Emilia Zaprianova

Serum ganglioside patterns in multiple sclerosis.

The relative distribution of gangliosides was determined in the serum of 37 patients with multiple sclerosis (MS) and of 30 healthy subjects. There was a significant increase of GM1 and GD1a, and a decrease of GM3 proportion in the serum of relapsing-remitting MS patients (RRMS) during their first MS attack. The RRMS patients in relapse with a long duration of the disease had a significant decrease of GM1 and an increase of GD1a portion in the serum. An increase of GD1a, one of the major brain neuron ganglioside fraction, suggested the neuron injury in the early and with a long duration RRMS. The finding of an increase of GM1, the main human myelin ganglioside, during the first MS attack in RRMS patients confirms previous evidence for the possible involvement of gangliosides in the early pathological course of demyelination in MS.

Direct Thin-Layer Chromatographic Method for Isolation of Gangliosides

Abstract A simple procedure is developed for obtaining a ganglioside mixture. The scheme of the method is as follows: the sample was homogenized and extracted twice with cyclohexane (first extraction). The residue was reextracted with a mixture of chloroform /methanol = 1:3 (v/v) (second extraction). The extracts were collected after solvent evaporation. The residue was dissolved and applied onto a thin layer of silica gel according to the requirements of the preparative thin-layer chromatography. After the zone separation the visualized spots were scrapped and transferred to a column and eluted with a mixture of chloroform/methanol with increasing polarity. The eluate was concentrated and controlled for purity and composition by HPTLC. This procedure has been successfully applied to fractionation of ganglioside mixtures from human, calf and rat brains.

Ganglioside spinal cord changes in chronic relapsing experimental allergic encephalomyelitis induced in the Lewis rats.

Chronic relapsing experimental allergic encephalomyelitis (CREAE) was induced in Lewis rats by inoculation with guinea-pig myelin and complete Freunds adjuvant followed by treatment with low-dose cyclosporin A. Rats were sacrified at different phases of the disease (just before the onset of clinical signs, during the first clinical episode of CREAE and during the first recovery). Gangliosides were extracted from the spinal cord, analysed after purification by two-dimensional chromatography and quantified densitometrically. An increase of GM1, the main rat myelin ganglioside, and a decrease of GT1b, suggested to play a role in mediating the interactions between oligodendroglia and axons, were observed during the development of the CREAE. These findings indicating significant ganglioside changes in CREAE give further support to the concept concerning the involvement of gangliosides in autoimmune demyelination.

A Variant for Isolation of Serum Gangliosides

Abstract The serum gangliosides are of great diagnostic importance. A step-by-step procedure for their isolation from serum is described and it includes the following stages: a) dehydration of the sample; b) total lipids extraction; c) non-polar lipids removal by preparative TLC; d) elimination of the blood sugar by Sep-Pak technique; e) TLC of the ganglioside fractions. Azeotropic distillation of the mixture of serum water/n-propanol was used for sample dehydration. Triple extraction was carried out: with cyclohexane (I); chloroform : methanol = 1 : 1 (v/v) (II) and chloroform : methanol = 1 : 2 (v/v) (III). The extracts were combined and the non-polar lipids were removed. The eluate composition was checked out by HPTLC with a modified mobile phase: chloroform:methanol:0.1 M sodium lactate = 55 : 40 : 10 (v/v/v). Comparative data for the Rf values of the ganglioside fractions as well as recovery data are presented. This method is also applicable for milk samples.

A method for determination of lipid-bound sialic acid after chromatographic isolation of brain gangliosides

Abstract The lipid-bound sialic acid determination gives an indirect idea of the ganglioside composition. An attempt at adapting the ‘resorcinol method’ of Svennerholm using thin-layer chromatography is described in this article. An aliquot of a lipid extract from bovine brain was chromatographed and the ganglioside carrying area was visualised. The sorbent was scrapped and eluted. The sialic acid was hydrolysed with a mixture of resorcinol/hydrochloric acid and the color product formed was extracted with butanol/butylacetate. The color intensity was measured at 580 nm. It was found that the lipid-bound sialic acid in bovine brain was about 780 μg/g fresh tissue. The recovery data calculated by means of a standart solution were 97–102%. This method is particularly useful for some routine diagnostic studies.

Antibodies against gangliosides GM1 of patients with multiple sclerosis alter the response of neurons to synaptic activation

Earlier, we showed in electrophysiological experiments performed with a demyelinated preparation (neural abdominal chain ganglion of the leech) that a 40-min incubation in the presence of rabbit antiganglioside serum caused significant changes in the electric activity of the Retzius neurons [3, 5]. We found that these changes resulted from the interaction between gangliosides contained in the Retzius neuron membrane and antibodies against gangliosides, leading to disturbance of the functional activity of Na-channels for incoming current, which are responsible for the generation of spikes by this cell [5]. As serum of patients with multiple sclerosis often has a high titer of antibodies against gangliosides GM1 [7, 8, 11–13], we assumed that this serum, as well as serum of immunized rabbits, which contains a mixture of four major gangliosides (GM1, GD1a, GD1b, and GT1b), may have a similar effect of neuronal electrogenesis.

Ganglioside Changes in Brain in Chronic Relapsing Experimental Allergic Encephalomyelitis Induced in the Lewis Rat

Chronic relapsing experimental allergic encephalomyelitis (CREAE) was induced in Lewis rats by inoculation with guinea-pig myelin and complete Freunds adjuvant followed by treatment with low-dose cyclosporin A. Rats were sacrified at different phases of the disease (just before the onset of clinical signs, during the first clinical episode of CREAE and during the first recovery). Gangliosides were extracted from the brain, analysed after purification by HPTLC fractionation and quantified densitometrically. An increase of GM1, the main rat myelin ganglioside, and a decrease of GT1b, suggested to play a role in mediating the interactions between oligodendroglia and axons, were observed during the development of the CREAE. These findings indicating significant ganglioside changes in CREAE give further support to the concept concerning the involvement of gangliosides in autoimmune demyelination.