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.

An ultradian oscillation of protein synthesis was detected by synchronization of metabolic activity in rat hepatocyte cultures. This oscillation occurs in dense cultures in fresh medium, but not in sparse ones. Metabolic synchronization of sparse cultures, however, was initiated by conditioned medium or addition of 0.3-0.5 microm of a mixture of bovine brain gangliosides to fresh culture medium along with either 0.06-0.2 microm GM1 or 0.1-0.2 microm GDIa. GTIb and GDIb did not produce oscillations, nor did human liver ganglioside GM3. High expression of GM1 ganglioside determinants in hepatocytes maintained in the conditioned medium purified polyclonal antibodies to GM1 was coupled with protein synthetic oscillatory activity, i.e. metabolic synchronization. Incubation of dense cultures with GM1-antibodies for 24 h decreased the amplitude of these oscillations. In sparse cultures maintained in fresh medium where protein synthesis showed no oscillatory pattern, GM1 expression was low.

The ganglioside composition of Ehrlich ascites carcinoma (EAC) cells and the role of the individual gangliosides in binding and penetration into the cell of influenza virus were determined. EAC gangliosides identical with or close t o G M~. GMZ, G M~, G T~~ and G T I~ were characterized by thin-layer chromarography, compositional analyses, methylation analysis and mass-spectrometry. The ganglioside uptake capacity of native and neuraminidase-treated EAC cells was studied with tritium-labeled gangliosides of definite structure and the binding of influenza virus to cells was determinated by using ["Hluridine-labeled virus and by heinagglutination studies. Treatment of the cells with Vihrio c l t o l~~r n~~ neuraminidase largely decreased binding of the virus. Exogenous gangliosides with a terminal galactose unit or a penultimate galactose masked by neuraminic acid were able to restore the virus-binding capacity of ncuraminidase-treated cells, however, the main ganglioside of EAC cells, Ghfz, which carbohydrate chain is terminated by N-acetylgalactosamine, was completely ineffective. The common carbohydrate sequence of the gangliosides showing binding activityis proposed to be the main recognition structure of the influenza virus receptor on the surface of EAC cells. Penetration of labeled influenza virus into the nuclei of EAC cells was evaluated by measuring the radioactivity of the nuclei of neuraminidase-treated ganglioside-loaded cells after exposition to the labeled virus. Of all gangliosides tested only trisialogangliosidcs of the G~l b type were able to induce increased entry of the virus into the cells and accumulation of its radioactive component into the nuclei. It is suggested that G T I~ gangliosides react specifically with the virus protein responsible for membrane fusion (apparently the hemagglutinin HA2 subunit) and thus are involved in virus penetration and delivery of the virus genome to the nuclei.When interacting with cells ortho-and paramyxoviruses first adsorb to specific host receptors and then penetrate thc cell. While nothing is known about the nature of the cell surface components involved in the penetration process, some information about the receptors responsible for adsorbtion of the viruses is available. The receptors contain neuraminic acid and their receptor activity is destroyed by neuraminidase [2]. It is therefore thought that glycoproteins [3 -51 or sialoglycolipids (gangliosides) [6 -81 serve as receptors.Comparing the virus-binding activity of glycoproteins isolated from HeLa cells and brain gangliosides, Wu et al.", I showed that adsorbtion of Sendai virus required much less glycoprotein than glycolipid and concluded that the former are responsible for virus attachment. However recently it hdS been demonstrated that di-, tri-and tetrasialogangliosides can function as receptors for Sendai virus in cultured bovine kidney cells [lo, 1 11. Thus, the existing evidence of participitation of gangliosides in virus-cell association is contradictive. Moreover, the data obtained so far d...

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.

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The ganglioside composition of mouse ascites hepatoma (MAH) cells, the ascites fluid and cell-conditioned media were determined and found to be qualitatively identical, but quantitatively different.The ganglioside content of the ascites fluid and the medium conditioned by MAH-cells at the native cell concentration (1 O8 cells/ml) comprised respectively 74.9 :d and 23 % of the cell-associated gangliosides. When incubated at lower cell-density (lo6 cells/ml) the cells were found to be release about three-times higher amounts of ganglioside per cell than during incubation at the native concentration. Centrifugation of the dense-cell-conditioned medium revealed the major part of the released gangliosides to be associated with a 150000 x g pellet that probably contains shed plasma membrane fragments. In the 150000 x g pellet of the extracellular fluids the relative content of the most polar cell ganglioside corresponding chromatographically to GT1, was about ten-times higher than in the cells. The possibility is raised that the more intense shedding of gangliosides from less crowded MAH cells may play a role in the self protection of the tumor from host immune rejection during initial stages of growth.Gangliosides repeatedly have been suggested to exert immunosuppressing activity [I -61. Although the major part of the cell gangliosides are believed to reside in the outer leaflet of the plasma membrane certain amounts are found in non-cell associated form in the blood serum, where they may be present as free molecules, micelles, protein-bound complexes or membrane fragments. It has been speculated that serum gangliosides released by tumor cells may participate in the selfprotection of those cells from host immune rejection [I].However up to now only few data on the shedding of gangliosides from tumor cells have been published [7 -91.In the present study we examine the gangliosides secreted by mouse ascites hepatoma (MAH) cells and demonstrate that the amount of gangliosides shed depends specifically on the cell density. A preliminary communication has been published [lo]. MATERIALS AND METHODSwere redistillated prior to use. All chemicals were of analytical reagent grade and solvents CellsMale CBAxC5,BL/6F1 mice of 18-22 g weight were used throughout. The MAH-cells were grown in the peritoneal cavity and maintained for 9 days. The ascites fluid was separated from the cells by centrifugation at 800 x g (3000 rpm) for 3 min. The cell pellet was washed three times with physiological salt solution. For subsequent incubation the cell pellet was suspended in Eagle's complete medium and diluted with physiological salt solution to obtain a final concentration of 17 mg protein/ml or 0.17 mg protein/ml corresponding to lo8 or 106 celIs/mI respectively.Abbreviations. MAH, mouse ascite hepatoma 22a; the designation of gangliosides follows the nomenclature system of Svennerholm as listed in [IS].The intensity of protein synthesis was determined by studying incorporation of [14C]leucine (Amersham; specific activity 330 Ci/mol; acti...

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.

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