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F Persat

Publications and source records attributed to F Persat.

34 records · Page 2Linked to original sources

Inhibition of human peripheral blood mononuclear cell proliferative response by glycosphingolipids from metacestodes of Echinococcus multilocularis.

The effect on human peripheral blood mononuclear cells (PBMCs) of neutral glycosphingolipids extracted from metacestodes of the parasite Echinococcus multilocularis was investigated. Neutral glycosphingolipids inhibited [3H]thymidine uptake by human PBMCs upon stimulation by mitogens such as phytohemagglutinin A and pokeweed mitogen or by allogeneic Burkitt B cells. This effect was dose dependent and was related to a decrease in interleukin 2 (IL-2) synthesis, the expression of IL-2 receptors (CD25) being unmodified. Addition of exogenous recombinant IL-2 restored the cell proliferation. Partial inhibition of immunoglobulin G (IgG), IgA, and IgM synthesis was observed in the supernatant of cell culture in association with the inhibitory effect. Identification of active subfractions contained in the neutral glycosphingolipid fraction was also studied in relation to cell viability. The free ceramide fraction had an inhibitory effect, in part related to cell lysis, particularly at high concentration, while the monogalactosylceramides had a paradoxical effect: as an activator at low concentrations and as an inhibitor at high concentrations, with limited cell survival. The immunogenic neutral glycosphingolipids containing at least two carbohydrate residues, all having a structure based on Gal beta 1-->6Gal, were inhibitors of PBMC proliferation and showed good cell survival. These results suggest that parasite neutral glycosphingolipids may play an immunologically relevant role in alveolar hydatid disease.

Animals↗

Free ceramides of Echinococcus multilocularis metacestodes.

Free ceramides were isolated and purified from the metacestodes of Echinococcus multilocularis. Two different fractions were obtained by preparative thin-layer chromatography. Their structure was determined by gas chromatography and electron impact mass spectrometry of trimethylsilylated derivatives. The ceramide with the higher thin-layer chromatographic migration rate contained exclusively erythro-sphinganine associated with saturated C16, C18 and very-long-chain fatty acids (up to C30) and unsaturated C24 fatty acid. The second ceramide contained 90.3% sphingosine and 9.7% sphinganine associated with saturated C16 and C24 and unsaturated C18 and C24 fatty acids. These findings were discussed with regard to the structure and metabolic pathway of neutral and acid glycosphingolipids found in the metacestodes.

Animals↗

Gangliosides of Echinococcus multilocularis metacestodes.

Gangliosides, glycosphingolipids with sialic acid, were found in metacestodes of Echinococcus multilocularis in low quantities. All gangliosides were resolved after preparative high-performance thin layer chromatography into four fractions. Cholera toxin was specifically bound to the major ganglioside, allowing the identification of it as a GM1. Precise structure of the four fractions was determined by sequential degradation by exoglycosidases, gas chromatography, electron impact mass spectrometry and liquid secondary ion-mass spectrometry. Beside GM1, the other fractions were GM3, GD1a and, at a lesser percentage, GM2, all belonging to the same a-ganglio-series. The ceramide part of these parasite gangliosides contained sphingosine associated to unsaturated n24, saturated n24 and n16 fatty acids.

Animals↗

Glycosphingolipids with Gal beta 1----6Gal sequences in metacestodes of the parasite Echinococcus multilocularis.

Neutral glycosphingolipids of the metacestodes of Echinococcus multilocularis, an animal and human parasite, were resolved by high performance thin layer chromatography into 12 fractions. Nine of these fractions were permethylated, analyzed by electron impact-mass spectrometry, and submitted to methylation analysis by gas chromatography-mass spectrometry. Native fractions were analyzed by liquid secondary ion-mass spectrometry and degraded sequentially by exoglycosidases. In addition to a previously described galactosylceramide, a di-, a tri-, and a tetragalactosyl-ceramide having Gal beta 1-6Gal internal linkages were characterized. This type of carbohydrate chain has been described in glycolipids of a marine mollusk, Turbo cornutus (Matsubara, T., and Hayashi, A. (1981) J. Biochem. (Tokyo), 89, 645-650). In addition two novel fucolipids were found with the following structures: Fuc alpha 1-3Gal beta 1-6Gal-Cer and Gal beta 1-6(Fuc alpha 1-3)Gal beta 1-6Gal-Cer. Ceramides contained sphinganine and either nonhydroxy fatty acids with 16, 18, 26, and 28 carbon atoms, or hydroxy fatty acids, with 16 and 18 carbon atoms. Di-, tri-, and tetragalactosylceramides containing the Gal beta 1-6Gal disaccharide were found to be immunogenic in humans.

Animals↗

Plasma itraconazole concentrations in neutropenic patients after repeated high-dose treatment.

Two different treatments with repeated oral high doses of itraconazole were tested for 10 days in 20 neutropenic patients, 10 receiving 400 mg per day and 10 receiving 600 mg per day. In each group 5 patients were treated for acute leukaemia and 5 patients were recipients of autologous bone-marrow transplantation (ABMT). Itraconazole plasma concentrations were assayed by high-performance liquid chromatography. Statistical analysis disclosed a significant interaction between the dispensed dose and the patient types. The difference between the two doses of itraconazole was greater in the ABMT than in the leukaemia patients. After 10 days at 600 mg per day all the ABMT patients had an itraconazole plasma concentration higher than 250 micrograms/l. Therefore, 600 mg per day seems more efficient to obtain a therapeutic level of itraconazole in ABMT patients but this needs to be confirmed for all the neutropenic patients.

Acute Disease↗

Detection of antibodies against glycolipids of Echinococcus multilocularis metacestodes in sera of patients with alveolar hydatid disease.

Glycosphingolipids extracted from Echinococcus multilocularis metacestodes were tested against sera from patients with alveolar hydatid disease (AHD). The tests were performed first by ELISA then by immunostaining on thin layer chromatography. A binding between the parasite glycolipids and antibodies of AHD human sera was observed by ELISA. The glycolipids related to this binding were specifically parasite neutral glycosphingolipids. No reaction was observed with parasite acid glycolipids nor with neutral glycolipid extracts from human or mongolian gerbil erythrocytes. Comparison between absorbance values of 25 AHD sera and 20 control sera showed significant differences. Similar results were obtained with sera from hydatid cyst patients. Sera from patients with other parasitoses (schistosomiasis, strongyloidosis or paludism) were also tested. The reactive fractions were identified by immunostaining on thin layer chromatography with AHD sera, they were neutral glycosphingolipids containing at least two carbohydrate residues. These results were compared with chromatograms obtained with hydatid cyst human sera and discussed.

Animals↗

Glycosphingolipids of Echinococcus multilocularis metacestodes.

Neutral and acid glycosphingolipids of Echinococcus multilocularis metacestodes that were obtained after intraperitoneal infection of Meriones unguiculatus have been analyzed by thin layer chromatography. Neutral and acid glycosphingolipids accounted for 95% and 5% of total glycosphingolipids, respectively. 12 different fractions were observed in the neutral glycosphingolipids extracts of the parasite. The most important was a monohexosylceramide fraction accounting for 56.4% of neutral glycosphingolipids. 9 different fractions were detected in gangliosides (acid glycosphingolipids). The fact that these glycosphingolipids were specific to the parasite was established by the analysis of different cell populations of the host. Glycosphingolipids were purified from control and parasite-infected gerbil blood cells as well as from peritoneal exudate cells of healthy gerbils after a non-specific immunostimulation. The chromatograms obtained with these extracts were totally different from the parasite. In addition, parasitosis was found to have no effect on the host blood cell glycosphingolipids.

Animals↗

Analysis of the monohexosylceramide fraction of Echinococcus multilocularis metacestodes.

Monohexosylceramides of Echinococcus multilocularis metacestodes have been isolated and analyzed by thin-layer chromatography, gas-liquid chromatography and mass spectrometry. 90.9% of the parasite fraction was galactosylceramide; glucosylceramide was present at only 9.1%. The most important fatty acids were normal C16:0 and C26:0 fatty acids. The hydroxylated fatty acids of the ceramide part constituted 20.1% of the total, their major constituents were C18:0 and C26:0. The sphinganine accounted for 70.4% of long-chain bases, phytosphingosine and sphingosine were also detected. The importance of the long chain fatty acids and the presence of sphinganine in the monohexosylceramide fraction were discussed.

Animals↗

[Praziquantel in human hydatidosis. Evaluation by preoperative treatment].

The efficacy of praziquantel has been studied in human hydatic disease due to E. granulosus, using parasitological and pharmacological criteria of improvement. This evaluation uses a prospective therapeutic trial in 15 patients with one or several hydatic cysts in different sites. Before surgery, nine of these received daily 75 mg/kg of praziquantel in 2 courses of 10 days each. The other six patients are considered as controls. The protoscolices vitality is determined by direct optic microscopy and by intraperitoneal mouse inoculation. According to the results of optic microscopy, praziquantel sterilizes hydatic cysts: 19 sterilized cysts out of 26 coming from treated patients, and 4 out of 11 in control group. This effect seems to be emphasized in hepatic localization. Nevertheless, according to the results of mouse inoculation, praziquantel does not significantly reduce the pathogenicity of inoculated proscolices and germinal layer: 11 mice out of 39 are healthy in the treated group, and 5 out of 15 in controls. Moreover, the determination with a fluorimetric method has not detected praziquantel in the hydatid liquid issued from treated patients. The drug does not pass through the cyst wall, and so, cannot have a scolicidal activity. Praziquantel cannot be considered as a medical treatment in human hydatidosis.

Animals↗

A comparative study of UTP-D-glucose-1-phosphate uridylyl transferase in the cysts of Echinococcus multilocularis and the livers of infected and control Meriones unguiculatus.

Kinetic and physical parameters of UDP-glucose pyrophosphorylase were determined in Meriones unguiculatus infected with Echinococcus multilocularis metacestodes (cestoda). Studies were carried out on parasite cysts, and on livers from control and infected animals after purification of the enzyme by affinity chromatography on UTP-agarose. The enzyme from infected and control livers had km values for UTP of 0.01 mM and 0.5 mM, respectively; for glucose-1-phosphate values were 0.46 mM and 0.07 mM, respectively. On the other hand the enzyme from cysts was found to have a higher Km for UTP (1 mM) and for glucose-1-phosphate (1.5 mM) than from infected or non-infected livers. Physical characteristics (pI = 6 and Mr = 160,000) of UDP-glucopyrophosphorylases were the same in controls and infected host livers but were different from the cyst enzyme (pI = 7 and Mr = 251,000). These results provide evidence for the existence of significant differences between parasitic and host enzymes, which could possibly be exploited in chemotherapy.

Animals↗

Purification of a UTP:D-glucose-1-phosphate uridylyltransferase from Golgi apparatus of cat liver by affinity chromatography on UTP-agarose.

UDP-glucose pyrophosphorylase from Golgi apparatus solubilized by detergent has been purified 100-fold from microsomes by affinity chromatography on UTP-agarose. The purified enzyme has apparent Mr 270,000 and isoelectric pH 3.9 against 360,000 and 4.2 for soluble enzyme. According to these characteristics, UDP-glucose pyrophosphorylase from Golgi apparatus is different from cytosolic enzyme.

Animals↗

Evidence for coupling between transport of UDP-glucose and its synthesis by membrane-bound pyrophosphorylase in Golgi apparatus of cat liver.

Incubation of sealed vesicles of cat-liver Golgi apparatus with UDP[14C]glucose showed that the vesicles accumulated radioactivity. After Triton X-100 treatment or sonication of washed vesicles, soluble radiolabeled species were released and identified by paper chromatography as UDP[14C]glucose, [14C]glucose 1-phosphate and free glucose. In the incubation medium, UDPglucose was effectively protected by addition of dimercaptopropanol and UTP. Presence of glucose 1-phosphate and glucose within the vesicles most probably arose from luminal pyrophosphatase and phosphatase. A portion of the [14C]glucose moiety became covalently linked to endogenous acceptors. Uptake of UDPglucose was saturable and dependent on time and on the concentration of sugar nucleotide. Together, these results were consistent with a transport system for UDPglucose in Golgi vesicles. Furthermore, penetration rate was considerably higher with UDPglucose synthetized in situ from glucose 1-phosphate by membrane-bound pyrophosphorylase than from added UDPglucose: Vmax values were respectively 10 and 2 pmol/15 min per mg protein. This result allows the conclusion that a coupling between translocase and synthetase is involved in UDPglucose transport through Golgi apparatus membranes. The mechanism of this 'kinetic advantage' is discussed.

Animals↗

Properties of uridine diphosphate glucose pyrophosphorylase from Golgi apparatus of liver.

Golgi apparatus isolated from cat liver contained UDPglucose pyrophosphorylase (UTP:alpha-D-glucose-1-phosphate uridylyltransferase, EC 2.7.7.9) activity. The results of washing suggested that pyrophosphorylase was bound firmly to Golgi membranes. Moreover, the enzyme was activated by Triton X-100 in the same extent as galactosyltransferase, a typical Golgi apparatus enzyme. Two-substrate kinetic studies were performed with the enzymes from cytosol and Golgi fractions. The soluble enzyme showed an apparent 2.5-fold greater activity for the glucose 1-phosphate than for UTP, while pyrophosphorylase of Golgi apparatus had the same affinity for the two substrates. A random mechanism was observed with a direct dependence of apparent Michaelis constant values on the concentration of second substrate for soluble enzyme. In contrast, with Golgi enzyme one ligand had no effect on the binding of the other.

Animals↗

Specific antibody detection in human aspergillosis: a GEMO* multicentre evaluation of a rapid immunoelectrophoresis method (Paragon). Group d'Etude des Mycoses Opportunistes.

A new immunoelectrophoresis system, the Paragon system, was evaluated in three different hospital centres with the aim of improving standardization of the serodiagnosis of human aspergillosis. To select the most efficient antigen, various commercial and home-made antigens were first tested on 19 sera from 19 patients with highly probable aspergillosis. The value measured using the Paragon anti-Aspergillus antibody detection system was then compared with the results obtained by conventional serological diagnostic methods (conventional immunoelectrophoresis, enzyme linked-immunosorbent assay, indirect immunofluorescence): this step was performed using the first 19 sera as well as 16 other sera from 13 patients with suspected aspergillosis. Concordant results were obtained in 28 cases. The discrepancies observed with seven sera were probably related to differences in the nature of the antigens. Paragon immunoelectrophoresis proved to be a practicable technique requiring only a small amount of serum and giving results within a shorter time than competitive methods (24-48 h). Its major drawbacks compared with conventional immunoelectrophoresis are some difficulties in reading, fewer precipitin lines and the relatively high cost of routine analysis.

Antibodies, Fungal↗