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Melanomas reprogram their metabolism to rapidly adapt to therapy-induced stress conditions, allowing them to persist and ultimately develop resistance. We report that a subpopulation of melanoma cells tolerate MAPK pathway inhibitors (MAPKis) through a concerted metabolic reprogramming mediated by peroxisomes and UDP-glucose ceramide glycosyltransferase (UGCG). Compromising peroxisome biogenesis, by repressing PEX3 expression, potentiated the proapoptotic effects of MAPKis via an induction of ceramides, an effect limited by UGCG-mediated ceramide metabolism. Cotargeting PEX3 and UGCG selectively eliminated a subset of metabolically active, drug-tolerant [CD36.sup.+] melanoma persister cells, thereby sensitizing melanoma to MAPKis and delaying resistance. Increased levels of peroxisomal genes and UGCG were found in patient-derived MAPKi-relapsed melanomas, and simultaneously inhibiting PEX3 and UGCG restored MAPKi sensitivity in multiple models of therapy resistance. Finally, combination therapy consisting of a newly identified inhibitor of the PEX3PEX19 interaction, a UGCG inhibitor, and MAPKis demonstrated potent antitumor activity in preclinical melanoma models, thus representing a promising approach for melanoma treatment., Introduction Melanoma is the deadliest form of skin cancer. Current treatments for patients with metastatic melanoma include therapies that target the MAPK pathway (BRAF and MEK inhibitors) and immunotherapies targeting [...]

Cardiolipidology - a new direction in cardiology - is developing intensively owing to the method of mass spectrometry. This method has acquired particular importance for the determination of molecular types of sphingolipids, the metabolism of which is closely related to the metabolism of cholesterol. Changes in the level of a number of sphingolipids claiming to be markers of cardiovascular diseases (CVDs) (molecular forms of sphingomyelins, ceramides, glucosylceramides, sphingosine, and sphinganin), in the blood plasma of patients with hereditary forms of early atherosclerosis (familial hyperlipidemia, FHL) were studied by chromatography-mass spectrometry. The study group (52 people) consisted of patients with acute manifestations of atherosclerosis, with postinfarction cardiosclerosis, and with dyslipidemia or early CVD. In patients with FHL, there was an increase in the proportion of long-chain sphingomyelin SM18:1/22:0 and the level of ceramides with carbon chains C20-1 and C22-1. An increase in the level of sphingosine, possessing proapoptotic properties, which can be considered as a marker of an additional risk of cardiovascular complications, was revealed in patients with a high clinical probability of FHL. The search for new CVD markers will improve personalized approach to the management of such patients and improve the results of their treatment., Author(s): A. V. Alessenko [sup.1], M. A. Shupik [sup.1], U. A. Gutner [sup.1], D. A. Zateyshchikov [sup.2], L. O. Minushkina [sup.2], A. A. Rogozhina [sup.2], A. T. Lebedev [sup.3], O. [...]

Risk for childhood asthma is conferred by alleles within the 17q21 locus affecting ORMDL sphingolipid biosynthesis regulator 3 (ORMDL3) expression. ORMDL3 inhibits sphingolipid de novo synthesis. Although the effects of 17q21 genotypes on sphingolipid synthesis in human asthma remain unclear, both decreased sphingolipid synthesis and ORMDL3 overexpression are linked to airway hyperreactivity. To characterize the relationship of genetic asthma susceptibility with sphingolipid synthesis, we analyzed asthma-associated 17q21 genotypes (rs7216389, rs8076131, rs4065275, rs12603332, and rs8067378) in both children with asthma and those without asthma, quantified plasma and whole-blood sphingolipids, and assessed sphingolipid de novo synthesis in peripheral blood cells by measuring the incorporation of stable isotopelabeled serine (substrate) into sphinganine and sphinganine-1-phosphate. Whole-blood dihydroceramides and ceramides were decreased in subjects with the 17q21 asthma-risk alleles rs7216389 and rs8076131. Children with nonallergic asthma had lower dihydroceramides, ceramides, and sphingomyelins than did controls. Children with allergic asthma had higher dihydroceramides, ceramides, and sphingomyelins compared with children with nonallergic asthma. Additionally, de novo sphingolipid synthesis was lower in children with asthma compared with controls. These findings connect genetic 17q21 variations that are associated with asthma risk and higher ORMDL3 expression to lower sphingolipid synthesis in humans. Altered sphingolipid synthesis may therefore be a critical factor in asthma pathogenesis and may guide the development of future therapeutics., Introduction Asthma in children is characterized by airway hyperreactivity and a strong genetic predisposition. The asthma-associated susceptibility locus 17q21 is strongly linked to childhood-onset asthma (1), and asthma-associated genotypes in [...]

Adipose tissue plays important roles in regulating whole-body energy metabolism through its storage function in white adipocytes and its dissipating function in brown and beige adipocytes. Adipose tissue also produces a variety of secreted factors called adipocytokines, including leptin and adiponectin. Furthermore, recent studies have suggested the important roles of extracellular vesicles of endosomal origin termed exosomes, which are secreted from adipocytes and other cells in adipose tissue and influence whole-body glucose and lipid metabolism. Adiponectin is known to be a pleiotropic organ-protective protein that is exclusively produced by adipocytes and decreased in obesity. Adiponectin accumulates in tissues such as heart, muscle, and vascular endothelium through binding with T-cadherin, a glycosylphosphatidylinositol-anchored (GPI-anchored) cadherin. Recently, adiponectin was found to enhance exosome biogenesis and secretion, leading to a decrease in cellular ceramides, excess of which is known to cause insulin resistance and cardiovascular disease phenotypes. These findings support the hypothesis that adipose tissue metabolism systemically regulates exosome production and whole-body metabolism through exosomes. This review focuses on intra-adipose and interorgan communication by exosomes, adiponectin-stimulated exosome production, and their dysregulation in metabolic diseases., Introduction Obesity is associated with an array of complications, including cardiovascular diseases, type 2 diabetes (T2D), fatty liver disease, osteoporosis, airway disease, degenerative disorders including dementia, and certain cancers. In [...]

Nearly all stress stimuli (e.g., inflammatory cytokines, glucocorticoids, chemotherapeutics, etc.) induce sphingolipid synthesis, leading to the accumulation of ceramides and ceramide metabolites. While the role of these lipids in the [...]

The present study tested the hypotheses that 1) short-term dietary deficiency of magnesium (21 days) in rats would result in the upregulation of sphingomyelin synthase (SMS) and p53 in cardiac and vascular (aortic) smooth muscles, 2) low levels of [Mg.sup.2+] added to drinking water would either prevent or greatly reduce the upregulation of both SMS and p53, 3) exposure of primary cultured vascular smooth muscle cells (VSMCs) to low extracellular [Mg.sup.2+] concentration ([[[Mg.sup.2]].sub.o]) would lead to the de novo synthesis of ceramide, 4) inhibition of either SMS or p53 in primary culture VSMCs exposed to low [[[Mg.sup.2+]].sub.o] would lead to reductions in the levels of de novo ceramide synthesis, and 5) inhibition of sphingomyelin palmitoyl-CoA transferase (SPT) or ceramide synthase (CS) in primary cultured VSMCs exposed to low [[[Mg.sup.2+]].sub.o] would lead to a reduction in the levels of de novo ceramide synthesis. The data indicated that short-term magnesium deficiency (10% normal dietary intake) resulted in the upregulation of SMS and p53 in both ventricular and aortic smooth muscles; even very low levels of water-borne [Mg.sup.2+] (e.g., 15 mg x [l.sup.-1] x [day.sup.-1]) either prevented or ameliorated the upregulation in SMS and p53. Our experiments also showed that VSMCs exposed to low [[[Mg.sup.2+]].sub.o] resulted in the de novo synthesis of ceramide; the lower the [[[Mg.sup.2+]].sub.o], the greater the synthesis of ceramide. In addition, the data indicated that inhibition of either SMS, p53, SPT, or CS in VSMCs exposed to low [[[Mg.sup.2+]].sub.o] resulted in marked reductions in the de novo synthesis of ceramide. ceramide synthase; de novo ceramide; water-borne magnesium; cardiac muscle; vascular muscle doi: 10.1152/ajpheart.00671.2010.

Although obesity is associated with multiple features of the metabolic syndrome (insulin resistance, leptin resistance, hepatic steatosis, chronic inflammation, etc.), the molecular changes that promote these conditions are not completely understood. Here, we tested the hypothesis that elevated ceramide biosynthesis contributes to the pathogenesis of obesity and the metabolic syndrome. Chronic treatment for 8 wk of genetically obese (ob/ob), and, high-fat diet-induced obese (DIO) mice with myriocin, an inhibitor of de novo ceramide synthesis, decreased circulating ceramides. Decreased ceramide was associated with reduced weight, enhanced metabolism and energy expenditure, decreased hepatic steatosis, and improved glucose hemostasis via enhancement of insulin signaling in the liver and muscle. Inhibition of de novo ceramide biosynthesis decreased adipose expression of suppressor of cytokine signaling-3 (SOCS-3) and induced adipose uncoupling protein-3 (UCP3). Moreover, ceramide directly induced SOCS-3 and inhibited UCP3 mRNA in cultured adipocytes suggesting a direct role for ceramide in regulation of metabolism and energy expenditure. Inhibition of de novo ceramide synthesis had no effect on adipose tumor necrosis factor-[alpha] (TNF-[alpha]) expression but dramatically reduced adipose plasminogen activator inhibitor-1 (PAI-1) and monocyte chemoattactant protein-1 (MCP-1). This study highlights a novel role for ceramide biosynthesis in body weight regulation, energy expenditure, and the metabolic syndrome. sphingolipids; insulin resistance; leptin resistance; insulin resistance; suppressor of cytokine signaling-3; tumor necrosis factor-[alpha]; plasminogen activator inhibitor-1; monocyte chemoattactant protein-1

Ceramides are central intermediates of sphingo-lipid metabolism with critical functions in cell organization and survival. They are synthesized on the cytosolic surface of the endoplasmic reticulum (ER) and transported by ceramide transfer protein to the Golgi for conversion to sphingomyelin (SM) by SM synthase SMS1. In this study, we report the identification of an SMS1-related (SMSr) enzyme, which catalyses the synthesis of the SM analogue ceramide phosphoethanolamine (CPE)in the ER lumen. Strikingly, SMSr produces only trace amounts of CPE, i.e., 300-fold less than SMS1-derived SM. Nevertheless, blocking its catalytic activity causes a substantial rise in ER ceramide levels and a structural collapse of the early secretory pathway. We find that the latter phenotype is not caused by depletion of CPE but rather a consequence of ceramide accumulation in the ER. Our results establish SMSr as a key regulator of ceramide homeostasis that seems to operate as a sensor rather than a converter of ceramides in the ER.

The molecular events leading to emphysema development include generation of oxidative stress and alveolar cell apoptosis. Oxidative stress upregulates ceramides, proapoptotic signaling sphingolipids that trigger further oxidative stress and alveolar space enlargement, as shown in an experimental model of emphysema due to VEGF blockade. As alveolar cell apoptosis and oxidative stress mutually interact to mediate alveolar destruction, we hypothesized that the oxidative stress generated by ceramide is required for its pathogenic effect on lung alveoli. To model the direct lung effects of ceramide, mice received ceramide intratracheally ([Cer.sub.12:0] or [Cer.sub.8:0]; 1 mg/kg) or vehicle. Apoptosis was inhibited with a general caspase inhibitor. Ceramide augmentation shown to mimic levels found in human emphysema lungs increased oxidative stress, and decreased, independently of caspase activation, the lung superoxide dismutase activity at 48 h. In contrast to their wild-type littermates, transgenic mice over-expressing human Cu/Zn SOD were significantly protected from ceramide-induced superoxide production, apoptosis, and air space enlargement. Activation of lung acid sphingomyelinase in response to ceramide treatment was abolished in the Cu/Zn SOD transgenic mice. Since cigarette smoke-induced emphysema in mice is similarly ameliorated by the Cu/Zn SOD overexpression, we hypothesized that cigarette smoke may induce ceramides in the mouse lung. Utilizing tandem mass spectrometry, we documented increased lung ceramides in adult mice exposed to cigarette smoke for 4 wk. In conclusion, ceramide-induced superoxide accumulation in the lung may be a critical step in ceramide's proapoptotic effect in the lung. This work implicates excessive lung ceramides as amplifiers of lung injury through redox-dependent mechanisms. sphingolipids; cell death; lung; oxidative stress

Activation of cells by receptor- and nonreceptor-mediated stimuli not only requires a change in the activity of signaling proteins but also requires a reorganization of the topology of the signalosom in the cell. The cell membrane contains distinct domains, rafts that serve the spatial organization of signaling molecules in the cell. Many receptors or stress stimuli transform rafts by the generation of ceramide. These stimuli activate the acid sphingomyelinase and induce a translocation of this enzyme onto the extracellular leaflet of the cell membrane. Surface acid sphingomyelinase generates ceramide that serves to fuse small rafts and to form large ceramide-enriched membrane platforms. These platforms cluster receptor molecules, recruit intracellular signaling molecules to aggregated receptors, and seem to exclude inhibitory signaling factors. Thus ceramide-enriched membrane platforms do not seem to be part of a specific signaling pathway but may facilitate and amplify the specific signaling elicited by the cognate stimulus. This general function may enable these membrane domains to be critically involved in the induction of apoptosis by death receptors and stress stimuli, bacterial and viral infections of mammalian cells, and the regulation of cardiovascular functions. signal transduction; acid sphingomyelinase; rafts; membrane platforms

Ceramide, the basic structural unit of sphingolipids, controls the balance between cell growth and death by inducing apoptosis. We have previously shown that accumulation of ceramide, triggered by hydrogen peroxide ([H.sub.2][O.sub.2]) or by short-chain ceramide analogs, induces apoptosis of lung epithelial cells. Here we elucidate the link between caspase-3 activation, at the execution phase, and ceramide accumulation, at the commitment phase of apoptosis in A549 human lung adenocarcinoma cells. The induction of ceramide accumulation by various triggers of ceramide generation, such as [H.sub.2][O.sub.2], [C.sub.6]-ceramide, or UDP-glucose-ceramide glucosyltransferase inhibitor DL-threo-1-phenyl-2-decanoylamino-3-morpholino-1-propanol, triggered the activation of caspase-3. This ceramide elevation also induced the cleavage of the death substrate poly(ADP-ribose) polymerase and was followed by apoptotic cell death. Ceramide-mediated apoptosis was blocked by a general caspase inhibitor, Boc-D-fluoromethylketone, and by overexpression of the antiapoptotic protein Bcl-2. Notably, overexpression of Bcl-2 reduced the basal cellular levels of ceramide and prevented the induction of ceramide generation by [C.sub.6]-ceramide, which implies ceramide generation as a possible target for the antiapoptotic effects of Bcl-2. Bcl-2 overexpression; hydrogen peroxide; sphingolipid signaling

Diacylglycerol (DAG) and ceramide are important second messengers affecting cell growth, differentiation, and apoptosis. Balb/c-3T3 fibroblast cells expressing dopamine-D2S (short) receptors (Balb-D2S cells) provide a model of G protein-mediated cell growth and transformation. In Balb-D2S cells, apomorphine (E[C.sub.50] = 10 nM) stimulated DAG and ceramide formation by 5.6- and 4.3-fold, respective]y, maximal at 1 h and persisting over 6 h. These actions were blocked by pretreatment with pertussis toxin (PTX), implicating [G.sub.i]/[G.sub.o] proteins. To address which G proteins are involved, Balb-D2S clones expressing individual PTX-insensitive G[[alpha].sub.i] proteins were treated with PTX and tested for apomorphine-induced responses. Neither PTX-insensitive G[[alpha].sub.i2] nor G[[alpha].sub.i3] rescued D2S-induced DAG or ceramide formation. Both D2S-induced DAG and ceramide signals required G[beta][gamma]-subunits and were blocked by inhibitors of phospholipase C [1-(6-[([17[beta]]-3-methoxyestra-1,2,3[10]-trien-17yl) amino]hexyl)-1H-pyrrole-2,5-dione (U-73122) and partially by D609]. The similar G protein specificity of D2S-induced calcium mobilization, DAG, and ceramide formation indicates a common G[beta][gamma]-dependent phospholipase C-mediated pathway. Both D2 agonists and ceramide specifically induced mitogen-activated protein kinase (ERK1/2), suggesting that ceramide mediates a novel pathway of D2S-induced ERK1/2 activation, leading to cell growth. phospholipase C; G protein; mitogen-activated protein kinase

Research has been conducted on GM1 ganglioside. The role of GM1 ganglioside's ceramide moiety in its association with detergent resistant membrane domains in human leukemia HL-60 cells has been investigated, and the results are reported.

The physiologic role of leptin in the liporegulation is reviewd. Research shows that leptin functions as a liporegulatory hormone and maintains intracellular homeostasis under conditions of varied calorie intake. Data indicate that leptin facilitates surplus fat deposition in adipocytes, suggesting that in diet-induced obesity, leptin resistance may be due to increased leptin resistance factors.