A recent study by Castillo and colleagues (24) also implied the neutral N-SMase to be involved in airway epithelial cell death. This study indicated that reactive oxygen species, in particular H2O2, activate the N-SMase2, whereas peroxynitrate stimulated the A-SMase (24). Collectively, these studies strongly suggest endothelial ceramide as a novel target to prevent or treat emphysema and COPD. It will be interesting to determine whether sphingolipids also play a role in other cells relevant for the development of COPD, in particular, in macrophages and natural killer T cells thought to be critical for the development of COPD (201).
Children with cystic fibrosis (CF) very often develop infections with P. aeruginosa and once past childhood almost all patients with cystic fibrosis suffer from a chronic pneumonia with P. aeruginosa, Burkholderia cepacia, and/or S. aureus. Although the life expectancy of patients with CF has increased, these bacterial lung infections are key to the development of the disease and very often result in destruction of the lung. CF is caused by a mutation of CFTR and occurs with a frequency of 1:2,500 births, at least in Western countries. Several recent studies suggested a proinflammatory status in the lung, and possibly also other organs, of patients with CF that triggers chronic inflammation even without a bacterial or viral infection. Thus, it was shown that even noninfected Cftr-deficient mice suffer from increased IL-8 concentrations in the trachea (202, 203). Further, studies on aborted embryos with CF and on BAL fluids from patients with CF as young as 4 weeks with negative cultures for CF-related bacteria, virus, and fungi, revealed a significant increase of proinflammatory mediators in the lungs (204, 205). These studies suggest that patients with CF suffer from an uncontrolled inflammation in the lung that might be critical for the propensity of these patients to develop infections with P. aeruginosa and other bacteria.
Recent studies imply sphingolipids (206), and in particular ceramide, as critical regulators for the development of the high sensitivity of Cftr-deficient mice to P. aeruginosa infections (207). These studies demonstrated in different Cftr-deficient mouse strains that ceramide accumulation in respiratory epithelial cells and in the submucosal glands of uninfected Cftr-deficient mice is age dependent (207). Cftr deficiency alters the acidification of intracellular vesicles, most likely prelysosomes and lysosomes, in respiratory epithelial cells, which results in alkalinization of these vesicles. It should be noted that other vesicles, in particular endosomes, with a different pattern of ion channel expression than prelysosomes and lysosomes are not alkalinized in CF (208–210). The inappropriate alkalinization of (pre)lysosomes and other acidic vesicles may result in an imbalance of enzymes that release (acid sphingomyelinase) and consume ceramide (acid ceramidase) and an accumulation of ceramide in respiratory epithelial cells from Cftr-deficient mice. A similar accumulation was observed in respiratory epithelial cells and lung specimens from patients with CF, suggesting that the mouse data also apply to humans (207). The increased ceramide concentration in airway epithelial cells of Cftr-deficient mice triggered chronic pulmonary inflammation, death of respiratory epithelial cells, and deposition of DNA in bronchi and thus explains the proneness of Cftr-deficient mice to develop pulmonary P. aeruginosa infections. Extracellular DNA on epithelial cells may facilitate bacterial adherence in the respiratory tract, whereas an increased and chronic inflammatory response may affect lung functions, block the mucociliary clearance, alter the innate immune response, change the airway architecture, and finally result in pulmonary fibrosis. Partial inhibition of A-SMase that was achieved by heterozygosis of the A-SMase in Cftr-deficient mice (Cftr−/−/Smpd1+/− mice) or by pharmacological treatment of Cftr-deficient mice with the functional A-SMase inhibitor amitriptyline, normalized pulmonary ceramide, abrogated the above-mentioned consequences and, most important, prevented severe pulmonary infections with P. aeruginosa.
Although these studies indicate that ceramide also accumulates in CFTR-deficient epithelial cells from patients with CF, it remains to be determined whether the data obtained in a murine system can be transferred to the human situation. Furthermore, the role of sphingolipids in other cells that normally express Cftr—such as macrophages, endothelial cells, and neutrophils—for the development of inflammation and infection susceptibility in CF remains to be determined. Finally, one of the major problems of children with CF is the development of fibrosis along with the inflammation and infection. Fibrosis might be a consequence of chronic infection, but animal models suggest that fibrosis also occurs in Cftr-deficient mice without infection and may therefore be an independent part of the disease that needs further attention.
These results indicate a novel concept for explaining the pathogenesis of lung disease in CF, i.e., an increase of ceramide caused by an imbalance of enzyme activities upon intravesicular pH changes. On the other hand, the role of sphingolipids in infection, as described above (3.4. Sphingolipids in Pulmonary Infections), maintains that a normal defense against P. aeruginosa requires intact membrane rafts, at least some activity of the A-SMase, and normal ceramide values. Thus, any future treatment of CF with A-SMase blockers must likely be personalized and tightly controlled to prevent too strong an inhibition of the acid sphingomyelinase. We speculate that inhibition of the A-SMase to treat of CF might be facilitated by the development of inhalation drugs that block the A-SMase and, thus, will improve CF.
Several studies indicate that rafts are not only critically involved in the infection of respiratory cells with pathogenic bacteria, but also with viruses that affect the respiratory tract. Thus, the infection of human epithelial cells with pathogenic rhinoviruses that cause the common cold results in the formation of large membrane platforms derived from small rafts (115, 211). Destruction of these domains by extraction of cholesterol prevented internalization of the virus indicating the significance of rafts for the infection (115, 211). Further studies revealed a rapid activation of the A-SMase, release of ceramide, and formation of large ceramide-enriched membrane platforms upon infection with human rhinoviruses. Pharmacological or small interfering RNA-mediated inhibition of the A-SMase prevented infection of human epithelial cells with rhinovirus, supporting the notion that raft-derived domains and ceramide-enriched domains are important for infection of human epithelial cells with rhinoviruses.
Sphingolipids, in particular sphingomyelin, are present in pulmonary surfactant (212) and are, together with some phospholipids, increased in A-SMase–deficient mice (213). Of note, the BAL of patients with ARDS shows an elevated sphingomyelin:phosphatidycholine ratio (214). The relevance of this observation is not known, but interestingly, instillation of TNF leads to increased alveolar sphingomyelin levels and sphingomyelinase activity that may, in turn, raise ceramide concentrations that finally impair surfactant functions (139) and possibly also production (215).
Because so many cellular activities are affected by sphingolipids, it comes as no surprise that we learn about more lung diseases in which sphingolipds play a prominent role. And it seems likely that, similar to the situation of arachidonic acid metabolites 30 years ago, more sphingolipid entities with important biological functions will be discovered such as ceramide-1-phosphate (216), sphingosylphosphorylcholine (217), or dihydrosphingosine-1-phosphate (218). In mouse models, inhibition of ceramide provided a novel treatment for several disorders, and it is now of great importance to transfer these insights into the development of novel therapeutic opportunities for humans, some of which have been addressed here. Furthermore, specific inhibitors of the A-SMase/ceramide pathway or modifiers of S1P receptors will be critical for transferring our basic knowledge of lung diseases onto future treatments.
However, the multitude of physiological roles assumed by sphingolipids also raises the issue of side effects. The development of successful drugs will therefore depend on an improved understanding of the roles of sphingolipids at the cellular, pulmonary, and systemic levels. At the cellular level, one fundamental question concerns the exact role of cellular ceramide and the conditions when it acts as a second messenger, when its mode of action consists mainly in altering the biophysical membrane properties, or when it modulates ion channels. These insights will help to further develop approaches directed at the inhibition of sphingomyelinase or ceramide synthase isoenzymes. At the pulmonary levels, comprehensive information on the distribution and function of S1P receptors in the lung, and further studies on the role of sphingolipids in the regulation of immune processes, will be required to define therapeutic opportunities and to avoid severe pulmonary side effects. And finally, progress in understanding sphingolipid biology in other organs will raise the awareness of possible extrapulmonary side effects.
However, several deliberations favor the chance that it may be possible to develop relatively safe sphingolipid-targeted drugs. First, ceramide, which seems to be predominantly involved in stress responses, might prove an excellent target for novel treatment strategies of diseases such as sepsis, bacterial infections, ARDS, COPD, and CF. Its safety is suggested by the fact that mice lacking the A-SMase do not suffer from acute disorders (though they develop a chronic storage disease) and that many patients have already been treated with A-SMase inhibitors in the form of tricyclic antidepressants such as imipramine. Second, incomplete inhibition of the A-SMase may be sufficient for the treatment of chronic diseases such as CF and COPD, thus further reducing the likelihood of side effects. Third, it might be possible to take advantage of the fact that several enzymes, such as acid sphingomyelinase (A-SMase) (see Table 1), acid ceramidase (219), neutral ceramidase (220), and SphK1 (39) are found in the extracellular space, allowing for the design of drugs that need not enter cells and might thus have fewer side effects. Fourth, the considerable number of S1P receptors suggests that it may be possible to develop receptor-specific drugs with reduced side effects. And finally, side effects can be minimized by local treatment of the lungs using inhalation drugs.
We suggest that the field of sphingolipids offers many promising and relatively safe avenues to the treatment of pulmonary disorders.
|1.||Hannun YA, Luberto C, Argraves KM. Enzymes of sphingolipid metabolism: from modular to integrative signaling. Biochemistry 2001;40:4893–4903.|
|2.||Futerman AH, Hannun YA. The complex life of simple sphingolipids. EMBO Rep 2004;5:777–782.|
|3.||Pyne S, Pyne NJ. Sphingosine 1 phosphate signalling in mammalian cells. Biochem J 2000;349:385–402.|
|4.||Spiegel S, Milstien S. Sphingosine 1 phosphate: an enigmatic signalling lipid. Nat Rev Mol Cell Biol 2003;4:397–407.|
|5.||Bielawska AE, Shapiro JP, Jiang L, Melkonyan HS, Piot C, Wolfe CL, Tomei LD, Hannun YA, Umansky SR. Ceramide is involved in triggering of cardiomyocyte apoptosis induced by ischemia and reperfusion. Am J Pathol 1997;151:1257–1263.|
|6.||Lang PA, Kempe DS, Tanneur V, Eisele K, Klarl BA, Myssina S, Jendrossek V, Ishii S, Shimizu T, Waidmann M, et al. Stimulation of erythrocyte ceramide formation by platelet activating factor. J Cell Sci 2005;118:1233–1243.|
|7.||Göggel R, Winoto Morbach S, Vielhaber G, Imai Y, Lindner K, Brade L, Brade H, Ehlers S, Slutsky AS, Schütze S, et al. PAF-mediated pulmonary edema: a new role for acid sphingomyelinase and ceramide. Nat Med 2004;10:155–160.|
|8.||Mathias S, Younes A, Kan CC, Orlow I, Joseph C, Kolesnick RN. Activation of the sphingomyelin signaling pathway in intact EL4 cells and in a cell free system by IL 1 beta. Science 1993;259:519–522.|
|9.||Schütze S, Potthoff K, Machleidt T, Berkovic D, Wiegmann K, Krönke M. TNF activates NF κB by phosphatidylcholine specific phospholipase C induced “acidic” sphingomyelin breakdown. Cell 1992;71:765–776.|
|10.||Auge N, Andrieu N, Negre Salvayre A, Thiers JC, Levade T, Salvayre R. The sphingomyelin ceramide signaling pathway is involved in oxidized low density lipoprotein induced cell proliferation. J Biol Chem 1996;271:19251–19255.|
|11.||Grammatikos G, Teichgraber V, Carpinteiro A, Trarbach T, Weller M, Hengge UR, Gulbins E. Overexpression of acid sphingomyelinase sensitizes glioma cells to chemotherapy. Antioxid Redox Signal 2007;9:1449–1456.|
|12.||Lacour S, Hammann A, Grazide S, Lagadic Gossmann D, Athias A, Sergent O, Laurent G, Gambert P, Solary E. manche Boitrel MT. Cisplatin induced CD95 redistribution into membrane lipid rafts of HT29 human colon cancer cells. Cancer Res 2004;64:3593–3598.|
|13.||Bezombes C, Grazide S, Garret C, Fabre C, Quillet Mary A, Muller S, Jaffrezou JP, Laurent G. Rituximab antiproliferative effect in B lymphoma cells is associated with acid sphingomyelinase activation in raft microdomains. Blood 2004;104:1166–1173.|
|14.||Zhang DX, Zou AP, Li PL. Ceramide reduces endothelium dependent vasodilation by increasing superoxide production in small bovine coronary arteries. Circ Res 2001;88:824–831.|
|15.||Santana P, Pena LA, Haimovitz Friedman A, Martin S, Green D, McLoughlin M, Cordon Cardo C, Schuchman EH, Fuks Z, Kolesnick R. Acid sphingomyelinase deficient human lymphoblasts and mice are defective in radiation induced apoptosis. Cell 1996;86:189–199.|
|16.||Hanada K, Kumagai K, Tomishige N, Kawano M. CERT and intracellular trafficking of ceramide. Biochim Biophys Acta 2007;1771:644–653.|
|17.||Okino N, He X, Gatt S, Sandhoff K, Ito M, Schuchman EH. The reverse activity of human acid ceramidase. J Biol Chem 2003;278:29948–29953.|
|18.||Marchesini N, Hannun YA. Acid and neutral sphingomyelinases: roles and mechanisms of regulation. Biochem Cell Biol 2004;82:27–44.|
|19.||Schissel SL, Keesler GA, Schuchman EH, Williams KJ, Tabas I. The cellular trafficking and zinc dependence of secretory and lysosomal sphingomyelinase, two products of the acid sphingomyelinase gene. J Biol Chem 1998;273:18250–18259.|
|20.||Koval M, Pagano RE. Intracellular transport and metabolism of sphingomyelin. Biochim Biophys Acta 1991;1082:113–125.|
|21.||Marathe S, Schisse DL, Yellin MJ, Beatini N, Mintzer R, Williams KJ, Tabas I. Human vascular endothelial cells are a rich and regulatable source of secretory sphingomyelinase. Implications for early atherogenesis and ceramide mediated cell signaling. J Biol Chem 1998;273:4081–4088.|
|22.||Takahashi I, Takahashi T, Abe T, Watanabe W, Takada G. Distribution of acid sphingomyelinase in human various body fluids. Tohoku J Exp Med 2000;192:61–66.|
|23.||Goldkorn T, Ravid T, Khan EM. Life and death decisions: ceramide generation and EGF receptor trafficking are modulated by oxidative stress. Antioxid Redox Signal 2005;7:119–128.|
|24.||Castillo SS, Levy M, Thaikoottathil JV, Goldkorn T. Reactive nitrogen and oxygen species activate different sphingomyelinases to induce apoptosis in airway epithelial cells. Exp Cell Res 2007;313:2680–2686.|
|25.||Krut O, Wiegmann K, Kashkar H, Yazdanpanah B, Krönke M. Novel tumor necrosis factor responsive mammalian neutral sphingomyelinase 3 is a C tail anchored protein. J Biol Chem 2006;281:13784–13793.|
|26.||Chalfant CE, Spiegel S. Sphingosine 1 phosphate and ceramide 1 phosphate: expanding roles in cell signaling. J Cell Sci 2005;118:4605–4612.|
|27.||Mitra P, Maceyka M, Payne SG, Lamour N, Milstien S, Chalfant CE, Spiegel S. Ceramide kinase regulates growth and survival of A549 human lung adenocarcinoma cells. FEBS Lett JT . FEBS Lett 2007;581:735–740.|
|28.||Graf C, Zemann B, Rovina P, Urtz N, Schanzer A, Reuschel R, Mechtcheriakova D, Muller M, Fischer E, Reichel C, et al. Neutropenia with impaired immune response to streptococcus pneumoniae in ceramide kinase deficient mice. J Immunol 2008;180:3457–3466.|
|29.||Alemany R, van Koppen CJ, Danneberg K, Ter BM, Meyer Zu HD. Regulation and functional roles of sphingosine kinases. Naunyn Schmiedebergs Arch Pharmacol 2007;374:413–428.|
|30.||Watterson K, Sankala H, Milstien S, Spiegel S. Pleiotropic actions of sphingosine 1 phosphate. Prog Lipid Res 2003;42:344–357.|
|31.||Spiegel S, English D, Milstien S. Sphingosine 1 phosphate signaling: providing cells with a sense of direction. Trends Cell Biol 2002;12:236–242.|
|32.||Yang AH, Ishii I, Chun J. In vivo roles of lysophospholipid receptors revealed by gene targeting studies in mice. Biochim Biophys Acta 2002;1582:197–203.|
|33.||Murakami M, Shiraishi A, Tabata K, Fujita N. Identification of the orphan GPCR, P2Y(10) receptor as the sphingosine 1 phosphate and lysophosphatidic acid receptor. Biochem Biophys Res Commun 2008;371:707–712.|
|34.||Allende ML, Proia RL. Sphingosine 1 phosphate receptors and the development of the vascular system. Biochim Biophys Acta 2002;1582:222–227.|
|35.||Murata N, Sato K, Kon J, Tomura H, Yanagita M, Kuwabara A, Ui M, Okajima F. Interaction of sphingosine 1 phosphate with plasma components, including lipoproteins, regulates the lipid receptor mediated actions. Biochem J 2000;352:809–815.|
|36.||Olivera A, Kohama T, Edsall L, Nava V, Cuvillier O, Poulton S, Spiegel S. Sphingosine kinase expression increases intracellular sphingosine 1 phosphate and promotes cell growth and survival. J Cell Biol 1999;147:545–558.|
|37.||Yatomi Y, Ozaki Y, Ohmori T, Igarashi Y. Sphingosine 1 phosphate: synthesis and release. Prostaglandins Other Lipid Mediat 2001;64:107–122.|
|38.||Pappu R, Schwab SR, Cornelissen I, Pereira JP, Regard JB, Xu Y, Camerer E, Zheng YW, Huang Y, Cyster JG, et al. Promotion of lymphocyte egress into blood and lymph by distinct sources of sphingosine 1 phosphate. Science 2007;316:295–298.|
|39.||Venkataraman K, Thangada S, Michaud J, Oo ML, Ai Y, Lee YM, Wu M, Parikh NS, Khan F, Proia RL, et al. Extracellular export of sphingosine kinase 1a contributes to the vascular S1P gradient. Biochem J 2006;397:461–471.|
|40.||Zhao Y, Kalari SK, Usatyuk PV, Gorshkova I, He D, Watkins T, Brindley DN, Sun C, Bittman R, Garcia JG, et al. Intracellular generation of sphingosine 1 phosphate in human lung endothelial cells: role of lipid phosphate phosphatase 1 and sphingosine kinase 1. J Biol Chem 2007;282:14165–14177.|
|41.||Taha TA, Hannun YA, Obeid LM. Sphingosine kinase: biochemical and cellular regulation and role in disease. J Biochem Mol Biol 2006;39:113–131.|
|42.||Pfaff M, Powaga N, Akinci S, Schutz W, Banno Y, Wiegand S, Kummer W, Wess J, Haberberger RV. Activation of the SPHK/S1P signalling pathway is coupled to muscarinic receptor dependent regulation of peripheral airways. Respir Res 2005;6:48.|
|43.||Choi OH, Kim JH, Kinet JP. Calcium mobilization via sphingosine kinase in signalling by the Fc epsilon RI antigen receptor. Nature 1996;380:634–636.|
|44.||Oskeritzian CA, Alvarez SE, Hait NC, Price MM, Milstien S, Spiegel S. Distinct roles of sphingosine kinases 1 and 2 in human mast cell functions. Blood 2008;111:4193–4200.|
|45.||Cowart LA, Szulc Z, Bielawska A, Hannun YA. Structural determinants of sphingolipid recognition by commercially available anti ceramide antibodies. J Lipid Res 2002;43:2042–2048.|
|46.||Goni FM, Contreras FX, Montes LR, Sot J, Alonso A. Biophysics (and sociology) of ceramides. Biochem Soc Symp 2005;177–188.|
|47.||Kroesen BJ, Jacobs S, Pettus BJ, Sietsma H, Kok JW, Hannun YA, de Leij LF. BcR induced apoptosis involves differential regulation of C16 and C24 ceramide formation and sphingolipid dependent activation of the proteasome. J Biol Chem 2003;278:14723–14731.|
|48.||Desai K, Sullards MC, Allegood J, Wang E, Schmelz EM, Hartl M, Humpf HU, Liotta DC, Peng Q, Merrill AH Jr. Fumonisins and fumonisin analogs as inhibitors of ceramide synthase and inducers of apoptosis. Biochim Biophys Acta 2002;1585:188–192.|
|49.||Hurwitz R, Ferlinz K, Sandhoff K. The tricyclic antidepressant desipramine causes proteolytic degradation of lysosomal sphingomyelinase in human fibroblasts. Biol Chem Hoppe Seyler 1995;375:447–450.|
|50.||Zeidan YH, Pettus BJ, Elojeimy S, Taha T, Obeid LM, Kawamori T, Norris JS, Hannun YA. Acid ceramidase but not acid sphingomyelinase is required for tumor necrosis factor-α–induced PGE2 production. J Biol Chem 2006;281:24695–24703.|
|51.||Luberto C, Hannun YA. Sphingomyelin synthase, a potential regulator of intracellular levels of ceramide and diacylglycerol during SV40 transformation. Does sphingomyelin synthase account for the putative phosphatidylcholine specific phospholipase C? J Biol Chem 1998;273:14550–14559.|
|52.||Hannun YA, Luberto C. Ceramide in the eukaryotic stress response. Trends Cell Biol 2000;10:73–80.|
|53.||Zemann B, Urtz N, Reuschel R, Mechtcheriakova D, Bornancin F, Badegruber R, Baumruker T, Billich A. Normal neutrophil functions in sphingosine kinase type 1 and 2 knockout mice. Immunol Lett 2007;109:56–63.|
|54.||French KJ, Schrecengost RS, Lee BD, Zhuang Y, Smith SN, Eberly JL, Yun JK, Smith CD. Discovery and evaluation of inhibitors of human sphingosine kinase. Cancer Res 2003;63:5962–5969.|
|55.||Lee C, Xu DZ, Feketeova E, Kannan KB, Yun JK, Deitch EA, Fekete Z, Livingston DH, Hauser CJ. Attenuation of shock induced acute lung injury by sphingosine kinase inhibition. J Trauma 2004;57:955–960.|
|56.||Park JH, Schuchman EH. Acid ceramidase and human disease. Biochim Biophys Acta 2006; 1758:2133–2138.|
|57.||Mandala S, Hajdu R, Bergstrom J, Quackenbush E, Xie J, Milligan J, Thornton R, Shei GJ, Card D, Keohane C, et al. Alteration of lymphocyte trafficking by sphingosine 1 phosphate receptor agonists. Science 2002;296:346–349.|
|58.||Peng X, Hassoun PM, Sammani S, McVerry BJ, Burne MJ, Rabb H, Pearse D, Tuder RM, Garcia JG. Protective effects of sphingosine 1 phosphate in murine endotoxin induced inflammatory lung injury. Am J Respir Crit Care Med 2004;169:1245–1251.|
|59.||Sawicka E, Zuany Amorim C, Manlius C, Trifilieff A, Brinkmann V, Kemeny DM, Walker C. Inhibition of Th1 and Th2 mediated airway inflammation by the sphingosine 1 phosphate receptor agonist FTY720. J Immunol 2003;171:6206–6214.|
|60.||Idzko M, Hammad H, van Nimwegen M, Kool M, Möller T, Soullié T, Willart MA, Hijdra D, Hoogsteden HC, Lambrecht BN. Local application of FTY720 to the lung abrogates experimental asthma by altering dendritic cell function. J Clin Invest 2006;116:2935–2944.|
|61.||Dudek SM, Camp SM, Chiang ET, Singleton PA, Usatyuk PV, Zhao Y, Natarajan V, Garcia JG. Pulmonary endothelial cell barrier enhancement by FTY720 does not require the S1P1 receptor. Cell Signal 2007;19:1754–1764.|
|62.||Paugh SW, Cassidy MP, He H, Milstien S, Sim Selley LJ, Spiegel S, Selley DE. Sphingosine and its analog, the immunosuppressant 2 amino 2 (2 [4 octylphenyl]ethyl) 1,3 propanediol, interact with the CB1 cannabinoid receptor. Mol Pharmacol 2006;70:41–50.|
|63.||Payne SG, Oskeritzian CA, Griffiths R, Subramanian P, Barbour SE, Chalfant CE, Milstien S, Spiegel S. The immunosuppressant drug FTY720 inhibits cytosolic phospholipase A2 independently of sphingosine 1 phosphate receptors. Blood 2007;109:1077–1085.|
|64.||Sanna MG, Liao J, Jo E, Alfonso C, Ahn MY, Peterson MS, Webb B, Lefebvre S, Chun J, Gray N, et al. Sphingosine 1 phosphate (S1P) receptor subtypes S1P1 and S1P3, respectively, regulate lymphocyte recirculation and heart rate. J Biol Chem 2004;279:13839–13848.|
|65.||Sanna MG, Wang SK, Gonzalez Cabrera PJ, Don A, Marsolais D, Matheu MP, Wei SH, Parker I, Jo E, Cheng WC, et al. Enhancement of capillary leakage and restoration of lymphocyte egress by a chiral S1P1 antagonist in vivo. Nat Chem Biol 2006;2:434–441.|
|66.||Osada M, Yatomi Y, Ohmori T, Ikeda H, Ozaki Y. Enhancement of sphingosine 1 phosphate induced migration of vascular endothelial cells and smooth muscle cells by an EDG 5 antagonist. Biochem Biophys Res Commun 2002;299:483–487.|
|67.||Horinouchi K, Erlich S, Perl DP, Ferlinz K, Bisgaier CL, Sandhoff K, Desnick RJ, Stewart CL, Schuchman EH. Acid sphingomyelinase deficient mice: a model of types A and B Niemann-Pick disease. Nat Genet 1995;10:288–293.|
|68.||Kuemmel TA, Thiele J, Schroeder R, Stoffel W. Pathology of visceral organs and bone marrow in an acid sphingomyelinase deficient knock out mouse line, mimicking human Niemann-Pick disease type A. A light and electron microscopic study. Pathol Res Pract 1997;193:663–671.|
|69.||Dhami R, Passini MA, Schuchman EH. Identification of novel biomarkers for Niemann Pick disease using gene expression analysis of acid sphingomyelinase knockout mice. Mol Ther 2006;13:556–564.|
|70.||Clarke CJ, Hannun YA. Neutral sphingomyelinases and nSMase2: bridging the gaps. Biochim Biophys Acta 2006;1758:1893–1901.|
|71.||Stoffel W, Jenke B, Block B, Zumbansen M, Koebke J. Neutral sphingomyelinase 2 (smpd3) in the control of postnatal growth and development. Proc Natl Acad Sci USA 2005;102:4554–4559.|
|72.||Liu Y, Wada R, Yamashita T, Mi Y, Deng CX, Hobson JP, Rosenfeldt HM, Nava VE, Chae SS, Lee MJ, et al. Edg 1, the G protein coupled receptor for sphingosine 1 phosphate, is essential for vascular maturation. J Clin Invest 2000;106:951–961.|
|73.||Ishii I, Friedman B, Ye X, Kawamura S, McGiffert C, Contos JJ, Kingsbury MA, Zhang G, Brown JH, Chun J. Selective loss of sphingosine 1 phosphate signaling with no obvious phenotypic abnormality in mice lacking its G protein coupled receptor, LP(B3)/EDG 3. J Biol Chem 2001;276:33697–33704.|
|74.||Ishii I, Ye X, Friedman B, Kawamura S, Contos JJ, Kingsbury MA, Yang AH, Zhang G, Brown JH, Chun J. Marked perinatal lethality and cellular signaling deficits in mice null for the two sphingosine 1 phosphate (S1P) receptors, S1P(2)/LP(B2)/EDG 5 and S1P(3)/LP(B3)/EDG 3. J Biol Chem 2002;277:25152–25159.|
|75.||MacLennan AJ, Carney PR, Zhu WJ, Chaves AH, Garcia J, Grimes JR, Anderson KJ, Roper SN, Lee N. An essential role for the H218/AGR16/Edg 5/LP(B2) sphingosine 1 phosphate receptor in neuronal excitability. Eur J Neurosci 2001;14:203–209.|
|76.||Kono M, Belyantseva IA, Skoura A, Frolenkov GI, Starost MF, Dreier JL, Lidington D, Bolz SS, Friedman TB, Hla T, et al. Deafness and stria vascularis defects in S1P2 receptor null mice. J Biol Chem 2007;282:10690–10696.|
|77.||Allende ML, Sasaki T, Kawai H, Olivera A, Mi Y, van Echten Deckert G, Hajdu R, Rosenbach M, Keohane CA, Mandala S, et al. Mice deficient in sphingosine kinase 1 are rendered lymphopenic by FTY720. J Biol Chem 2004;279:52487–52492.|
|78.||Kharel Y, Lee S, Snyder AH, Sheasley O'Neill SL, Morris MA, Setiady Y, Zhu R, Zigler MA, Burcin TL, Ley K, et al. Sphingosine kinase 2 is required for modulation of lymphocyte traffic by FTY720. J Biol Chem 2005;280:36865–36872.|
|79.||Mizugishi K, Yamashita T, Olivera A, Miller GF, Spiegel S, Proia RL. Essential role for sphingosine kinases in neural and vascular development. Mol Cell Biol 2005;25:11113–11121.|
|80.||Eliyahu E, Park JH, Shtraizent N, He X, Schuchman EH. Acid ceramidase is a novel factor required for early embryo survival. FASEB J 2007;21:1403–1409.|
|81.||Hannun YA, Obeid LM. The Ceramide centric universe of lipid mediated cell regulation: stress encounters of the lipid kind. J Biol Chem 2002;277:25847–25850.|
|82.||Taha TA, Mullen TD, Obeid LM. A house divided: ceramide, sphingosine, and sphingosine 1 phosphate in programmed cell death. Biochim Biophys Acta 2006;1758:2027–2036.|
|83.||Medler TR, Petrusca DN, Lee PJ, Hubbard WC, Berdyshev EV, Skirball J, Kamocki K, Schuchman E, Tuder RM, Petrache I. Apoptotic sphingolipid signaling by ceramides in lung endothelial cells. Am J Respir Cell Mol Biol 2008;38:639–646.|
|84.||Xia P, Wang L, Gamble JR, Vadas MA. Activation of sphingosine kinase by tumor necrosis factor alpha inhibits apoptosis in human endothelial cells. J Biol Chem 1999;274:34499–34505.|
|85.||Monick MM, Mallampalli RK, Bradford M, McCoy D, Gross TJ, Flaherty DM, Powers LS, Cameron K, Kelly S, Merrill AH Jr, et al. Cooperative prosurvival activity by ERK and Akt in human alveolar macrophages is dependent on high levels of acid ceramidase activity. J Immunol 2004;173:123–135.|
|86.||Castillo SS, Levy M, Wang C, Thaikoottathil JV, Khan E, Goldkorn T. Nitric oxide enhanced caspase 3 and acidic sphingomyelinase interaction: a novel mechanism by which airway epithelial cells escape ceramide induced apoptosis. Exp Cell Res 2007;313:816–823.|
|87.||Gulbins E, Li PL. Physiological and pathophysiological aspects of ceramide. Am J Physiol Regul Integr Comp Physiol 2006;290:R11–R26.|
|88.||Thon L, Mohlig H, Mathieu S, Lange A, Bulanova E, Winoto Morbach S, Schutze S, Bulfone Paus S, Adam D. Ceramide mediates caspase independent programmed cell death. FASEB J 2005;19:1945–1956.|
|89.||Hughes SE. Detection of apoptosis using in situ markers for DNA strand breaks in the failing human heart. fact or epiphenomenon? J Pathol 2003;201:181–186.|
|90.||Maceyka M, Sankala H, Hait NC, Le SH, Liu H, Toman R, Collier C, Zhang M, Satin LS, Merrill AH Jr, et al. SphK1 and SphK2, sphingosine kinase isoenzymes with opposing functions in sphingolipid metabolism. J Biol Chem 2005;280:37118–37129.|
|91.||Min J, Van VP, Zhang L, Hanigan MH, Alexander H, Alexander S. Sphingosine 1 phosphate lyase regulates sensitivity of human cells to select chemotherapy drugs in a p38 dependent manner. Mol Cancer Res 2005;3:287–296.|
|92.||Hait NC, Oskeritzian CA, Paugh SW, Milstien S, Spiegel S. Sphingosine kinases, sphingosine 1 phosphate, apoptosis and diseases. Biochim Biophys Acta 2006;1758:2016–2026.|
|93.||Johnson KR, Johnson KY, Crellin HG, Ogretmen B, Boylan AM, Harley RA, Obeid LM. Immunohistochemical distribution of sphingosine kinase 1 in normal and tumor lung tissue. J Histochem Cytochem 2005;53:1159–1166.|
|94.||Kono Y, Nishiuma T, Nishimura Y, Kotani Y, Okada T, Nakamura S, Yokoyama M. Sphingosine kinase 1 regulates differentiation of human and mouse lung fibroblasts mediated by TGF beta1. Am J Respir Cell Mol Biol 2007;37:395–404.|
|95.||Singer SJ, Nicolson GL. The fluid mosaic model of the structure of cell membranes. Science 1972;175:720–731.|
|96.||Brown DA, London E. Functions of lipid rafts in biological membranes. Annu Rev Cell Dev Biol 1998;14:111–136.|
|97.||Kolesnick RN, Goni FM, Alonso A. Compartmentalization of ceramide signaling: physical foundations and biological effects. J Cell Physiol 2000;184:285–300.|
|98.||Simons K, Ikonen E. Functional rafts in cell membranes. Nature 1997;387:569–572.|
|99.||Xu X, Bittman R, Duportail G, Heissler D, Vilcheze C, London E. Effect of the structure of natural sterols and sphingolipids on the formation of ordered sphingolipid/sterol domains (rafts). Comparison of cholesterol to plant, fungal, and disease associated sterols and comparison of sphingomyelin, cerebrosides, and ceramide. J Biol Chem 2001;276:33540–33546.|
|100.||Liu P, Anderson RGW. Compartmentalized production of ceramide at the cell surface. J Biol Chem 1995;45:27129–27185.|
|101.||Gulbins E, Kolesnick R. Raft ceramide in molecular medicine. Oncogene 2003;22:7070–7077.|
|102.||Gulbins E, Dreschers S, Wilker B, Grassme H. Ceramide, membrane rafts and infections. J Mol Med 2004;82:357–363.|
|103.||Parton RG, Simons K. The multiple faces of caveolae. Nat Rev Mol Cell Biol 2007;8:185–194.|
|104.||Predescu SA, Predescu DN, Malik AB. Molecular determinants of endothelial transcytosis and their role in endothelial permeability. Am J Physiol Lung Cell Mol Physiol 2007;293:L823–L842.|
|105.||Megha LE. Ceramide selectively displaces cholesterol from ordered lipid domains (rafts): implications for lipid raft structure and function. J Biol Chem 2004;279:9997–10004.|
|106.||Nurminen TA, Holopainen JM, Zhao H, Kinnunen PK. Observation of topical catalysis by sphingomyelinase coupled to microspheres. J Am Chem Soc 2002;124:12129–12134.|
|107.||Grassme H, Jekle A, Riehle A, Schwarz H, Berger J, Sandhoff K, Kolesnick R, Gulbins E. CD95 signaling via ceramide rich membrane rafts. J Biol Chem 2001;276:20589–20596.|
|108.||Grassme H, Jendrossek V, Bock J, Riehle A, Gulbins E. Ceramide rich membrane rafts mediate CD40 clustering. J Immunol 2002;168:298–307.|
|109.||Dumitru CA, Gulbins E. TRAIL activates acid sphingomyelinase via a redox mechanism and releases ceramide to trigger apoptosis. Oncogene 2006;25:5612–5625.|
|110.||Abdel Shakor AB, Kwiatkowska K, Sobota A. Cell surface ceramide generation precedes and controls FcgammaRII clustering and phosphorylation in rafts. J Biol Chem 2004;279:36778–36787.|
|111.||Pfeiffer A, Bottcher A, Orso E, Kapinsky M, Nagy P, Bodnar A, Spreitzer I, Liebisch G, Drobnik W, Gempel K, et al. Lipopolysaccharide and ceramide docking to CD14 provokes ligand specific receptor clustering in rafts. Eur J Immunol 2001;31:3153–3164.|
|112.||Grassme H, Jendrossek V, Riehle A. von KG, Berger J, Schwarz H, Weller M, Kolesnick R, Gulbins E. Host defense against Pseudomonas aeruginosa requires ceramide rich membrane rafts. Nat Med 2003;9:322–330.|
|113.||Esen M, Schreiner B, Jendrossek V, Lang F, Fassbender K, Grassme H, Gulbins E. Mechanisms of Staphylococcus aureus induced apoptosis of human endothelial cells. Apoptosis 2001;6:431–439.|
|114.||Grassme H, Gulbins E, Brenner B, Ferlinz K, Sandhoff K, Harzer K, Lang F, Meyer TF. Acidic sphingomyelinase mediates entry of N. gonorrhoeae into nonphagocytic cells. Cell 1997;91:605–615.|
|115.||Grassme H, Riehle A, Wilker B, Gulbins E. Rhinoviruses infect human epithelial cells via ceramide enriched membrane platforms. J Biol Chem 2005;280:26256–26262.|
|116.||Zhang Y, Mattjus P, Schmid PC, Dong Z, Zhong S, Ma WY, Brown RE, Bode AM, Schmid HH, Dong Z. Involvement of the acid sphingomyelinase pathway in uva-induced apoptosis. J Biol Chem 2001;276:11775–11782.|
|117.||Kashkar H, Wiegmann K, Yazdanpanah B, Haubert D, Krönke M. Acid sphingomyelinase is indispensable for UV light induced Bax conformational change at the mitochondrial membrane. J Biol Chem 2005;280:20804–20813.|
|118.||Rotolo JA, Zhang J, Donepudi M, Lee H, Fuks Z, Kolesnick R. Caspase dependent and independent activation of acid sphingomyelinase signaling. J Biol Chem 2005;280:26425–26434.|
|119.||Lang PA, Schenck M, Nicolay JP, Becker JU, Kempe DS, Lupescu A, Koka S, Eisele K, Klarl BA, Rubben H, et al. Liver cell death and anemia in Wilson disease involve acid sphingomyelinase and ceramide. Nat Med 2007;13:164–170.|
|120.||Morita Y, Perez GI, Paris F, Miranda SR, Ehleiter D, Haimovitz Friedman A, Fuks Z, Xie Z, Reed JC, Schuchman EH, et al. Oocyte apoptosis is suppressed by disruption of the acid sphingomyelinase gene or by sphingosine 1 phosphate therapy. Nat Med 2000;6:1109–1114.|
|121.||Scheel Toellner D, Wang K, Craddock R, Webb PR, McGettrick HM, Assi LK, Parkes N, Clough LE, Gulbins E, Salmon M, et al. Reactive oxygen species limit neutrophil life span by activating death receptor signaling. Blood 2004;104:2557–2564.|
|122.||Heinrich M, Wickel M, Schneider Brachert W, Sandberg C, Gahr J, Schwandner R, Weber T, Saftig P, Peters C, Brunner J, et al. Cathepsin D targeted by acid sphingomyelinase derived ceramide. EMBO J 1999;18:5252–5263.|
|123.||Huwiler A, Johansen B, Skarstad A, Pfeilschifter J. Ceramide binds to the CaLB domain of cytosolic phospholipase A2 and facilitates its membrane docking and arachidonic acid release. FASEB J 2001;15:7–9.|
|124.||Zhang Y, Yao B, Delikat S, Bayoumy S, Lin XH, Basu S, McGinley M, Chan Hui PY, Lichenstein H, Kolesnick R. Kinase suppressor of Ras is ceramide activated protein kinase. Cell 1997;89:63–72.|
|125.||Dobrowsky RT, Hannun YA. Ceramide activated protein phosphatase: partial purification and relationship to protein phosphatase 2A. Adv Lipid Res 1993;25:91–104.|
|126.||Muller G, Ayoub M, Storz P, Rennecke J, Fabbro D, Pfizenmaier K. PKC ζ is a molecular switch in signal transduction of TNF α, bifunctionally regulated by ceramide and arachidonic acid. EMBO J 1995;14:1961–1969.|
|127.||Yao B, Zhang Y, Delikat S, Mathias S, Basu S, Kolesnick R. Phosphorylation of Raf by ceramide activated protein kinase. Nature 1995;378:307–310.|
|128.||Gulbins E, Szabo I, Baltzer K, Lang F. Ceramide induced inhibition of T lymphocyte voltage gated potassium channel is mediated by tyrosine kinases. Proc Natl Acad Sci USA 1997;94:7661–7666.|
|129.||Lepple Wienhues A, Belka C, Laun T, Jekle A, Walter B, Wieland U, Welz M, Heil L, Kun J, Busch G, et al. Stimulation of CD95 (Fas) blocks T lymphocyte calcium channels through sphingomyelinase and sphingolipids. Proc Natl Acad Sci USA 1999;96:13795–13800.|
|130.||Siskind LJ, Kolesnick RN, Colombini M. Ceramide channels increase the permeability of the mitochondrial outer membrane to small proteins. J Biol Chem 2002;277:26796–26803.|
|131.||Schuchman EH. The pathogenesis and treatment of acid sphingomyelinase deficient Niemann Pick disease. J Inherit Metab Dis 2007;30:654–663.|
|132.||Mendelson DS, Wasserstein MP, Desnick RJ, Glass R, Simpson W, Skloot G, Vanier M, Bembi B, Giugliani R, Mengel E, et al. Niemann Pick disease: findings at chest radiography, thin section CT, and pulmonary function testing. Radiology 2006;238:339–345.|
|133.||McGovern MM, Wasserstein MP, Giugliani R, Bembi B, Vanier MT, Mengel E, Brodie SE, Mendelson D, Skloot G, Desnick RJ, et al. A prospective, cross-sectional survey study of the natural history of Niemann-Pick disease type B. Pediatrics 2008;122:e341–e349.|
|134.||Simonaro CM, Desnick RJ, McGovern MM, Wasserstein MP, Schuchman EH. The demographics and distribution of type B Niemann Pick disease: novel mutations lead to new genotype/phenotype correlations. Am J Hum Genet 2002;71:1413–1419.|
|135.||Guillemot N, Troadec C, de Villemeur TB, Clement A, Fauroux B. Lung disease in Niemann Pick disease. Pediatr Pulmonol 2007;42:1207–1214.|
|136.||Miranda SR, He X, Simonaro CM, Gatt S, Dagan A, Desnick RJ, Schuchman EH. Infusion of recombinant human acid sphingomyelinase into niemann pick disease mice leads to visceral, but not neurological, correction of the pathophysiology. FASEB J 2000;14:1988–1995.|
|137.||Niessen F, Schaffner F, Furlan Freguia C, Pawlinski R, Bhattacharjee G, Chun J, Derian CK, Andrade Gordon P, Rosen H, Ruf W. Dendritic cell PAR1–S1P3 signalling couples coagulation and inflammation. Nature 2008;252:654–658.|
|138.||Romiti E, Vasta V, Meacci E, Farnararo M, Linke T, Ferlinz K, Sandhoff K, Bruni P. Characterization of sphingomyelinase activity released by thrombin stimulated platelets. Mol Cell Biochem 2000;205:75–81.|
|139.||Ryan AJ, McCoy DM, McGowan SE, Salome RG, Mallampalli RK. Alveolar sphingolipids generated in response to TNF α modifies surfactant biophysical activity. J Appl Physiol 2003;94:253–258.|
|140.||Wong ML, Xie B, Beatini N, Phu P, Marathe S, Johns A, Gold PW, Hirsch E, Williams KJ, Licinio J, et al. Acute systemic inflammation up regulates secretory sphingomyelinase in vivo: a possible link between inflammatory cytokines and atherogenesis. Proc Natl Acad Sci USA 2000;97:8681–8686.|
|141.||Claus RA, Bunck AC, Bockmeyer CL, Brunkhorst FM, Lösche W, Kinscherf R, Deigner H P. Role of increased sphingomyelinase activity in apoptosis and organ failure of patients with severe sepsis. FASEB J 2005;19:1719–1721.|
|142.||von Bismarck P, Garcia Wistadt CF, Klemm K, Winoto Morbach S, Uhlig U, Schütze S, Adam D, Lachmann B, Uhlig S, Krause MF. Improved pulmonary function by acid sphingomyelinase inhibition in a newborn piglet lavage model. Am J Respir Crit Care Med 2008;177:1233–1241.|
|143.||von Bismarck P, Klemm K, Wistadt CF, Winoto Morbach S, Uhlig U, Schutze S, Uhlig S, Lachmann B, Krause MF. Surfactant “fortification” by topical inhibition of nuclear factor κB activity in a newborn piglet lavage model. Crit Care Med 2007;35:2309–2318.|
|144.||Drobnik W, Liebisch G, Audebert FX, Frohlich D, Gluck T, Vogel P, Rothe G, Schmitz G. Plasma ceramide and lysophosphatidylcholine inversely correlate with mortality in sepsis patients. J Lipid Res 2003;44:754–761.|
|145.||Delogu G, Famularo G, Amati F, Signore L, Antonucci A, Trinchieri V, Di ML, Cifone MG. Ceramide concentrations in septic patients: a possible marker of multiple organ dysfunction syndrome. Crit Care Med 1999;27:2413–2417.|
|146.||Machleidt T, Kramer B, Adam D, Neumann B, Schütze S, Wiegmann K, Kronke M. Function of the p55 tumor necrosis factor receptor “death domain” mediated by phosphatidylcholine specific phospholipase C. J Exp Med 1996;184:725–733.|
|147.||Haimovitz Friedman A, Cordon Cardo C, Bayoumy S, Garzotto M, McLoughlin M, Gallily R, Edwards CK, Schuchman EH, Fuks Z, Kolesnick R. Lipopolysaccharide induces disseminated endothelial apoptosis requiring ceramide generation. J Exp Med JT . J Exp Med 1997;186:1831–1841.|
|148.||Vandivier RW, Henson PM, Douglas IS. Burying the dead: the impact of failed apoptotic cell removal (efferocytosis) on chronic inflammatory lung disease. Chest 2006;129:1673–1682.|
|149.||Uhlig S, Burdon D. Pro- and anti-inflammatory Cytokines and apoptosis in acute lung injury. In Baue AE, Berlot G, Gullo A, Vincent J L, editors. Sepsis and organ dysfunction: from chaos to rationale. Milano: Springer Verlag; 2002. pp. 221–234.|
|150.||Matute Bello G, Martin TR. Science review: apoptosis in acute lung injury. Crit Care 2003;7:355–358.|
|151.||Martin TR, Hagimoto N, Nakamura M, Matute Bello G. Apoptosis and epithelial injury in the lungs. Proc Am Thorac Soc 2005;2:214–220.|
|152.||Kuwano K. Involvement of epithelial cell apoptosis in interstitial lung diseases. Intern Med 2008;47:345–353.|
|153.||Abraham E. Neutrophils and acute lung injury. Crit Care Med 2003;31:S195–S199.|
|154.||McVerry BJ, Garcia JG. Endothelial cell barrier regulation by sphingosine 1 phosphate. J Cell Biochem 2004;92:1075–1085.|
|155.||Vandenbroucke E, Mehta D, Minshall R, Malik AB. Regulation of endothelial junctional permeability. Ann N Y Acad Sci 2008;1123:134–145.|
|156.||Lee MJ, Thangada S, Claffey KP, Ancellin N, Liu CH, Kluk M, Volpi M, Sha'afi RI, Hla T. Vascular endothelial cell adherens junction assembly and morphogenesis induced by sphingosine 1 phosphate. Cell 1999;99:301–312.|
|157.||Garcia JG, Liu F, Verin AD, Birukova A, Dechert MA, Gerthoffer WT, Bamberg JR, English D. Sphingosine 1 phosphate promotes endothelial cell barrier integrity by Edg dependent cytoskeletal rearrangement. J Clin Invest 2001;108:689–701.|
|158.||Okazaki M, Kreisel F, Richardson SB, Kreisel D, Krupnick AS, Patterson GA, Gelman AE. Sphingosine 1 phosphate inhibits ischemia reperfusion injury following experimental lung transplantation. Am J Transplant 2007;7:751–758.|
|159.||McVerry BJ, Peng X, Hassoun PM, Sammani S, Simon BA, Garcia JG. Sphingosine 1 phosphate reduces vascular leak in murine and canine models of acute lung injury. Am J Respir Crit Care Med 2004;170:987–993.|
|160.||Sanchez T, Skoura A, Wu MT, Casserly B, Harrington EO, Hla T. Induction of vascular permeability by the sphingosine 1 phosphate receptor 2 (S1P2R) and its downstream effectors ROCK and PTEN. Arterioscler Thromb Vasc Biol 2007;27:1312–1318.|
|161.||Gon Y, Wood MR, Kiosses WB, Jo E, Sanna MG, Chun J, Rosen H. S1P3 receptor induced reorganization of epithelial tight junctions compromises lung barrier integrity and is potentiated by TNF. Proc Natl Acad Sci USA 2005;102:9270–9275.|
|162.||Singleton PA, Dudek SM, Ma SF, Garcia JG. Transactivation of sphingosine 1 phosphate receptors is essential for vascular barrier regulation. Novel role for hyaluronan and CD44 receptor family. J Biol Chem 2006;281:34381–34393.|
|163.||Singleton PA, Moreno Vinasco L, Sammani S, Wanderling SL, Moss J, Garcia JG. Attenuation of vascular permeability by methylnaltrexone: role of mOP R and S1P3 transactivation. Am J Respir Cell Mol Biol 2007;37:222–231.|
|164.||Feistritzer C, Riewald M. Endothelial barrier protection by activated protein C through PAR1 dependent sphingosine 1 phosphate receptor 1 crossactivation. Blood 2005;105:3178–3184.|
|165.||Finigan JH, Dudek SM, Singleton PA, Chiang ET, Jacobson JR, Camp SM, Ye SQ, Garcia JG. Activated protein C mediates novel lung endothelial barrier enhancement: role of sphingosine 1 phosphate receptor transactivation. J Biol Chem 2005;280:17286–17293.|
|166.||Lindner K, Uhlig U, Uhlig S. Ceramide alters endothelial cell permeability by a nonapoptotic mechanism. Br J Pharmacol 2005;145:132–140.|
|167.||Uhlig S, Göggel R, Engel S. Mechanisms of platelet activating factor (PAF) mediated responses in the lung. Pharmacol Rep 2005;57:206–221.|
|168.||Balsinde J, Balboa MA, Dennis EA. Inflammatory activation of arachidonic acid signaling in murine P388D1 macrophages via sphingomyelin synthesis. J Biol Chem 1997;272:20373–20377.|
|169.||Göggel R, Hoffman S, Nüsing R, Narumiya S, Uhlig S. PAF induced pulmonary edema is partly mediated by PGE2, EP3 receptors and potassium channels. Am J Respir Crit Care Med 2002;166:657–662.|
|170.||Mallampalli RK, Mathur SN, Warnock LJ, Salome RG, Hunninghake GW, Field FJ. Betamethasone modulation of sphingomyelin hydrolysis up regulates CTP:cholinephosphate cytidylyltransferase activity in adult rat lung. Biochem J 1996;318:333–341.|
|171.||Göggel R, Uhlig S. The inositol trisphosphate pathway mediates platelet activating factor induced pulmonary oedema. Eur Respir J 2005;25:849–857.|
|172.||Oskeritzian CA, Milstien S, Spiegel S. Sphingosine 1 phosphate in allergic responses, asthma and anaphylaxis. Pharmacol Ther 2007;115:390–399.|
|173.||Ammit AJ, Hastie AT, Edsall LC, Hoffman RK, Amrani Y, Krymskaya VP, Kane SA, Peters SP, Penn RB, Spiegel S, et al. Sphingosine 1 phosphate modulates human airway smooth muscle cell functions that promote inflammation and airway remodeling in asthma. FASEB J 2001;15:1212–1214.|
|174.||Idzko M, Panther E, Corinti S, Morelli A, Ferrari D, Herouy Y, Dichmann S, Mockenhaupt M, Gebicke Haerter P, Di VF, et al. Sphingosine 1 phosphate induces chemotaxis of immature and modulates cytokine release in mature human dendritic cells for emergence of Th2 immune responses. FASEB J 2002;16:625–627.|
|175.||Matloubian M, Lo CG, Cinamon G, Lesneski MJ, Xu Y, Brinkmann V, Allende ML, Proia RL, Cyster JG. Lymphocyte egress from thymus and peripheral lymphoid organs is dependent on S1P receptor 1. Nature 2004;427:355–360.|
|176.||Field KA, Holowka D, Baird B. Structural aspects of the association of FcepsilonRI with detergent resistant membranes. J Biol Chem 1999;274:1753–1758.|
|177.||Olivera A, Mizugishi K, Tikhonova A, Ciaccia L, Odom S, Proia RL, Rivera J. The sphingosine kinase sphingosine 1 phosphate axis is a determinant of mast cell function and anaphylaxis. Immunity 2007;26:287–297.|
|178.||Olivera A, Rivera J. Sphingolipids and the balancing of immune cell function: lessons from the mast cell. J Immunol 2005;174:1153–1158.|
|179.||Nishiuma T, Nishimura Y, Okada T, Kuramoto E, Kotani Y, Jahangeer S, Nakamura S. Inhalation of sphingosine kinase inhibitor attenuates airway inflammation in asthmatic mouse model. Am J Physiol Lung Cell Mol Physiol 2008;294:L1085–L1093.|
|180.||Beaven MA. Division of labor: specialization of sphingosine kinases in mast cells. Immunity 2007;26:271–273.|
|181.||Chiba N, Masuda A, Yoshikai Y, Matsuguchi T. Ceramide inhibits LPS induced production of IL 5, IL 10, and IL 13 from mast cells. J Cell Physiol 2007;213:126–136.|
|182.||Watterson KR, Ratz PH, Spiegel S. The role of sphingosine 1 phosphate in smooth muscle contraction. Cell Signal 2005;17:289–298.|
|183.||Rosenfeldt HM, Amrani Y, Watterson KR, Murthy KS, Panettieri RJ, Spiegel S. Sphingosine 1 phosphate stimulates contraction of human airway smooth muscle cells. FASEB J 2003;17:1789–1799.|
|184.||Roviezzo F, Di LA, Bucci M, Brancaleone V, Vellecco V, De NM, Orlotti D, De PR, Rossi F, D'Agostino B, et al. Sphingosine 1 phosphate/sphingosine kinase pathway is involved in mouse airway hyperresponsiveness. Am J Respir Cell Mol Biol 2007;36:757–762.|
|185.||Kume H, Takeda N, Oguma T, Ito S, Kondo M, Ito Y, Shimokata K. Sphingosine 1 phosphate causes airway hyper reactivity by rho mediated myosin phosphatase inactivation. J Pharmacol Exp Ther 2007;320:766–773.|
|186.||Ediger TL, Toews ML. Synergistic stimulation of airway smooth muscle cell mitogenesis. J Pharmacol Exp Ther 2000;294:1076–1082.|
|187.||Masini E, Giannini L, Nistri S, Cinci L, Mastroianni R, Xu W, Comhair SA, Li D, Cuzzocrea S, Matuschak GM, et al. Ceramide: a key signaling molecule in a Guinea pig model of allergic asthmatic response and airway inflammation. J Pharmacol Exp Ther 2008;324:548–557.|
|188.||Wenzel S, Holgate ST. The mouse trap: it still yields few answers in asthma. Am J Respir Crit Care Med 2006;174:1173–1176.|
|189.||Ressmeyer AR, Larsson AK, Vollmer E, Dahlen SE, Uhlig S, Martin C. Characterisation of guinea pig precision cut lung slices: comparison with human tissues. Eur Respir J 2006;28:603–611.|
|190.||Melendez AJ, Khaw AK. Dichotomy of Ca2+ signals triggered by different phospholipid pathways in antigen stimulation of human mast cells. J Biol Chem 2002;277:17255–17262.|
|191.||Grassme H, Kirschnek S, Riethmueller J, Riehle A, von Kürthy G, Lang F, Weller M, Gulbins E. CD95/CD95 ligand interactions on epithelial cells in host defense to Pseudomonas aeruginosa. Science 2000;290:527–530.|
|192.||Pier GB, Grout M, Zaidi TS, Olsen JC, Johnson LG, Yankaskas JR, Goldberg JB. Role of mutant CFTR in hypersusceptibility of cystic fibrosis patients to lung infections. Science 1996;271:64–67.|
|193.||Pier GB, Grout M, Zaidi TS. Cystic fibrosis transmembrane conductance regulator is an epithelial cell receptor for clearance of Pseudomonas aeruginosa from the lung. Proc Natl Acad Sci USA 1997;94:12088–12093.|
|194.||Kowalski MP, Pier GB. Localization of cystic fibrosis transmembrane conductance regulator to lipid rafts of epithelial cells is required for Pseudomonas aeruginosa induced cellular activation. J Immunol 2004;172:418–425.|
|195.||Garg SK, Santucci MB, Panitti M, Pucillo L, Bocchino M, Okajima F, Bisen PS, Saltini C, Fraziano M. Does sphingosine 1 phosphate play a protective role in the course of pulmonary tuberculosis? Clin Immunol 2006;121:260–264.|
|196.||Thompson CR, Iyer SS, Melrose N, VanOosten R, Johnson K, Pitson SM, Obeid LM, Kusner DJ. Sphingosine kinase 1 (SK1) is recruited to nascent phagosomes in human macrophages: inhibition of SK1 translocation by Mycobacterium tuberculosis. J Immunol 2005;174:3551–3561.|
|197.||Garg SK, Volpe E, Palmieri G, Mattei M, Galati D, Martino A, Piccioni MS, Valente E, Bonanno E, De VP, et al. Sphingosine 1 phosphate induces antimicrobial activity both in vitro and in vivo. J Infect Dis 2004;189:2129–2138.|
|198.||Petrache I, Natarajan V, Zhen L, Medler TR, Richter AT, Cho C, Hubbard WC, Berdyshev EV, Tuder RM. Ceramide upregulation causes pulmonary cell apoptosis and emphysema like disease in mice. Nat Med 2005;11:491–498.|
|199.||Vit JP, Rosselli F. Role of the ceramide signaling pathways in ionizing radiation induced apoptosis. Oncogene 2003;22:8645–8652.|
|200.||Petrache I, Medler TR, Richter AT, Kamocki K, Chukwueke U, Zhen L, Gu Y, Adamowicz J, Schweitzer KS, Hubbard WC, et al. Superoxide dismutase protects against apoptosis and alveolar enlargement induced by ceramide. Am J Physiol Lung Cell Mol Physiol 2008;295:L44–L53.|
|201.||Kim EY, Battaile JT, Patel AC, You Y, Agapov E, Grayson MH, Benoit LA, Byers DE, Alevy Y, Tucker J, et al. Persistent activation of an innate immune response translates respiratory viral infection into chronic lung disease. Nat Med 2008;14:633–640.|
|202.||Weber AJ, Soong G, Bryan R, Saba S, Prince A. Activation of NF κB in airway epithelial cells is dependent on CFTR trafficking and Cl channel function. Am J Physiol Lung Cell Mol Physiol 2001;281:L71–L78.|
|203.||Joseph T, Look D, Ferkol T. NF κB activation and sustained IL 8 gene expression in primary cultures of cystic fibrosis airway epithelial cells stimulated with Pseudomonas aeruginosa. Am J Physiol Lung Cell Mol Physiol 2005;288:L471–L479.|
|204.||Zahm JM, Gaillard D, Dupuit F, Hinnrasky J, Porteous D, Dorin JR, Puchelle E. Early alterations in airway mucociliary clearance and inflammation of the lamina propria in CF mice. Am J Physiol 1997;272:C853–C859.|
|205.||Tirouvanziam R, de Bentzmann S, Hubeau C, Hinnrasky J, Jacquot J, Péault B, Puchelle E. Inflammation and infection in naive human cystic fibrosis airway grafts. Am J Respir Cell Mol Biol 2000;23:121–127.|
|206.||Boujaoude LC, Bradshaw Wilder C, Mao C, Cohn J, Ogretmen B, Hannun YA, Obeid LM. Cystic fibrosis transmembrane regulator regulates uptake of sphingoid base phosphates and lysophosphatidic acid: modulation of cellular activity of sphingosine 1 phosphate. J Biol Chem 2001;276:35258–35264.|
|207.||Teichgraber V, Ulrich M, Endlich N, Riethmuller J, Wilker B, De Oliveira Munding CC, van Heeckeren AM, Barr ML, von Kurthy G, Schmid KW. Ceramide accumulation mediates inflammation, cell death and infection susceptibility in cystic fibrosis. Nat Med 2008;14:382–391.|
|208.||Root KV, Engelhardt JF, Post M, Wilson JW, Van Dyke RW. CFTR does not alter acidification of L cell endosomes. Biochem Biophys Res Commun 1994;205:396–401.|
|209.||Seksek O, Biwersi J, Verkman AS. Evidence against defective trans Golgi acidification in cystic fibrosis. J Biol Chem 1996;271:15542–15548.|
|210.||Haggie PM, Verkman AS. Cystic fibrosis transmembrane conductance regulator independent phagosomal acidification in macrophages. J Biol Chem 2007;282:31422–31428.|
|211.||Dreschers S, Franz P, Dumitru C, Wilker B, Jahnke K, Gulbins E. Infections with human rhinovirus induce the formation of distinct functional membrane domains. Cell Physiol Biochem 2007;20:241–254.|
|212.||Griese M. Pulmonary surfactant in health and human lung disease: state of the art. Eur Respir J 1999;13:1455–1476.|
|213.||Buccoliero R, Ginzburg L, Futerman AH. Elevation of lung surfactant phosphatidylcholine in mouse models of Sandhoff and of Niemann Pick A disease. J Inherit Metab Dis 2004;27:641–648.|
|214.||Hallman M, Spragg R, Harrell JH, Moser KM. Evidence of lung surfactant abnormality in respiratory failure. J Clin Invest 1982;70:673–683.|
|215.||Vivekananda J, Smith D, King RJ. Sphingomyelin metabolites inhibit sphingomyelin synthase and CTP:phosphocholine cytidylyltransferase. Am J Physiol Lung Cell Mol Physiol 2001;281:L98–L107.|
|216.||Hannun YA, Obeid LM. Principles of bioactive lipid signalling: lessons from sphingolipids. Nat Rev Mol Cell Biol 2008;9:139–150.|
|217.||Nixon GF, Mathieson FA, Hunter I. The multi functional role of sphingosylphosphorylcholine. Prog Lipid Res 2008;47:62–75.|
|218.||Berdyshev EV, Gorshkova IA, Usatyuk P, Zhao Y, Saatian B, Hubbard W, Natarajan V. De novo biosynthesis of dihydrosphingosine 1 phosphate by sphingosine kinase 1 in mammalian cells. Cell Signal 2006;18:1779–1792.|
|219.||He X, Okino N, Dhami R, Dagan A, Gatt S, Schulze H, Sandhoff K, Schuchman EH. Purification and characterization of recombinant, human acid ceramidase. Catalytic reactions and interactions with acid sphingomyelinase. J Biol Chem 2003;278:32978–32986.|
|220.||Romiti E, Meacci E, Donati C, Formigli L, Zecchi Orlandini S, Farnararo M, Ito M, Bruni P. Neutral ceramidase secreted by endothelial cells is released in part associated with caveolin 1. Arch Biochem Biophys 2003;417:27–33.|
|221.||Marathe S, Kuriakose G, Williams KJ, Tabas I. Sphingomyelinase, an enzyme implicated in atherogenesis, is present in atherosclerotic lesions and binds to specific components of the subendothelial extracellular matrix. Arterioscler Thromb Vasc Biol 1999;19:2648–2658.|
|222.||Takahashi T, Abe T, Sato T, Miura K, Takahashi I, Yano M, Watanabe A, Imashuku S, Takada G. Elevated sphingomyelinase and hypercytokinemia in hemophagocytic lymphohistiocytosis. J Pediatr Hematol Oncol 2002;24:401–404.|
|223.||Doehner W, Bunck AC, Rauchhaus M, von Haehling S, Brunkhorst FM, Cicoira M, Tschope C, Ponikowski P, Claus RA, Anker SD. Secretory sphingomyelinase is upregulated in chronic heart failure: a second messenger system of immune activation relates to body composition, muscular functional capacity, and peripheral blood flow. Eur Heart J 2007;28:821–828.|
|224.||Kornhuber J, Medlin A, Bleich S, Jendrossek V, Henkel AW, Wiltfang J, Gulbins E. High activity of acid sphingomyelinase in major depression. J Neural Transm 2005;112:1583–1590.|
|225.||Gorska M, Baranczuk E, Dobrzyn A. Secretory Zn2+ dependent sphingomyelinase activity in the serum of patients with type 2 diabetes is elevated. Horm Metab Res 2003;35:506–507.|
|226.||Quintern LE, Zenk TS, Sandhoff K. The urine from patients with peritonitis as a rich source for purifying human acid sphingomyelinase and other lysosomal enzymes. Biochim Biophys Acta 1989;1003:121–124.|
|227.||Karuna R, Sashidhar RB. The mycotoxin fumonisin B1 inhibits eukaryotic protein synthesis: in vitro and in vivo studies. Mycopathologia 2008;165:37–49.|
|228.||Nara F, Tanaka M, Hosoya T, Suzuki Konagai K, Ogita T. Scyphostatin, a neutral sphingomyelinase inhibitor from a discomycete, Trichopeziza mollissima: taxonomy of the producing organism, fermentation, isolation, and physico chemical properties. J Antibiot (Tokyo) 1999;52:525–530.|
|229.||Sakata A, Yasuda K, Ochiai T, Shimeno H, Hikishima S, Yokomatsu T, Shibuya S, Soeda S. Inhibition of lipopolysaccharide induced release of interleukin 8 from intestinal epithelial cells by SMA, a novel inhibitor of sphingomyelinase and its therapeutic effect on dextran sulphate sodium induced colitis in mice. Cell Immunol 2007;245:24–31.|
|230.||Arenz C, Gartner M, Wascholowski V, Giannis A. Synthesis and biochemical investigation of scyphostatin analogues as inhibitors of neutral sphingomyelinase. Bioorg Med Chem 2001;9:2901–2904.|
|231.||Schwab SR, Pereira JP, Matloubian M, Xu Y, Huang Y, Cyster JG. Lymphocyte sequestration through S1P lyase inhibition and disruption of S1P gradients. Science 2005;309:1735–1739.|
|232.||Rosen H, Sanna G, Alfonso C. Egress: a receptor regulated step in lymphocyte trafficking. Immunol Rev 2003;195:160–177.|
|233.||Gorshkova I, He D, Berdyshev E, Usatuyk P, Burns M, Kalari S, Zhao Y, Pendyala S, Garcia JG, Pyne NJ, et al. Protein kinase Cε regulates sphingosine 1 phosphate mediated migration of human lung endothelial cells through activation of phospholipase D2, protein kinase Cζ, and Rac1. J Biol Chem 2008;283:11794–11806.|
|234.||Ikegami M, Dhami R, Schuchman EH. Alveolar lipoproteinosis in an acid sphingomyelinase deficient mouse model of Niemann Pick disease. Am J Physiol Lung Cell Mol Physiol 2003;284:L518–L525.|