[The role of sphingolipids in pathogenesis of amyotrophic lateral sclerosis]. / Rol' sfingolipidov v patogeneze bokovogo amiotroficheskogo skleroza.

Amyotrophic lateral sclerosis (ALS) is an incurable neurodegenerative disease characterized by selective degeneration of motor neurons of the spinal cord and motor cortex and brain stem. The key features of the course of this disease are excitotoxicity, oxidative stress, mitochondrial dysfunction, neuro-inflammatory and immune reactions. Recently, the mechanisms of programmed cell death (apoptosis), which may be responsible for the degeneration of motor neurons in this disease, have been intensively studied. In this regard, sphingolipids, which are the most important sources of secondary messengers that transmit cell proliferation, differentiation and apoptosis signals, and are involved in the development of neuroinflammatory and immune responses, are of particular interest in the context of ALS pathogenesis. The review provides information from domestic and foreign authors on the involvement of various sphingolipids (sphingomyelins, ceramides, sphingosine, sphinganin, sphingosine-1-phosphate, galactosylceramides, glucosylceramides, gangliosides) in the development of pro-inflammatory reactions and apoptosis of motor neurons in ALS. The authors discuss the prospects of using new drugs that control the metabolism of sphingolipids for the treatment of ALS.

[The role of lipids in the pathogenesis of lateral amyotrophic sclerosis]. / Rol' lipidov v patogeneze bokovogo amiotroficheskogo skleroza.

Amyotrophic lateral sclerosis (ALS) is an incurable neurodegenerative disease characterized by selective degeneration of motor neurons of the motor cortex, brain stem and brain stem. Mutations in genes coding for SOD1, C9ORF72, TDP-43, FUS and others are associated with ALS and result in abnormal processing and transport of RNA as well as changes in the dynamics of cytoskeleton. In addition, a sharp change in the metabolism of various lipid classes, including phospholipids, fatty acids, sphingolipids, etc., was detected. This review describes changes in lipid content and activity of enzymes involved in their metabolism in ALS animal models as well as in patients. Changes in the metabolism of fatty acids, phospholipids, cholesterol and its derivatives are reviewed in detail. The prospects of searching for new drugs among modulators of lipid metabolism enzymes are discussed.

Changes in the Metabolism of Sphingoid Bases in the Brain and Spinal Cord of Transgenic FUS(1-359) Mice, a Model of Amyotrophic Lateral Sclerosis.

The aim of this study was to evaluate changes in the content of sphingoid bases - sphingosine (SPH), sphinganine, and sphingosine-1-phosphate (SPH-1-P) - and in expression of genes encoding enzymes involved in their metabolism in the brain structures (hippocampus, cortex, and cerebellum) and spinal cord of transgenic FUS(1-359) mice. FUS(1-359) mice are characterized by motor impairments and can be used as a model of amyotrophic lateral sclerosis (ALS). Lipids from the mouse brain structures and spinal cord after 2, 3, and 4 months of disease development were analyzed by chromatography/mass spectrometry, while changes in the expression of the SPHK1, SPHK2, SGPP2, SGPL1, ASAH1, and ASAH2 genes were assayed using RNA sequencing. The levels of SPH and sphinganine (i.e., sphingoid bases with pronounced pro-apoptotic properties) were dramatically increased in the spinal cord at the terminal stage of the disease. The ratio of the anti-apoptotic SPH-1-P to SPH and sphinganine sharply reduced, indicating massive apoptosis of spinal cord cells. Significant changes in the content of SPH and SPH-1-P and in the expression of genes related to their metabolism were found at the terminal ALS stage in the spinal cord. Expression of the SGPL gene (SPH-1-P lyase) was strongly activated, while expression of the SGPP2 (SPH-1-P phosphatase) gene was reduced. Elucidation of mechanisms for the regulation of sphingolipid metabolism in ALS will help to identify molecular targets for the new-generation drugs.

[Participation of Sphingolipids in the Pathogenesis of Atherosclerosis].

Lipid metabolism disorders are the most significant risk factor of development of cardiovascular diseases (CVD). In the process of diagnosing ischemic heart disease and other cardiovascular pathologies, levels of total cholesterol, low- and high- density lipoprotein cholesterol, triglycerides are determined. However, in recent years, close attention has been paid to the intersection of the metabolic pathways of the biosynthesis of cholesterol and sphingolipids. Sphingolipids - a group of lipids, which include a molecule of aliphatic alcohol sphingosine. This group includes sphingomyelins, cerebrosides, gangliosides and ceramides, sphingosines and sphingosine-1-phosphate (S-1-P). Ceramides and sphingosines have pro-apoptotic properties, and S-1-P protects cells from apoptosis. Particular attention as inducer CVD attracts ceramide. It has been established that aggregated lipoproteins isolated from atherosclerotic zones are enriched with ceramides. The level of ceramide and sphingosine increases with ischemia/reperfusion of the heart, in the infarction zone and in the blood, and also in hypertensive disease. S-1-P has a pronounced cardioprotective properties. Its content sharply decreases with ischemia and myocardial infarction. S-1-P performs a special function in the structure of high-density lipoproteins (HDL), being one of the main lipid components of these lipoproteins, which determines their multiple functions. Recently, work has been underway to create drugs that can correct the metabolism of S-1-P. The most successful drugs are those that use the S-1-P receptor as a target, since all of its actions are carried out through receptors. Increasing ceramide and sphingosine and reducing blood plasma level of S-1-P can be an important factor in the development of atherosclerosis. It is proposed to use the determination of the level of sphingolipids in blood plasma for early diagnosis of cardiac ischemia and in arterial hypertension. Chromatography-mass spectrometry has been suggested as the main method for testing these lipids.

[Dimebon correction of changes in phospholipid composition induced by tumor necrosis factor-alpha in experement]. / Korrektsiia dimebonom narushenii sostava fosfolipidov v strukturakh mozga, vyzvannykh faktorom nekroza opukholi α v éksperimente.

AIM: To investigate the ability of the neuroprotector dimebon to prevent alterations in brain lipid metabolism caused byTNF-α. MATERIAL AND METHODS: The ability of dimebon (2,8-Dimethyl-5-[2-(6-methyl-3-pyridinyl)ethyl]-2,3,4,5-tetrahydro-1H-pyrido[4,3-b]indole hydrochloride) to prevent alterations in brain lipid metabolism caused byTNF-α was studied in 65 male mice (20+2g weight). TNF-α (10 mkg/mouse), dimebon (0.2 mg/kg) and their combination were injected intraperitoneally. Thirty min, 2, 4 and 24 h after injection, lipid level alterations in total fractions and molecular species of phospholipids (phosphatidylcholine, lysophosphatidylcholine, sphingomyelin and phosphatidylethanolamine) were measured with mass-spectrometry in the hippocampus, cortex and cerebellum. RESULTS AND CONCLUSION: After injection of TNF-α into mice, there are significant changes in the level of all tested phospholipids. Dimebon at a dose of 0.2 mg/kg alone does not cause any changes in the content of all tested phospholipids, but injected together with TNF-α prevents cytokine induced alterations in the lipid content. The selectivity of TNF-α and dimebon influence on certain molecular species of various phospholipids in different parts of mouse brain is found. The presented data suggest protective properties of dimebon preventing the development of proinflammatory syndrome induced by TNF-α in the animal brain.

[The role of sphingolipids in cardiovascular pathologies]. / Rol' sfingolipidov v serdechno-sosudistykh patologiiakh.

Cardiovascular diseases (CVD) remain the leading cause of death in industrialized countries. One of the most significant risk factors for atherosclerosis is hypercholesterolemia. Its diagnostics is based on routine lipid profile analysis, including the determination of total cholesterol, low and high density lipoprotein cholesterol, and triglycerides. However in recent years, much attention has been paid to the crosstalk between the metabolic pathways of the cholesterol and sphingolipids biosynthesis. Sphingolipids are a group of lipids, containing a molecule of aliphatic alcohol sphingosine. These include sphingomyelins, cerebrosides, gangliosides and ceramides, sphingosines, and sphingosine-1-phosphate (S-1-P). It has been found that catabolism of sphingolipids is associated with catabolism of cholesterol. However, the exact mechanism of this interaction is still unknown. Particular attention as CVD inducer attracts ceramide (Cer). Lipoprotein aggregates isolated from atherosclerotic pluques are enriched with Cer. The level of Cer and sphingosine increases after ischemia reperfusion of the heart, in the infarction zone and in the blood, and also in hypertension. S-1-P exhibits pronounced cardioprotective properties. Its content sharply decreases with ischemia and myocardial infarction. S-1-P presents predominantly in HDL, and influences their multiple functions. Increased levels of Cer and sphingosine and decreased levels of S-1-P formed in the course of coronary heart disease can be an important factor in the development of atherosclerosis. It is proposed to use determination of sphingolipids in blood plasma as markers for early diagnosis of cardiac ischemia and for hypertension in humans. There are intensive studies aimed at correction of metabolism S-1-P. The most successful drugs are those that use S-1-P receptors as a targets, since all of its actions are receptor-mediated.

[Tumor necrosis faсtor-alpha - potential target for neuroprotector dimebon]. / Faktor nekroza opukholei-al'fa - potentsial'naia mishen' dlia neiroprotektora dimebona.

Dimebon (Dimebolin) is an antihistamine drug which has been used in Russia since 1983. Recently Dimebolin has attracted renewed interest after being shown to have positive effects on persons suffering from Alzheimer's disease. Animal studies have shown that dimebon acts through multiple mechanisms, both blocking the action of neurotoxic beta-amyloid peptides and inhibiting L-type calcium channels, modulating the action of AMPA and NMDA glutamate receptors. Our experiments with cell culture L929 and mice have shown that dimebon may exert its neuroprotective effect by blocking cytotoxic signals induced by proinflammatory cytokines such as TNF-a which are believed to play a central role in Alzheimer's disease. Dimebon (10 mg/ml) protected mouse fibroblasts L929 against the toxic action of TNF-a. Our study included 65 male mice. TNF-a (10 mg per mouse), dimebon (0,2 mg/kg) and their combination were injected intraperitonealy. Changes in the level of molecular species of sphingomyelin and galactosyl ceramide in hippocampus, cerebellum and cerebral cortex within 30 min, 2 h, 4 h, and 24 h after injection were detected by chromato-mass-spectrometry. Maximal changes in sphingomyelin and galactosyl ceramides contents of different molecular species after single TNF-a administration were found in the hippocampus, and were less expressed in the cerebral cortex and cerebellum after 24 h. Dimebon itself did not induce changes in the sphingolipid spectrum in brain sections, but protected them against disorders induced by TNF-a in the brain. Modern strategies in the search of new therapeutic approaches are based on the multitarget properties of new drugs. According to our results TNF-a may serve as a new target for dimebon.

Interaction of the nitric oxide signaling system with the sphingomyelin cycle and peroxidation on transmission of toxic signal of tumor necrosis factor-α in ischemia-reperfusion.

This review discusses the functional role of nitric oxide in ischemia-reperfusion injury and mechanisms of signal transduction of apoptosis, which accompanies ischemic damage to organs and tissues. On induction of apoptosis an interaction is observed of the nitric oxide signaling system with the sphingomyelin cycle, which is a source of a proapoptotic agent ceramide. Evidence is presented of an interaction of the sphingomyelin cycle enzymes and ceramide with nitric oxide and enzymes synthesizing nitric oxide. The role of a proinflammatory cytokine TNF-α in apoptosis and ischemia-reperfusion and mechanisms of its cytotoxic action, which involve nitric oxide, the sphingomyelin cycle, and lipid peroxidation are discussed. A comprehensive study of these signaling systems provides insight into the molecular mechanism of apoptosis during ischemia and allows us to consider new approaches for treatment of diseases associated with the activation of apoptosis.

Relationship between parameters of lipid peroxidation during obstructive jaundice and after bile flow restoration.

Restoration of bile flow after 9-day cholestasis in rat liver normalized the content of lipid peroxidation products. The removal of the cholestatic factor after 12-day cholestasis was not followed by recovery of these parameters. We showed that measurement of serum concentration of lipid peroxidation products in patients with cholelithiasis during the preoperative period holds promise for selection of the optimum time for surgical treatment and prediction of the risk of postoperative complications.

The relation between sphingomyelinase activity, lipid peroxide oxidation and NO-releasing in mice liver and brain.

We used animal models to study connection between oxidating system and sphingomyelin signaling cascade, because this models are more close related to people disease. Activation of n-sphingomyelinase (n-SMase) in mice liver and brain is coincided in time with increased level of peroxide products (conjugated dienes) after injection of tumor necrosis factor alpha (TNF-alpha). We found that ceramide can induce peroxide oxidation and lead to accumulation of TNF-alpha in animal organs. Nitric oxide (NO) donors (S-nitrosoglutathione and dinitrosyl iron complex) reversibly inhibited activity of n-SMase and decreased level of lipid peroxidation products. This data proposed that both SMase and messengers of oxidative systems could be targets for NO-derived oxidants.

Connection of lipid peroxide oxidation with the sphingomyelin pathway in the development of Alzheimer's disease.

Alzheimer's disease (AD) is characterized by progressive decline in cognition, memory and intellect. It has been hypothesized that amyloid-beta peptide (A-beta) may have a prominent role in neurodegeneration. Oxidative mechanisms have been implicated in this pathway. There is substantial evidence that inflammatory mechanisms, induced by tumour necrosis factor alpha (TNF-alpha), are also involved in AD. TNF-alpha activates receptors linked to multiple effector systems, including a sphingomyelin pathway and peroxide oxidation. We have determined the changes of neutral sphingomyelinase activity, sphingomyelin and ceramide contents, and the level of lipid peroxide products (conjugated dienes), in the cerebral cortex, hippocampus and cerebellum of rats within 3 h and 7 days of intracerebral injection of A-beta and TNF-alpha. A single injection of A-beta and TNF-alpha has been shown to increase the level of peroxide products in the hippocampus and cerebral cortex within 3 h and 7 days. Sphingomyelinase activity and ceramide levels have been found to increase 7 days after A-beta administration. We found that activation of the sphingomyelin pathway lies downstream from the oxidative stress.

Synthesis of lipids in isolated nuclei from rat thymus and liver cells.

Synthesis of lipids was studied in isolated nuclei from rat thymus and liver cells. On incubation of the isolated nuclei with [2-14C]acetate and [1-14C]glycerol, the label was intensively incorporated into phospholipids and with a significantly lower intensity into fatty acids and cholesterol. Only trace amounts of radioactivity were detected in the lipids of chromatin prepared from isolated thymus nuclei after their incubation, and this suggested that lipids were mainly synthesized on the nuclear membrane. On the preincubation of thymus tissue homogenate with [2-14C]acetate and the subsequent isolation of the nuclei and chromatin, the radioactivity of chromatin lipids was comparable to the radioactivity of nuclear lipids. The findings suggested that in the isolated nuclei the newly synthesized lipids were not transported into chromatin from the nuclear membrane. The specific radioactivities of individual phospholipids and fatty acids were different in the isolated nuclei and in nuclei obtained from preincubated homogenate. Mechanisms of lipid synthesis in isolated nuclei and causes of the different radioactivities of lipids in the isolated nuclei and in the nuclei obtained from the preincubated homogenate are discussed.

Functional role of phospholipids in the nuclear events.

This review presents the structural and functional role of phospholipids in chromatin and nuclear matrix as well as the difference in composition and turnover compared to those present in the nuclear membrane. Nuclei have a very active lipid metabolism which seems to play an important role in the transduction of the signals to the genome in response to agonists acting at the plasma membrane level. The evidence on the presence of phospholipid-calcium-dependent protein kinase C (PKC) in nuclei and enzymes of phospholipids turnover is given. Protein kinase C interacts with nuclear phosphoinositol and sphingomyelin cycles products. This fact evidences about possibility that signal transduction events could also occur at the nuclear level during induction of cell proliferation, differentiation and apoptosis.

Role of tumor necrosis factor alpha and sphingomyelin cycle activation in the induction of apoptosis by ischemia/reperfusion of the liver.

The signal transduction pathways triggering apoptotic mechanisms after ischemia/reperfusion may involve TNF-alpha secretion, ceramide generation, and initiation of lipid peroxidation. In the present study involvement of the TNF-alpha, sphingomyelin cycle, and lipid peroxidation in the initiation of apoptosis induced in liver cells by ischemia and reperfusion was investigated. Wistar rats were subjected to total liver ischemia (for 15, 30 min, and 1 h) followed by subsequent reperfusion. Ischemia caused sharp decrease of neutral sphingomyelinase activity. Activity of acidic sphingomyelinase initially decreased (during 15-30 min ischemia) but then increased (after 1 h of ischemic injury). Reperfusion of the ischemic lobe of the liver caused increase in neutral sphingomyelinase activity and decrease in acidic sphingomyelinase activity. A small amount of TNF-alpha detected by immunoblotting analysis was accumulated in the ischemic area of liver rapidly and the content of this cytokine dramatically increased after the reperfusion. TNF-alpha is known to induce free radical production. We found that the accumulation of TNF and increase of sphingomyelinase activity during the development of ischemic/reperfusion injury coincided with increase in content of lipid peroxidation products (conjugated dienes) and DNA degradation detected by gel electrophoresis. Recently it was shown that superoxide radicals are used as signaling molecules within the sphingomyelin pathway. This suggests the existence of cross-talk between the oxidation system and the sphingomyelin cycle in cells, which may have important implications for the initial phase and subsequent development of post-ischemic injury.

Effect of bilirubin on lipid peroxidation, sphingomyelinase activity, and apoptosis induced by sphingosine and UV irradiation.

The effect of bilirubin (BR) on sphingomyelin cycle activity, lipid peroxidation (LPO), and apoptosis induced by sphingosine and UV irradiation has been studied in vivo. Neutral Mg(2+)-dependent sphingomyelinase (SMase) activity and LPO level were monitored in heart, kidney, and liver of mice after administration of BR. BR inhibited both LPO and SMase activities in heart and kidney. BR induced a mild increase in LPO level and moderate increase in lipid contents in liver, consistent with the functional role of liver in both BR and lipid metabolism. BR injected to mice causes simultaneous and unidirectional alterations in both LPO level and SMase activity with a significant (p < 0.05) positive linear correlation between these two parameters. Sphingosine administration results in increased lipid peroxidation in murine liver. Data on DNA fragmentation indicate that exogenous BR may effectively protect thymus cells against sphingosine- and UV-mediated apoptosis. These results have revealed a biochemical association between oxidative stress and BR on one hand and the sphingomyelin cycle and apoptotic cell death on the other hand. Our data show that BR as an antioxidant, due to its effect on the sphingomyelin cycle, can protect membrane lipids against peroxidation and cells against apoptosis induced by various factors.

Effects of reduced and oxidized glutathione on sphingomyelinase activity and contents of sphingomyelin and lipid peroxidation products in murine liver.

Like the phosphatidyl inositol cycle, the sphingomyelin cycle produces a series of the secondary messengers transmitting extracellular signals from the cytoplasmic membrane into the nucleus. Sphingomyelin, ceramide, sphingosine, sphingomyelinase, and ceramidase are the main components of the sphingomyelin cycle. In spite of numerous data on the functional properties of sphingomyelin cycle products, the activation mechanism for the key enzyme of the sphingomyelin cycle, sphingomyelinase (SMase), is not well understood. We have discovered effects of both reduced (GSH) and oxidized (GSSG) glutathione on the activity of neutral SMase in animals. GSH administration (18 mg per mouse) inhibits this enzymatic activity in liver for 2 h and increases the sphingomyelin level exactly as occurs in cell culture. The levels of diene conjugates and ketodienes decrease simultaneously during the experiment, thus indicating the ability of GSH to suppress oxidative processes in the cell. GSSG administration (18 mg per mouse) has no effect on the SMase activity during the first 15 min, but increases it twofold after 1 h. A short-term decrease in this activity after 30 min may depend on the conversion of excess GSSG into its reduced form by glutathione reductase. Unlike GSH, GSSG has no effect on the level of ketodienes after 1 h, but it induces the accumulation of diene conjugates. A strong correlation exists between the changes in SMase activity and in the level of oxidation products caused by either GSH or GSSG. These data indicate a relationship between SMase activity and the level of peroxidation products and possibly a relation between two signaling systems: the sphingomyelin cycle and the oxidative system.

Sphingolipids of transplantable rat nephroma-RA.

Contents of sphingolipids (ceramide, sphingomyelin, gangliosides) and the composition of their sphingoid bases were studied in the transplantable rat nephroma-RA and in rat kidneys. The content of sphingomyelin was about 1.3-fold decreased and the content of ceramide was about 1.4-fold increased in the nephroma compared to normal kidneys, and this correlated with a 1.4-fold increased activity of neutral sphingomyelinase; however, the activity of the acidic isoform of the enzyme was virtually unchanged. The content of gangliosides was also increased in the nephroma. Ceramide and sphingomyelin of the nephroma, in addition to sphingosine, contained a significant amount of sphinganine, although a considerable amount of the latter was also found in the renal ceramide. The ratio sphingosine/sphinganine in sphingomyelins changed from 65:1 in kidneys to 5:1 in the nephroma. Thus, the biosynthesis of sphingoid bases seems to be disturbed in the transplantable rat nephroma-RA compared to normal kidneys.

The role of sphingomyelin cycle metabolites in transduction of signals of cell proliferation, differentiation and death.

Sphingomyelin cycle metabolites ceramide, sphingosine and sphingosine 1-phosphate play an important role in cell proliferation, differentiation, reception, oncogenesis and apoptosis. Ceramide is an intracellular second messenger for apoptosis activating proteases and specific phosphatases. Sphingosine is an endogenous inhibitor of protein kinase C and has an inhibitory effect on many cell functions depending on the activity of this enzyme. On the other hand, sphingosine can activate other kinases depending on the concentration, cell type and nature of a stimulus and release Ca2+ from intracellular stores thereby regulating cell proliferation. Sphingosine induces apoptosis and its level is increased in cells as a result of action of apoptotic inducers. A phosphorylated product of sphingosine, sphingosine 1-phosphate, mediates the mitogenic signal, induces Ca2+ mobilization and protects cells from apoptosis resulting from elevation of ceramide. The quantitative levels of sphingomyelin metabolites in the cell determine the dynamic balance between the apoptotic and mitogenic signals.

Role of endogenous TNF-alpha and sphingosine in induced DNA synthesis in regenerating rat liver after partial hepatectomy.

Cytokine-stimulated metabolism of sphingomyelin results in the accumulation of ceramide and sphingosine which play a part in the regulation of cell proliferation, differentiation, and reception, as well as in oncogenesis. Formation of TNF-alpha (a member of the cytokine family), accumulation of sphingosine, and DNA synthesis (measured by immunoblotting, HPLC, and [3H]thymidine incorporation, respectively) were studied in rat liver after partial hepatectomy. The content of TNF-alpha was found to increase during 12 h following hepatectomy. The maximum of sphingomyelinase activity and accumulation of sphingosine precede the maximum of DNA synthesis. Sphingosine is known to inhibit protein kinase C. On the other hand, it stimulates the metabolism of phosphatidylinositol, thus causing accumulation of diacylglycerol and inositol-1,4,5-triphosphate, which in turn activate protein kinase C. Hence, the release of TNF-alpha in regenerating liver may modulate DNA synthesis through the accumulation of sphingosine which is involved in regulation of protein kinase C activity and of phosphatidylinositol turnover.