Effect of lysophosphatidylcholine on α-adrenoreactivity of rat aorta smooth muscles.

In experiments on rat aortic ring segments, lysophosphatidylcholine in concentrations of 2×10(-6), 2×10(-5), and 2×10(-4) M did not suppress the tonotropic effect of phenylephrine (6×10(-6) and 6×10(-5) M) and in concentration of 2×10(-5) M even potentiated it, which was noted for phenylephrine at a concentration of 6×10(-6) M. It was concluded that the chemomodulating effect of lysophosphatidylcholine depends on the type of receptors and cells.

Effects of LDL lipids on activity of group IIA secretory phospholipase A2.

Effects of phosphatidylcholine, oxidized phosphatidylcholine, sphyngomyelin, cholesterol, and cholesterol esters incorporated in LDL on activity of group IIA secretory phospholipase A2 from human cardiac myxoma were studied. Liposomes containing radioisotope-labeled phosphatidylethanolamine served as the substrate for group IIA secretory phospholipase A2. Oxidized phosphatidylcholine significantly stimulated activity of group IIA secretory phospholipase A2, while phosphatidylcholine in the same concentrations did not modify enzyme activity. Sphyngomyelin incorporated in LDL inhibited group IIA secretory phospholipase A2 activity. Cholesterol and cholesterol esters virtually did not modify enzyme activity. The results indicate that LDL phospholipids and their oxidized forms can be involved in regulation of group IIA secretory phospholipase A2. Study of the mechanisms regulating the proinflammatory group IIA secretory phospholipase A2 can promote the development of new approaches to the diagnosis and treatment of inflammatory processes.

Ganglioside GM3 and its biological functions.

Metabolism, topology, and possible mechanisms for regulation of the ganglioside GM3 content in the cell are reviewed. Under consideration are biological functions of GM3, such as involvement in cell differentiation, proliferation, oncogenesis, and apoptosis.

[Effect of atorvastatin therapy on the level of secretory phospholipase A2 group IIA and on the modification of low density lipoproteins in patients with ischemic heart disease].

We studied effect of atorvastatin on secretory phospholipase A2 group IIA (sPLA2-IIA) in blood serum of patients with ischemic heart disease (IHD), lipid composition of low density lipoproteins (LDL) and process of modification of LDL induced by sPLA2-IIA in 20 patients taking 20 mg/day of atorvastatin for 3 months. In patients with initially high level of sPLA2-IIA ( > 8 mcg/l) its concentration significantly decreased. Amount of total cholesterol, triglyceride, lecithin, and lysolecithin remained unchanged, however in equimolar relations there occurred decrease of amount of total cholesterol and increase of cholesterol esters. At incubation of LDL, extracted from patient s plasma before initiation of the study, with human sPLA2-IIA from cardiac myxoma, 3.5 nmol of lysolecithin per 1 mg of LDL protein was formed while at incubation of LDL of same patients, extracted after 3 months of atorvastatin administration, amount of lysolecithin was 1.54 nmol/mg LDL protein. Thus atorvastatin therapy causes lowering of sPLA2-IIA in patients with initially high blood level of the enzyme and to a great extent precludes sPLA2-IIA induced LDL modification.

Characteristics and regulation of ganglioside-induced elevation of free cytoplasmic Ca2+ in human blood platelets.

We have found that gangliosides GD3 and GM3 induced rapid, reversible elevation of free cytoplasmic Ca2+ in fura-2-loaded human blood platelets. The effect persisted in Ca(2+)-free medium, indicating that gangliosides stimulated mobilization of intracellular stores. The action of gangliosides was concentration-dependent with EC50 of about 1 microM. The Ca(2+)-mobilizing effects of gangliosides were potentiated by epinephrine and inhibited by substances inducing activation of protein kinase C and cAMP-dependent protein kinases. Acidic phospholipids partially mimicked the Ca(2+)-mobilizing effects of gangliosides indicating that lipid head charge is essential for this activity. While the elevation of [Ca2+]i produced by arachidonic acid was almost completely blocked by aspirin pretreatment, the effects of gangliosides were diminished only 2-fold, indicating that gangliosides activate both aspirin-sensitive and aspirin-insensitive mechanisms of [Ca2+]i elevation.

Diol lipids. Acylated ethyleneglycol glycosides. A new type of natural glycolipid.

A new type of glycolipid has been detected in ripening corn seeds. The presence of ethyleneglycol, galactose, glucose and fatty acids was shown by degradation studies. The products of alkaline deacylation were identified as galactosyl- and glycosylethyleneglycol by thin-layer chromatography and combined gas-liquid chromatography-mass spectrometry. The native ethyleneglycol galactolipid was isolated by distribution of the total lipids between heptane and 95% methanol, following silica gel column chromatography of the methanol soluble fraction. Analysis of the alkaline deacylation products of the isolated ethyleneglycol lipid as well as examination of the mass-spectrum of its tetraacetate showed the new lipid to have the structure of 1-acyl-2-(O-beta-D-galacto-pyranosyl)ethyleneglycol with palmitic, oleic and stearic acids as the main fatty acid components. The fragmentation patterns under electron impact of trimethylsilyl ethers of synthetic ethyleneglycol glycopyranosides and of 1-palmitoyl-2-(O-beta-D-tetraacetylgalactopyranosyl)ethyleneglycol are described.

Stimulation of platelet adhesion and activation by ganglioside GD3 adsorbed to plastic.

Platelet interaction with gangliosides GD3, GM3, GM1, GD1a and GT1b has been investigated. These gangliosides were previously identified in the vessel wall and ganglioside GD3 was found to accumulate selectively in the intima of atherosclerotic vessels. Gangliosides were adsorbed to plastic and incubated with 51Cr-labeled platelets. The adhesion of gel-filtered platelets to ganglioside GD3 was 3-4 times higher than to other immobilized gangliosides and to albumin-treated plastic. As was shown by scanning electron microscopy, GD3 stimulated intensive spreading of adherent platelets and formation of surface-bound aggregates, while only single unspread platelets were present on the surfaces coated with other gangliosides. GD3 isolated from milk and from human aorta possess the same stimulating activity. Platelet adhesion to GD3 decreased significantly in the presence of the stable prostacyclin analogue, carbacyclin.

Interaction of low-density lipoproteins with gangliosides.

The ganglioside uptake capacity of human serum low-density lipoproteins (LDL), the mode of ganglioside-LDL binding, and the influence of gangliosides on the floatation properties, size distribution, stability and fluorescence of LDL were investigated. The data obtained suggest that both hydrophobic and electrostatic forces are involved in formation of ganglioside-LDL complexes, but the former appear to be more important. Although association of gangliosides with LDL is predominantly unspecific, nonsaturable, and weak, a small saturable component due to specific ganglioside-apolipoprotein binding, also appears to be involved. In the presence of gangliosides the lipoprotein particles aggregate, the intrinsic fluorescence of LDL and their interaction with antibodies against apo-B change indicating that the state of apo-B [corrected] is modified by gangliosides.

Incorporation and localisation of ganglioside GM3 in human intimal atherosclerotic lesions.

Immunohistochemical examination showed that sections of intimal atherosclerotic plaques contained cells and cell clusters as well as areas of extracellular matrix specifically stained with antibodies against ganglioside GM3. No immunohistochemical staining was observed in areas bordering the plaques where there was no histological evidence of atherosclerosis. To determine whether the ganglioside GM3 deposits in the intimal plaques derived directly from plasma or were synthesised by intimal cells. intimal plaque and plasma LDL were assayed for ganglioside GM3 fatty acid composition. This assay showed that more than 50% of the fatty acids of GM3 isolated from both atherosclerotic and normal intima are either minor fatty acids or those absent from LDL GM3. We conclude that the GM3 deposits present in intimal plaque arise in intimal cells and do not derive from plasma LDL.

Phenotype determination of anti-GM3 positive cells in atherosclerotic lesions of the human aorta. Hypothetical role of ganglioside GM3 in foam cell formation.

Earlier we reported that atherosclerotic plaques contain cells which were specifically and very intensively stained with anti-GM3 antibodies although no GM3 positive cells were detected in the normal non-diseased arterial intima. Because of their lipid inclusions, GM3 positive cells in atherosclerotic lesions seemed to be foam cells but their origin needed clarification. Using an immunohistochemical technique in the present work, we showed that some of these foam cells contained CD68 antigen. However, the most intense accumulation of GM3 occurred in the areas composed of foam cells which did not stain with any cell type-specific antibodies, including antibodies to macrophages (anti-CD68) and smooth muscle cells (anti-smooth muscle alpha-actin), perhaps, because the cell type-specific antigens were lost during the transformation of intimal cells into foam cells. Ultrastructural analysis of the areas where foam cells overexpressed GM3 demonstrated that some foam cells lacked both a basal membrane and myofilaments but contained a large number of secondary lysosomes and phagolysosomes, morphological features which might indicate their macrophage origin. Other foam cells contained a few myofilaments and fragments of basal membrane around their plasmalemmal membrane, suggesting a smooth muscle cell origin. These observations indicate that accumulation of excessive amounts of GM3 occurs in different cell types transforming into foam cells. We suggest that up-regulation of GM3 synthesis in intimal cells might be an essential event in foam cell formation. Shedding of a large number of membrane-bound microvesicles from the cell surface of foam cells was observed in areas of atherosclerotic lesions corresponding to extracellular GM3 accumulation. We speculate that extracellularly localised GM3 might affect the differentiation and modification of intimal cells in atherosclerotic lesions.

Interaction of ganglioside-containing planar bilayers with serotonin and inorganic cations.

The binding of serotonin and inorganic cations K+, Na+, Ca2+, Mg2+ to planar bilayers formed from mixtures of phosphatidylcholine and mono-, di- and trisialogangliosides was studied by the potentiodynamic and nonactin-induced potassium conductivity method. The theoretical analysis of the results obtained was made taking into account (1) protrusion of the ganglioside charges from the membrane surface and (2) simultaneous adsorption of ions on the bilayer surface and on the ganglioside charges protruding into the solution. It was shown that there was no specific binding of K+ and Na+. The binding constants for Ca2+, Mg2+ were determined. These constants for all the gangliosides studied were equal to 500 M-1. The determined binding constants of serotonin to various gangliosides diminish in the following order: GD3 greater than GT1b greater than GD1a greater than GM1.

Action of influenza virus neuraminidase on gangliosides. Haemagglutinin inhibits viral neuraminidase.

The action of partly purified neuraminidase (NA) of influenza A virus, a mixture of detergent solubilized NA and haemagglutinin (HA) and of intact virions on gangliosides GT1b, GD1a, GD1b, GM1 was studied. The viral NA transformed GT1b mainly into GD1b with formation of only minor amounts of GM1. HA was found to inhibit the hydrolysis activity of viral NA. At the same time viral NA transformed GD1a quantitatively into GM1 which was not hydrolyzed by the enzyme. These results suggest that the function of NA is to transfer the 'primary' receptor (such as GT1b) into the proper carbohydrate sequence (GD1b-like) which is proposed to serve as the minimal structure required for influenza virus reception.

Interaction of prostaglandin E1 with human high density lipoproteins.

Prostaglandin E1 has been shown to interact with serum high density lipoproteins (HDL) in a manner resembling the interaction of a ligand with a high affinity binding site. The presence of 10(-12)-10(-10) M prostaglandin E1 induces a rearrangement of the HDL surface lipids and probably influences the biological functions of the lipoproteins.

Neutral glycosphingolipid content and composition of cells from normal and atherosclerotic human aorta.

We have investigated the content and composition of neutral glycosphingolipids (GSLs) in the cells isolated by enzyme digestion from elastic-hyperplastic and musculo-elastic intimal layers of grossly normal and atherosclerotic regions of human aorta. We have detected three types of neutral GSLs in the intimal cells identified as glucosylceramide, trihexosylceramide and tetrahexosylceramide. We failed to detect lactosylceramide in the intimal cells. The cells of the elastic-hyperplastic layer of grossly normal regions contained trihexosylceramide and tetrahexosylceramide, while glucosylceramide was not detected. Considerable amounts of glucosylceramide were found, and the trihexosylceramide and tetrahexosylceramide content was increased in the cells isolated from atherosclerotic regions. The cells of the musculo-elastic layer of grossly normal intimal regions contained glucosylceramide, trihexosylceramide and tetrahexosylceramide. Cells of the musculo-elastic layer of the fatty streak contained noticeable higher amounts of glucosylceramide, as well as greater amounts of trihexosylceramide and tetrahexosylceramide. Cells of the musculo-elastic layer of the plaque also appeared to contain more glucosylceramide, tetrahexosylceramide, but less trihexosylceramide as compared with grossly normal regions. In both cases cells of the fatty streak exhibited the highest total amount of neutral GSLs, but at the same time the neutral GSL composition of the fatty streak was not similar to GSL composition which is known for human blood monocytes. These findings indicate that elevation of neutral GSL level is observed in cells from atherosclerotic lesions of human aortic intima.

Ganglioside content and composition of cells from normal and atherosclerotic human aorta.

The ganglioside content and composition of cells obtained by enzyme digestion of 2 layers of human aortic intima were investigated. Five gangliosides were identified in cells isolated from the external musculo-elastic intimal layer adjacent to the media: GM3, GM1, GD3, GD1a, and GT1b. The same gangliosides plus ganglioside Gx, the chromatographic mobility of which corresponded to the mobility of ganglioside GD1b from human brain, were found in cells from the internal elastic-hyperplastic intimal layer adjacent to the vessel lumen. In both layers, the major cellular ganglioside was GM3 which represented 60% of the total cellular ganglioside content. The ganglioside content was lower in cells obtained from fatty streaks compared to cells isolated from unaffected intima. The amount of di- and trisialogangliosides in atherosclerotic plaque cells was lower, and that of monosialogangliosides higher than in cells isolated from unaffected intima. The amount of GM3 was mainly responsible for the difference in the total ganglioside content of cells obtained from different lesion types. On the whole, cells from fatty streaks contained smaller amounts of total gangliosides, whereas cells from plaques had greater total ganglioside content, than cells from unaffected intima.

Autoantibodies to gangliosides in sera of atherosclerotic patients.

Using ELISA we studied the levels and clinical correlation of serum antibodies against gangliosides and 5-hydroxytryptamine (5-HT) in patients with atherosclerosis and clinical manifestations of cardiovascular disease. A range of 70-80% of the patients showed higher titers of anti-GM3(L) and anti-5HT as compared to normal serum. The anti-GM3(L) antibodies appeared to be directed mainly against GM3 present in platelets and were much less reactive against GM3 isolated from the aorta. We concluded that the antigens responsible for the elevated anti-GM3(L) and anti 5-HT levels in atherosclerotic sera are released by vessel-wall activated platelets. These results provide further evidence of on-going autoimmune processes in atherosclerosis. The content of total sialic (TS) and lipid-bound sialic acid (LBS) was measured in sera of patients with IHD and of similar numbers of healthy donors. In the patient groups the average TS and LBS concentration was about 25% higher than in the control group. These changes appeared to be associated with higher degrees of protein sialylation and larger amounts of LDL in the patient sera than in those of healthy controls.

Phospholipids and glycosphingolipids in cultured skin fibroblasts from patients with progressive systemic sclerosis (scleroderma).

OBJECTIVE: A comparative investigation of glycosphingolipids and phospholipids of cultured skin fibroblasts from healthy donors and patients with progressive systemic sclerosis (SSc) was performed. METHODS: Culture techniques, qualitative and quantitative lipid analyses, determination of neuraminidase activity, and immunofluorescence microscopy were used. RESULTS: The quantitative content of individual phospholipids in the fibroblasts of SSc patients (PF) was equally elevated, on the average 2.8 times in comparison with the fibroblasts of healthy donors (DF). The total content of neutral glycosphingolipids was slightly elevated only because of a 1.5-fold increase of monoglucosylceramide in PF. A decrease in the amount of the gangliosides GM3 and GM1 and the absence of the disialogangliosides in PF in comparison with DF, were demonstrated. Immunofluorescent assay also showed a decrease of ganglioside epitopes on the cellular surface of PF in comparison with DF. The GM3-neuraminidase activity of PF homogenates was increased two-fold in comparison to normal values. CONCLUSIONS: These results are discussed in connection with abnormalities in membrane receptor functions, alterations in the cell phenotype and AMP-cyclase system activity, as well as hyperproduction of extracellular matrix proteins by PF.

Gangliosides and atherosclerosis.

The ganglioside levels in atherosclerotic lesions of human aorta are considerably higher than those in unaffected areas of aorta, and atherosclerotic patients frequently have increased concentrations of serum gangliosides. The present review summarizes recent findings that suggest the possible involvement of aortic gangliosides in platelet activation and adhesion of platelets to the vessel wall. The effect of gangliosides on the structure of low density lipoproteins (LDL), on the interaction of LDL with macrophages and hepatic cells and on the LDL-regulated biosynthesis of cholesterol is also discussed. In vitro experiments have demonstrated that a major ganglioside of the intima of atherosclerotic aorta induces rapid adhesion, aggregation and spreading of platelets. Moreover, gangliosides present in elevated amounts in the intercellular space of atherosclerotic aortic tissue modify the surface structure and stimulate aggregation of LDL. Ganglioside-modified LDL are readily recognized and taken up by macrophages, while preincubation of LDL with low concentrations of gangliosides inhibits LDL binding to hepatic cells. Thus, ganglioside enrichment of LDL is likely to interfere with LDL clearance via the hepatic cells. Thus, ganglioside enrichment of LDL is likely to interfere with LDL clearance via the hepatic LDL receptor, and to stimulate binding of LDL to the scavenger receptor of macrophages. It is postulated that high ganglioside levels in the aorta and serum may be an additional risk factor in atherosclerosis.

[Neutral glycosphingolipids in Ehrlich ascites carcinoma cells]. / Neitral'nye glikosfingolipidy kletokk astsitnoi kartsinomy Erlikha.

The structure of the neutral glycosphingolipids of the Ehrlich ascite carcinoma (EAC) cells was studied. The main four components were identified as glycosylceramide, lastosylceramide, N-acetylgalactosyllactosylceramide and galactosyl-N-acetyllactosylceramide (asialo-GM1). The neutral glycolipid pattern of the cells was found to depend on their density. Dilution of the cell suspension resulted in an increased content of asia-lo-GM1, whereas the content of the other neutral glycolipids remained unchanged. The possible connection between these changes and the earlier disclosed cell density dependence of the gangliosides in EAC cells is discussed.

[Glycosphingolipids of human aorta]. / Glikosfingolipidy aorty cheloveka.

The structures of the main gangliosides of human aorta (intima and media) were elucidated. The main component (67%) was identified as N-acetylneuraminosyl-lactosylceramide (ganglioside GM3). The aorta tissue contained also gangliosides GM1, GD3, GD1a, and GT1. All sialic acid residues in gangliosides were present as N-acetyl-neuraminosyl derivatives. Among neutral glycosphingolipids of human aorta, the main components were identified as glucosylceramide, lactosylceramide, globotriaosylceramide and globotetraosylceramide. The preliminary data suggest that the composition of the investigated glycosphingolipids in tissue might vary upon atherosclerosis lesions of aorta.