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B Cestaro

Publications and source records attributed to B Cestaro.

At least 19 recordsLinked to original sources
PubMed1992

Calcitonin-induced changes in the organization of sulfatide-containing membranes.

The interactions of salmon calcitonin with glycosphingolipid sulfatide are studied by right angle light scattering from the lipid suspension, by the excimer to monomer ratio (E/M) of the fluorescence intensity of pyrene phosphatidylcholine and pyrene sulfatide and by the leakage of carboxyfluorescein. It was found that calcitonin strongly modified the structure of the sulfatide aggregate, as indicated by the light scattering determinations. At a lipid peptide ratio 100:1 (molar ratio) light scattering from the suspension was negligible, indicating the formation of peptide-sulfatide complexes with a structure different from that of the lipid aggregate. The interactions of calcitonin with sulfatide when the latter is a component of a bilayer were also evaluated. A specific calcitonin-membrane sulfatide interaction was demonstrated by determining the temperature-dependent E/M of pyrene phosphatidylcholine and pyrene sulfatide in dipalmitoyl phosphatidylcholine/sulfatide (80:20, molar ratio) liposomes. The E/M curves were modified by calcitonin only when the liposomes were labelled with fluorescent sulfatide which probes the sulfatide behavior in the membrane. Furthermore, the addition of calcitonin to the incubation medium of liposomes containing sulfatide promoted the release of vesicle entrapped carboxyfluorescein without disrupting the bilayer structure, the release being correlated with the amount of sulfatide in the bilayer and the calcitonin concentration in the medium.

1,2-Dipalmitoylphosphatidylcholine↗
PubMed1991

Pyrene derivatives as markers of transbilayer effect of lipid peroxidation on neuronal membranes.

Two different pyrene derivatives, namely 12-(1-pyrene)dodecanoic acid (P12-FA) and N-(12-(1-pyrene)dodecanoyl)-galactosylsphingosine I3-sulfate (P12-CS) have been used to follow lipid peroxidation both in model and natural membranes. The malondialdehyde (MDA) production in small unilamellar vesicles of dipalmitoylphosphatidylcholine/arachidonic acid (80:20, molar ratio), symmetrically labelled with both probes determined a progressive decrease of pyrene fluorescence due to an involvement of pyrene in the peroxidative reaction. Nervous membranes are particularly sensitive to lipid oxidation which differentially acts on the two layers of the membrane determining a greater rigidity of the exofacial one. Thus, we consider the possibility to asymmetrically introduce the pyrene ring, as P12-FA or P12-CS, in synaptosomes for monitoring lipid peroxidation in each layer of the membrane. The amount of the two probes incorporated in the membrane was 20 +/- 3 and 10 +/- 2 nmol/mg of protein for P12-FA and P12-CS, respectively. P12-FA was symmetrically distributed in the two layers, whereas 95% of P12-CS was incorporated in the exofacial layer of the membrane as determined by TNBS measurements. The decrease in fluorescence of synaptosome associated pyrene was, in the early stages of lipid peroxidation, greater for P12-CS than for P12-FA labelled membranes, indicating a greater susceptibility of the exofacial layer to iron-induced peroxidation.

Animals↗
PubMed1991

Age-related differences in synaptosomal peroxidative damage and membrane properties.

Young, adult, and old rats were used to study the effect of age on the integrity and functioning of brain synaptosomes. An evaluation was made of the differences in lipid composition, membrane fluidity, Na+, K(+)-ATPase activity, and susceptibility to in vitro lipid peroxidation. There was an age-related increase in synaptosomal free fatty acids, with no modification in acyl chain composition, and a decrease in membrane phospholipids which increased the cholesterol/phospholipid mole ratio. With altered lipid composition, there was a corresponding age-dependent decrease in membrane fluidity, a reduction of Na+, K(+)-ATPase activity, and an overall greater susceptibility to in vitro lipid peroxidation. Furthermore, lipid peroxidation promoted strong modifications of the membrane fluidity, lipid composition, and Na+,K(+)-ATPase activity just as aging did, thus indicating a possible contribution of oxidative damage to ageing processes. The cases studied revealed that the greater responsiveness of old membranes to in vitro lipid peroxidation resulted in the highest degree of membrane alteration, indicating that all pathological states known to promote a peroxidative injury can have even more dramatic consequences when they take place in old brain.

Aging↗
PubMed1990

Studies on peroxidation processes of model membranes and synaptosomes: role of phosphatidic acid.

Dipalmitoylphosphatidic acid (DPPA) was found to exert a strong inhibitory effect on Fe-induced peroxidation of arachidonic acid inserted into liposomal dipalmitoylphosphatidylcholine (DPPC) vesicles. This inhibition was quite effective both below and above the phase transition temperature of the liposomes. Moreover, we demonstrated the antiperoxidative activity of phosphatidic acid (PA) in synaptosomal membranes. PA enriched synaptosomes were prepared by the stimulation of the endogenous phospholipase D activity or by the incubation of the synaptosomes with Streptomyces chromofuscus phospholipase D. The possible contribution of PA to the in vivo defense mechanism against free radical-induced damage is discussed.

Animals↗
PubMed1990

Studies on peroxidation processes of model membranes: role of pyrophosphate.

Phosphate-containing compounds of both a lipophilic (dipalmitoylphosphatidic acid, lysophosphatidic acid and dicetylphosphate) and hydrophilic nature (orthophosphate, 3-phosphoglyceric acid, 2,3-diphosphoglyceric acid and pyrophosphate in particular) have been found to inhibit to varying degrees the lipoperoxidation of liposomal arachidonic acid. Lipophilic compounds seem to act exclusively at the lipid component level of the membrane, giving rise to polyanionic complexes with free arachidonic acid or its radical derivatives that could bind the Fe2+ strongly (thereby inhibiting the iron pro-oxidation activity) or to minimize the lateral mobility of the lipid radicals (thereby reducing the propagation of lipid peroxidation). The high antiperoxidative power of hydrophilic compounds, and in particular of pyrophosphate, must, on the contrary, be primarily attributed to their ability to form very stable complexes with the Fe present in the solution surrounding the liposomal membranes. The possible contribution of these physiological compounds to the in vivo defense mechanism against radical-induced damage is discussed.

Arachidonic Acid↗
PubMed1989

Synthesis of pyrene derivatives of cerebroside sulfate and their use for determining arylsulfatase A activity.

Two fluorescent derivatives of cerebroside sulfate ('sulfatide') have been synthesized and used as substrates for determining arylsulfatase A activity. These were 12-(1-pyrene)dodecanoyl cerebroside sulfate (P12-sulfatide) and 12(1-pyrenesulfonylamido)dodecanoyl cerebroside sulfate (PSA12-sulfatide). When incubated at pH 5.0 in the presence of 5 mM MnCl2 and 5.5 mM of taurodeoxycholate, either substrate was hydrolyzed by arylsulfatase A of human leukocytes. The rate of hydrolysis was proportional to the incubation time and concentration of enzyme; Michaelis-Menten type kinetics were observed with increasing concentrations of substrate. For determining the rate of hydrolysis, each of the two products (i.e., P12- and PSA12-cerebrosides) were separated from the bulk of respective unreacted sulfatide on small columns of DEAE-Sephadex A-25 and their fluorescence intensities read at 343-378 and 350-380 nm for the excitation and emission wavelengths for P12- and PSA12-cerebrosides, respectively. When extracts of skin fibroblasts derived from normal individuals and patients with Maroteaux-Lamy (lacking arylsulfatase B) or metachromatic leukodystrophy (lacking arylsulfatase A) were used as source of enzyme, P12-sulfatide was hydrolyzed by the former two but not by the latter cell extract. Several derivatives of cerebroside sulfate were also synthesized and found to inhibit the hydrolysis of pyrenesulfatide by leukocyte arylsulfatase A. The results demonstrate that these two pyrene containing sulfatides can be effectively used as specific substrates for the determination of arylsulfatase A activity in extract of cells and most probably also of tissues.

Animals↗
PubMed1989

Correlation of the dispersion state of pyrene cerebroside sulfate and its uptake and degradation by cultured cells.

This study aimed at increasing the efficiency and shortening the time required for administering cerebroside sulfate to cultured cells. For this purpose several modes of dispersion of a fluorescent derivative of cerebroside sulfate (sulfatide), in which the natural fatty acid has been replaced by pyrenedodecanoic acid (P12), were incubated with the cells. This fluorescent derivative of cerebroside sulfate (P12-CS) was introduced into the growth medium of the cells using the three following modes of dispersion: (1) P12-CS was dissolved in dimethylsulfoxide and added to the medium, (2) it was precomplexed with serum albumin or (3) incorporated into small, unilamellar vesicles (SUV) of phosphatidylcholine. With each of these respective modes of dispersion, the P12-CS was incubated for periods up to 48 h with cultured lymphoblasts or fibroblasts. Uptake by the cells could be determined by recording directly the cell-associated fluorescence, using a suspension of washed intact cells. The cell lipids were subsequently extracted with mixtures of chloroform/methanol and their fluorescence recorded. When related to the incubation time, uptake of P12-CS by the cells increased continuously using each of the above dispersions. The appearance of fluorescence at 475 nm ('excimer') and the ratio of this to the monomolecular fluorescence at 378 nm ('E/M') could be used as a measure for the presence of the internalized P12-CS in aggregated or fully dispersed states. These values (i.e., E/M), recorded on the suspensions of intact cells were rather high using the aqueous dispersions, intermediate values were observed using the SUV and rather low E/M values (0.5 or less) were observed using the preformed albumin-(P12-CS) complexes. Increasing the mole ratio of albumin to P12-CS (i.e., from 1:2 to 2:1 m/m), decreased the quantity of sulfatide which was taken up by the cells but also further decreased the E/M ratio, suggesting a fully dispersed state of the pyrene lipid within the cell. This indicated that, using an optimal albumin to P12-CS ratio of 1-2 (or its equivalent values in fetal calf serum) permitted an influx of single molecules of P12-CS into the cells. After 48 h, about 50% of the fluorescence of skin fibroblasts was found in metabolic degradation products of P12-CS. The parallel value for fibroblasts derived from a patient with metachromatic leukodystrophy was only about 5%. Appearance of the excimeric emission of a dispersion of P12-CS in water permitted estimation of its critical micellar concentration as being 7.5 x 10(-7) M.

Cell Line↗
PubMed1988

Studies on peroxidation of arachidonic acid in different liposomes below and above phase transition temperature.

The mechanism of Fe-induced peroxidation of arachidonic acid (AA) in small unilamellar vesicles (SUV) of dipalmitoylphosphatidylcholine (DPPC) and dipalmitoylphosphatidylethanolamine (DPPE) was studied below and above gel to liquid-crystalline phase transition temperature (Tm). In both liposomes the AA peroxidation resulted higher in the temperature range below Tm, but the extent of malonyldialdehyde (MDA) formation was dramatically lower in DPPE vesicles when compared with corresponding DPPC liposomes. A possible explanation for this is discussed.

Arachidonic Acids↗
PubMed1988

N-pyrene dodecanoyl sulfatide as membrane probe: a study of glycolipid dynamic behavior in model membranes.

An N-linked pyrene-dodecanoyl sulfatide was employed to measure the ratio of excimer fluorescence to monomer fluorescence intensities (E/M). The E/M values provided information about both the dynamic behavior and the structural distribution of the labelled glycolipid in note dispersion of micellar sulfatides and multilamellar vesicles of different phospholipids. Most of the labelled sulfatide seems to be located in domains sequestered from the surrounding phospholipids still above the phase transition temperature of the vesicles. The glycolipids sequestered in these domain environments are less sensitive to the structural changes that the addition of cholesterol or Ca2+ can induce in the phospholipid regions during the phase transition.

Fluorescent Dyes↗
PubMed1988

Erythrocyte D-glucose transport activity in reconstituted model membranes of different lipid composition.

The stereospecific influx of D-glucose into liposomes formed on sonication of different glyco- and phospholipids with transport proteins from human erythrocyte ghosts solubilized with Triton x-100 was measured as an index of their total D-glucose transport activity. Specific D-glucose transport increased when acidic phospho- and glycolipids (especially sulfatide) were added to the phosphatidylcholine bilayers of the model membranes while cholesterol strongly inhibited the process. The modulation of D-glucose transport activity and its possible correlation with the lipid composition and the chemico-physical state of the erythrocytes is discussed.

Cerebrosides↗
PubMed1987

Interactions of insulin with sulfatide-containing vesicles of phosphatidylcholine at different pHs.

Positively charged insulin is described to induce aggregation of phosphatidylcholine vesicles containing 10 mol% sulfatide at acidic pH. Techniques including light-scattering, Sepharose chromatography, centrifugation, trapped volume determination, circular dichroism and fluorescence polarization, demonstrate that large amounts of negatively charged insulin remain firmly associated to the vesicles upon raising the pH to 7. This is surprising, since only trace amounts of insulin associate to the sulfatide-containing vesicles upon direct incubation at pH 7. The possible molecular explanation of the phenomenon and the relevance of these findings to the actions of insulin in vivo are discussed.

Circular Dichroism↗
PubMed1987

Chromosomal aberrations induced in human cultured cells by liposome-encapsulated deoxyribonuclease I.

Experiments of incorporation of a nucleolytic enzyme into human cells cultured in vitro have been carried out with the aim of inducing structural chromosome variations. Human heteroploid cells, either as asynchronous populations or enriched in mitoses, and PHA-stimulated lymphocytes were used as recipients. We found that all these cells when exposed to pancreatic DNAase I encapsulated in liposomes, either of multilamellar (MLV) or of small unilamellar (SUV) type, show an incidence of chromosome damage higher than that induced by the enzyme free in the incubation buffer. Our results indicate that liposomes are suitable vehicles for the transfer of an exogenous nuclease into human cultured cells. The enzyme remains functionally active and interacts with nuclear DNA, giving rise to chromosome lesions.

Aneuploidy↗
PubMed1987

Preparation of spin-labelled sulfatides for EPR studies on model membranes.

Using the N-hydroxysuccinimide ester of the fatty acids, galactosylceramide I3-sulfate containing a 5-or 16-doxyl-stearoyl residue was prepared in good yield by acylation of the galactosylsphingosine I3-sulfate (lysosulfatide) obtained from the saponification of the bovine brain sulfatide. The EPR behavior of the two semisynthetic sulfatides was analyzed in natural sulfatide micelles and in multilamellar vesicles of egg phosphatidylcholine. The evaluated parameters demonstrate that these spin-labelled sulfatides can be used for the study of sulfatide behavior in lipid structures.

Animals↗
PubMed1986

Fluorospectroscopic studies of mixtures of distearoylphosphatidylcholine and sulfatides with defined fatty acid compositions.

Simple study models characteristic for lamellar organization of distearoylphosphatidylcholine and sulfatide have been prepared for fluorospectroscopic investigations on the influence of these glycolipids on the chemico-physical properties of lecithin bilayers. The motion of 1,6-diphenyl-1,3,5-hexatriene in mixed lecithin-sulfatide bilayers changed with temperature, with the compositional ratio of the two lipids, with the presence of divalent cations such as Ca2+ and with the fatty acid composition of sulfatide moiety. Steady-state fluorescence measurements of the average motion of the fluorophore permit evaluation of the gel to liquid-crystalline phase transition in all these membrane models containing different sulfatides.

Fatty Acids↗
PubMed1986

Calorimetric properties of mixtures of distearoylphosphatidylcholine and sulfatides with definite fatty acid composition.

The thermotropic behavior of multilamellar liposomes prepared from mixtures of distearoyllecithin and sulfatide of definite fatty acid composition, has been studied by DTA calorimetry. The saturated stearoyl- and lignoceroyl-sulfatides do not modify significatively both the delta H and the Tm of the phase transition of the lecithins. These parameters, on the contrary, are decreased by all the sulfatides containing unsaturated fatty acids. A contrasting effect is at last exhibited by alpha-OH-stearoylsulfatide which promotes a decrease in delta H but not in Tm of the phase transition. The possibility that these sulfatide properties can influence the dynamic interactions and the sovramolecular organization of the phospholipid bilayers has been discussed.

Calorimetry↗
PubMed1985

Liposomes induce chromosome aberrations in human cultured cells.

The genotoxic effect of multilamellar lipid vesicles (MLV) was analysed on cultured heteroploid and diploid human cells. Dose-dependent reduction of cell survival and mitotic rate as well as induction of chromosome aberrations were observed. Chromatid and chromosome breaks and chromatid exchanges were found in 24-h culture after liposome treatment, whereas chromosome rearrangements were prevalent at 48 h. Neutral (PC/Chol) and positive (PC/SA) MLV showed a greater damage than negative (PS/PC; PS) MLV. Fibroblasts were the most sensitive cell type. In the case of PC/Chol MLV vesicles, control experiments with PC and Chol of controlled purity ruled out the possibility that the observed chromosome aberrations were caused by toxic oxidation products present in commercial preparations.

Cell Line↗