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P Viani

Publications and source records attributed to P Viani.

At least 19 recordsLinked to original sources
PubMed2001

GM3 ganglioside inhibits endothelin-1-mediated signal transduction in C6 glioma cells.

We found that sparse and confluent C6 glioma cells differ both in GM3 content, which increases with cell density, and in endothelin-1 (ET-1)-induced phosphoinositide hydrolysis, which was markedly higher in the sparse cells than in the confluent. Also after manipulation of the cellular GM3 content through treatment with exogenous GM3 or with drugs known to affect GM3 metabolism, the ET-1 effect was inversely related to GM3 cellular levels. Cell treatment with an anti-GM3 mAb resulted in the enhancement of ET-1-induced phospholipase C activation and restored the capacity of GM3-treated cells to respond to ET-1. These findings suggest that the GM3 ganglioside represents a physiological modulator of ET-1 signaling in glial cells.

Animals↗
PubMed2001

Nitric oxide production in living neurons is modulated by sphingosine: a fluorescence microscopy study.

An investigation was carried out into the possible effect of sphingosine (Sph) on nitric oxide (NO) production in living neurons. Differentiated granule cells were used in a dynamic videoimaging analysis of single cells labeled, simultaneously, with FURA-2 and the NO indicator 4,5-diaminofluorescein. The results demonstrate that Sph exerts a potent inhibitory effect on the Ca2+-dependent production of NO, without modifying the [Ca2+]i. The effect appears to be specific as neither ceramide nor Sph-1-phosphate had any effect on the NO and [Ca2+]i levels. The data demonstrate that Ca2+-dependent NO production is a specific Sph target in living granule cells, suggesting that this bioactive sphingoid plays a relevant role in neuronal NO signaling.

Animals↗
PubMed2001

Basic fibroblast growth factor-induced proliferation of primary astrocytes. evidence for the involvement of sphingomyelin biosynthesis.

We recently reported that the marked decrease in cellular ceramide in primary astrocytes is an early event associated with the mitogenic activity of basic fibroblast growth factor (bFGF) (Riboni, L., Viani, P., Bassi, R., Stabieini, A., and Tettamanti, G. (2000) GLIA 32, 137-145). Here we show that a rapid activation of sphingomyelin biosynthesis appears to be the major mechanism responsible for the fall in ceramide levels induced by bFGF. When quiescent astrocytes were treated with bFGF, an increased amount of newly synthesized ceramide (from either l-[(3)H]serine or [(3)H]sphingosine) was directed toward the biosynthesis of sphingomyelin. Conversely, bFGF did not appear to affect ceramide levels by other metabolic pathways involved in ceramide turnover such as sphingomyelin degradation and ceramide biosynthesis, degradation, and glucosylation. Enzymatic studies demonstrating a relevant and rapid increase in sphingomyelin synthase activity after bFGF treatment have provided a convincing explanation for the activation of sphingomyelin biosynthesis. The bFGF-induced increase in sphingomyelin synthase appears to depend on a post-translational activation mechanism. Moreover, in the presence of brefeldin A, the activation of sphingomyelin biosynthesis was abolished, suggesting that the enzyme is located in a compartment other than the Golgi apparatus. Also the phosphatidylcholine-specific phospholipase C inhibitor D609 exerted a potent inhibitory effect on sphingomyelin biosynthesis. Finally, we demonstrate that inhibition of sphingomyelin biosynthesis by brefeldin A or D609 led to a significant inhibition of bFGF-stimulated mitogenesis. All this supports that, in primary astrocytes, the early activation of sphingomyelin synthase is involved in the bFGF signaling pathway leading to proliferation.

Animals↗
PubMed2000

Biomodulatory role of ceramide in basic fibroblast growth factor-induced proliferation of cerebellar astrocytes in primary culture.

To evaluate the role of ceramide in glial growth, primary cultures of quiescent astrocytes from rat cerebellum were stimulated to proliferate by mitogenic doses of basic fibroblast growth factor (bFGF). Parallel to the bFGF mitogenic effect was a marked, and persistent, decrease in cellular ceramide levels. Both in vitro and in culture metabolic studies have led us to exclude both sphingomyelinase and ceramidase involvement in ceramide level variation. Instead, we found evidence of a functional connection between the decrease in ceramide levels and astrocyte proliferation. In fact, cell growth in bFGF-stimulated astrocytes was inhibited by exogenous ceramide and C2-ceramide, maximal inhibition being obtained at a ceramide concentration of 5-10 microM. Under the same conditions, the dihydroderivatives of ceramides were without effect. Following ceramide treatment, the phosphorylation of the MAP kinase isoforms ERK1/2, key components in bFGF-induced cell proliferation, was examined. The administration of antiproliferative doses of ceramide or C2-ceramide, but not of their dihydroderivatives, resulted in a significant inhibition of ERK1/2 activation. In conclusion, our data indicate that the prompt modulation of ceramide levels by bFGF is an early step associated with the signaling pathways responsible for the mitogenic activity of bFGF in astrocytes.

Animals↗
PubMed2000

Cultured granule cells and astrocytes from cerebellum differ in metabolizing sphingosine.

Sphingosine metabolism was studied in primary cultures of differentiated cerebellar granule cells and astrocytes. After a 2-h pulse with [C3-(3)H]sphingosine at different doses (0.1-200 nmol/mg of cell protein), both cell types efficiently incorporated the long chain base; the percentage of cellular [(3)H]sphingosine over total label incorporation was extremely low at sphingosine doses of <10 nmol/mg of cell protein and increased at higher doses. Most of the [(3)H]sphingosine taken up underwent metabolic processing by N-acylation, 1-phosphorylation, and degradation (assessed as (3)H(2)O released in the medium). The metabolic processing of exogenous sphingosine was extremely efficient in both cells, granule cells and astrocytes being able to metabolize, respectively, an amount of sphingosine up to 80- and 300-fold the cellular content of this long chain base in 2 h. At the different doses, the prevailing metabolic route of sphingosine was different. At lower doses and in a wide dose range, the major metabolic fate of sphingosine was N-acylation. With increasing doses, there was first increased sphingosine degradation and then increased levels of sphingosine-1-phosphate. The data demonstrate that, in neurons and astrocytes, the metabolic machinery devoted to sphingosine processing is different, astrocytes possessing an overall higher capacity to synthesize the bioactive compounds ceramide and sphingosine-1-phosphate.

Acylation↗
PubMed1999

Sphingosine inhibits nitric oxide synthase from cerebellar granule cells differentiated in vitro.

The effects of different bioactive sphingoid molecules on NOS activity of differentiated cerebellar granule cells were investigated by measuring the conversion of [3H]arginine to [3H]citrulline. Cytosolic Ca2+-dependent NOS activity was strongly inhibited in a dose-dependent manner by sphingosine in concentrations of 1-40 microM. This inhibition seems to be peculiar to sphingosine in that ceramide, N-acetylsphingosine, sphingosine-1P, sphinganine and tetradecylamine have no effect on the cytosolic enzyme at the considered concentrations, suggesting that it is the bulk of the sphingosine hydrophilic portion that is critical for cytosolic NOS inhibition. This inhibition of cytosolic NOS is not reversed by increasing the arginine concentration, so a competitive mechanism can be excluded. Instead, increasing the concentrations of calmodulin led to loss of sphingosine inhibition, suggesting that sphingosine interferes with the calmodulin-dependent activation of the enzyme by a competitive mechanism. Sphingosine and related compounds had no effect on the particulate Ca2+-independent NOS activity. The data obtained suggest that sphingosine could be involved in the regulation of NO production in neurons.

Animals↗
PubMed1999

Predominance of the acylation route in the metabolic processing of exogenous sphingosine in neural and extraneural cells in culture.

The metabolic fate of exogenous [3H]sphingosine was investigated in five types of cultured cells: primary cultures of neurons and astrocytes, murine and human neuroblastoma cells and human skin fibroblasts. After administration of 40 nM [3-3H]sphingosine into a cell-conditioned medium containing fetal calf serum, all cell types rapidly and efficiently incorporated the long-chain base in a time-dependent fashion. In all cases, after a 120 min pulse, the amount of radioactivity taken up was in the range of the endogenous sphingosine content. However, unchanged [3H]sphingosine represented only a very minor portion of the label incorporated into cells throughout the pulse period (10-120 min), indicating rapid and efficient sphingosine metabolism in these cells. Most of the [3H]sphingosine taken up was metabolically processed, either by degradation (assessed as 3H2O release into the culture medium) or by N-acylation (mainly to radioactive ceramide, sphingomyelin, neutral glycolipids and gangliosides). [3H]Sphingosine 1-phosphate accounted for less than 2% of the total radioactivity incorporated in all cases. Throughout the pulse period and in all cell types, 3H-labelled organic metabolites largely prevailed over 3H2O, indicating that N-acylation is the major metabolic fate of sphingosine in these cells under apparently physiological conditions. These results are consistent with the notion that sphingosine has a rapid turnover in the cells studied, and indicate that regulation of the basal level of this bioactive molecule occurs mainly through N-acylation.

Acylation↗
PubMed1999

A fluorescence method for the determination of plasma susceptibility to lipid peroxidation.

OBJECTIVE: We propose a fluorescence kinetics method for monitoring plasma susceptibility to peroxidation. DESIGN AND METHOD: Plasmatic peroxidation was induced by CuSO4 (500 microM), and fluorescence was measured every 30 min. Kinetics were represented by a sigmoidal curve from which it was possible to calculate the latency time (lag-time) and the propagation velocity (slope) of plasma peroxidation. RESULTS: The lag-time monitored by the fluorescence kinetics method corresponded to the formation of thiobarbituric acid reactive substances, and to progressive depletion of polyunsaturated fatty acids and alpha-tocopherol. The mechanism of reaction appeared to be dependent upon plasmatic hydroperoxides, and independent of oxygen radicals. Plasma storage is possible for at least two months at -80 degrees C, and reproducibility of the method is very good. CONCLUSIONS: Fluorescence kinetics provide a highly comprehensive picture of plasma susceptibility to peroxidation in comparison with the conventional measurements of anti- and pro-oxidant ratios.

Copper↗
PubMed1998

Metabolic fate of exogenous sphingosine in neuroblastoma neuro2A cells. Dose-dependence and biological effects.

The possible relationship between metabolism and biological effects of sphingosine was investigated in Neuro2a cells. [C3-3H]-sphingosine, administered at different doses (80 pmol-80 nmol/mg cell protein). Amounts up to hundredfold were rapidly taken up and metabolized, the intracellular content of sphingosine being processed within 2 h. At low doses, [3H]-sphingosine represented a minor portion of the cellular radiolabel, and N-acylated metabolites, particularly ceramide, prevailed over degradation products. Neuro2a cell differentiation took place in conjunction with ceramide increase. At increasing exogenous sphingosine/cell ratio, the acylation process became saturated while sphingosine degradation increased proportionally. From this point on [3H]-sphingosine accumulated and cell toxicity occurred. In conclusion, in Neuro2a cells the biological effects exerted by exogenous sphingosine are strictly connected to the exogenous sphingosine/cell ratio and to the capacity of the cell to metabolize sphingosine.

Animals↗
PubMed1998

The effects of exogenous sphingosine on Neuro2a cells are strictly related to the overall capacity of cells to metabolize sphingosine.

Neuro2a cells were exposed to different doses (1-40 nmol/10(6) cells) of [C3-3H]sphingosine and the relationship between metabolism and biological effects of sphingosine was investigated. Sphingosine appeared to be rapidly taken up and metabolized. The incorporation of sphingosine was not merely dependent on its concentration but primarily on the dose per cell of administered sphingosine. At low doses, [3H]sphingosine represented a minor portion of the cellular radioactivity, and N-acylated metabolites, particularly ceramide, largely prevailed over degradation products. Concomitantly with ceramide increase, Neuro2a differentiation took place. With increasing exogenous sphingosine/doses, the acylation process reached saturation. From this point on, [3H]sphingosine started accumulating and eventually cell toxicity occurred. In conclusion, the biological effects exerted by exogenous sphingosine on Neuro2a cells are not merely dependent on the long-chain base concentration in the culture medium, but are strictly related to the cellular dose of exogenous sphingosine and to the capacity of cells to metabolize sphingosine.

Animals↗
PubMed1997

pH sensitivity and plasma stability of liposomes containing N-stearoylcysteamine.

In this study, we investigated the pH sensitivity of different liposomal formulations containing 10 mol% N-stearoylcysteamine, as pH sensitive molecule. Liposome stability was monitored by determining the release of different entrapped water soluble molecules, 5,6-carboxyfluorescein (CF) being the marker of leakage mainly used. Small unilamellar vesicles composed of egg phosphatidylcholine (EPC) and N-stearoylcysteamine (9:1 molar ratio) incubated at 20 degrees C in citrate phosphate buffer released, at pH 6.8, 2.5 fold the amount of CF released at pH 7.4. The addition of plasma to the incubation medium and an increase of temperature to 37 degrees C led to significantly increased the CF release from EPC/N-stearoylcysteamine SUV, both at pH 7.4 and 6.8. The addition of cholesterol had a stabilizing effect on liposomal vesicles with respect to both temperature and plasma, without affecting pH sensitivity. In fact, at 37 degrees C and in 25% plasma the ternary mixture showed the highest CF release, as a consequence of the moderate acidification of the medium from 7.4 to 6.8. Thus, these liposome formulations are potentially a useful tool for specific drug delivery to pathological tissues such as tumours, inflammation sites and ischemic areas where it is known that a lowering of the pH can occur.

1,2-Dipalmitoylphosphatidylcholine↗
PubMed1997

Behaviour of nitric oxide synthase in rat cerebellar granule cells differentiating in culture.

The possible relation between nitric oxide synthase (NOS) activity and neural differentiation was investigated using primary cultures of rat cerebellar granule cells differentiating in culture. NOS activity was measured in the cytosolic and particulate fractions obtained from cell homogenate. In the experimental conditions used the optimal pH for NOS activity was about 6.4, the activity being about 3-fold higher than at pH 7.4. Cerebellar granule cell differentiation was associated with marked increases in NOS activity. In undifferentiated cells the enzyme was almost evenly distributed between the cytosolic and particulate fractions, during differentiation there was a 12-fold increase in activity in the cytosolic enzyme and a 3-fold increase in the particulate one. This indicates a marked preferential enrichment of the cytosolic enzyme during differentiation. Cerebellar granule cells produced and released NO in the culture medium; NO formation being markedly higher in differentiated cells (7-12 DIC) than in undifferentiated (2-3 DIC) ones. These data demonstrate a relationship between NOS expression and NO production and the differentiation of cerebellar granule cells, supporting the notion that NO may play a role in this process.

Animals↗
PubMed1996

Pyrene lipids as markers of peroxidative processes in different regions of low and high density lipoproteins.

Three different pyrene derivatives, pyrene decanoyl phosphatidylcholine (P10PC), pyrene dodecanoyl sulfatide (P12CS) and cholesteryl pyrenyl hexanoate (P6Chol), were used to follow lipid peroxidation in low and high density lipoproteins. Probe-labelled lipoproteins were subjected to Cu2+ catalyzed peroxidation. In all cases the fluorescence of the probes progressively decreased due to the involvement of pyrene in the peroxidative reaction. Thus, we used the fluorescence decrease of P6Chol to monitor the lipid peroxidation in the hydrophobic core of LDL and HDL, and that of the amphipatic probes, P10PC and P12CS, to follow lipid peroxidation in the envelope of both lipoproteins. The possibility of following lipid peroxidation in individual lipoprotein regions could lead to more detailed information on the oxidative modifications that play an important role in the altered cholesterol homeostasis involved in the formation of atherosclerotic lesions. No differences were observed in the peroxidation kinetics of the hydrophobic core of HDL and LDL monitored with P6Chol. On the contrary kinetics obtained with P10PC and P12 CS demonstrated the HDL envelope to be more susceptible to Cu2+ -dependent lipid peroxidation than that of the LDL. This could be due to a greater radical generating capacity of the HDL envelope and can be explained on the basis of low vitamin E levels and large amounts of polyunsaturated fatty acids esterified on phospholipids determined in HDL, and on literature evidence that indicates HDL as the principal vehicle of circulating plasma lipids peroxides.

Adult↗
PubMed1996

Sphingoid bioregulators in the differentiation of cells of neural origin.

The involvement of ceramide in the differentiation of two neuroblastoma cell lines, Neuro2a and SH-SY5Y, and cerebellar granule cells in primary culture was investigated. The following results were obtained: (a) the cellular content of ceramide markedly increased with induced differentiation of Neuro2a cells (inducers: RA, FCS deprivation), SH-SY5Y cells (inducers: RA, PMA), and spontaneous differentiation of cerebellar granule cells; (b) all the investigated cells in the differentiated form displayed a higher ability to produce ceramide from exogenously administered [3H]Sph-SM and expressed a higher content of neutral sphingomyelinase and, in the case of cerebellar granule cells, also of acidic sphingomyelinase; (c) inhibition of ceramide biosynthesis by Fumonisin B1 blocked the process of differentiation in Neuro2a and cerebellar granule cells; and (d) treatments capable of enhancing ceramide level (administration of sphingosine or C2-Ceramide) induced differentiation in both Neuro2a and SH-SY5Y cells. The data obtained support the notion that ceramide plays a general biomodulatory role in neural cell differentiation.

Animals↗
PubMed1995

Effect of endothelin-1 induced ischemia on peroxidative damage and membrane properties in rat striatum synaptosomes.

Synaptosomes obtained from rat striata lesioned by central injection of endothelin-1 (ET-1) were analyzed for the levels of lipid peroxidation products, the susceptibility to lipid peroxidation, the phospholipid and free fatty acid composition and the activity of Na+,K(+)-ATPase one hour after ET-1 treatment. The intrastriatal injection of ET-1 promoted an increase of endogenous thiobarbituric reactive substances (TBARS), as index of free radical mediated lipid damage, and a greater susceptibility to iron/ascorbate-induced lipid peroxidation. The pattern of free fatty acids showed a significant decrease of arachidonic and docosahexaenoic acid consequent to ET-1 treatment. The analysis of lipid composition showed a significant loss of phospholipids: among phospholipid species, sphingomyelin and phosphatidylethanolamine plasmalogen were particularly reduced by ET-1 treatment. The activity of membrane-bound Na+,K(+)-ATPase was also significantly reduced in synaptosomes obtained from ET-1 lesioned striata. Taken together these results indicate a significant modification of synaptosomal membrane of ET-1 treated rat striata, possibly due to a free radical mediated damage.

Animals↗
PubMed1995

Evaluation of the effect of alcohol consumption on erythrocyte lipids and vitamins in a healthy population.

There is epidemiological evidence that a moderate consumption of alcohol could reduce coronary heart disease. To corroborate this statement and to see how the consumption of red wine affects phospholipid and fatty acid patterns and antioxidant status, a survey was conducted on 58 adult males (20-75 years old). A questionnaire was used to discover their medical history, various constitutional and life-style factors, food habits, and nutrient intake; and the type and amount of alcoholic intake were investigated by means of a questionnaire. Subjects were divided into two groups of nondrinkers and average drinkers (mean 46.5 +/- 4.4, range 30.2-63.4 g/day), and the effect of alcohol was made on the following variables: gamma-glutamyltranspeptidase, apolipoprotein AI, apolipoprotein B, total and high-density lipoprotein-cholesterol, triglycerides, and antioxidant vitamins were determined in plasma; and phospholipids the fatty acids of phosphatidylcholine (PC) and phosphatidylethanolamine total cholesterol, alpha-tocopherol, beta-carotene, and retinol were determined in red blood cells (RBCs). There were no significant differences between the two groups in protein, fat, carbohydrate, retinol equivalent, alpha-tocopherol content, and cholesterol/saturated fat index of their diet. Analysis of the plasma levels of biochemical variables--adjusted for age, smoking (number of cigarettes/day), and body mass index--showed a significant increase of gamma-glutamyltranspeptidase and apolipoprotein AI concentration in the average drinkers compared with nondrinkers. Plasma alpha-tocopherol and retinol levels were also significantly higher in average drinkers. In the latter group, ANCOVA demonstrated a lower percentage of sphingomyelin in RBCs, with a lower sphingomyelin/PC ratio.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗