Sialidase and beta-galactosidase activities in serum of alcoholic patients.
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Adult↗
Biomedical subjects
H Stibler
Publications and source records attributed to H Stibler.
The sialic acid and galactose concentrations in erythrocyte membranes in patients with myotonic dystrophy, limb-girdle and facioscapulohumeral dystrophy.
Sialic acid is an important constituent of membrane-bound glycoproteins and glycolipids. It occurs linked to galactose at the surface of the membrane and is involved in, e.g., cation exchange, receptor function, maintenance of membrane polarity and intercellular interactions. In myotonic dystrophy there is evidence of an as yet basically undefined plasma-membrane abnormality. Considering the importance of sialic acid in membrane function, sialic acid as well as galactose concentrations were measured in erythrocyte membranes from 17 patients with myotonic dystrophy and compared to 17 matched healthy controls. There was a highly significant (P less than 0.0005) reduction of the sialic acid concentration in the patients, while no significant difference in galactose concentration was found. In 16 patients with limb-girdle and facioscapulohumeral dystrophy, sialic acid and galactose concentrations did not differ from matched controls. The possible importance of a reduced concentration of membrane-bound sialic acid in myotonic dystrophy is discussed in relation to previously reported biochemical membrane abnormalities in this disease.
Adult↗
Effect of ethanol on synaptosomal sialic acid metabolism in the developing rat brain.
The total, glycoprotein-bound and glycolipid-bound sialic acid concentration, ad the activities of ecto-sialyltransferase and neuraminidase were determined in synaptosomes from preweanling ethanol-treated and control rats. The period of treatment corresponded to that of maximal synaptogenesis and peak synthesis of sialoglycocompounds (days 27-37 postconception). The average of the peak blood ethanol concentration was 271 mg/100 ml. In the ethanol-treated animals the sialic acid concentration was significantly reduced (approximately 20%) with an equally distributed decrease of glycoprotein- and glycolipid-bound sialic acid. The activity of ecto-sialyltransferase with asialofetuin as exogeneous acceptor was significantly diminished (about 30%) in the ethanol-treated pups. Neuraminidase showed an unchanged activity after correction for the reduction of endogeneous sialic acid substrate concentration. The total protein and lipid concentrations of the synaptosomal preparations did not differ between the groups. These results suggest that ethanol treatment during on of the vulnerable periods of brain development causes an inhibition of the incorporation of sialic acid into synaptosomal membrane-bound sialoglycocompounds. Such an effect of ethanol exposure might disturb intercellular interactions and the functional performance of the membrane during development, and could be of importance in the pathogenesis of the central nervous system manifestations of the fetal alcohol syndrome.
Animals↗
Brain synaptosomal (Na+ and K+)ATPase activity as an index of tolerance to ethanol.
Variations of brain synaptosomal (Na+ and K+)ATPase activity and development of functional tolerance to ethanol were followed simultaneously throughout a chronic ethanol treatment. Inhibition of the enzyme activity by ethanol added in vitro in the presence of noradrenaline was also assayed. Adult male rats were rendered tolerant to ethanol by daily intragastric administration of doses of 3-6 g of ethanol/kg body wt. As assessed by the hypothermic effect and impairment of motor performance on a tilting plane after injection of a challenge dose of ethanol (3 g/kg body wt. IP), functional tolerance developed slowly, was demonstrable after 2 weeks of treatment and increased for up to 4 weeks. The two tolerance tests gave parallel and positively correlated results. Concurrently with the development of tolerance, the basal activity of brain synaptosomal (Na+ and K+)ATPase increased (7% to 18%) in preparations from ethanol treated animals when compared to those from starch-fed caloric controls, and there was less inhibition of the enzyme by ethanol added in vitro in the presence of noradrenaline. The time course of the appearance of these changes in enzyme activity were positively correlated with that of behavioral tolerance, strengthening a relationship between the phenomena events. The intensity of the noradrenaline-ethanol interaction with the membrane-bound (Na+ and K+)ATPase activity could be an index of the degree of tolerance to ethanol.
Alcoholism↗
Reduction of the sialic acid and galactose concentrations in erythrocyte membranes in alcoholics.
The sialic acid and galactose concentrations in erythrocyte membranes were examined in 32 and 17 alcoholics respectively, during current abuse, and the sialic acid concentration in 10 patients after one and four weeks of abstention. During current alcohol abuse the concentrations of these two surface carbohydrates were decreased by an average of 10% and 16% respectively, when compared to matched, healthy controls (p less than 0.0005). After one and four weeks of abstention the sialic acid values were not significantly different from those of the controls suggesting that the abnormal cells were eliminated within one week after cessation of alcohol intake. No correlations were found between the sialic acid or galactose concentrations and peripheral red cell characteristics or serum liver enzyme levels. Hemolysis in isotonic Tris was significantly increased during current abuse, and was highly dependent on the sialic acid concentration at one particular stage of Tris-induced lysis. The mechanism behind the change of the oligosaccharide concentration of the membrane may either be increased hydrolytic cleavage or inhibition of the synthesis of glycoproteins and glycolipids. Previous similar findings in serum transferrin in alcoholics and in synaptosomes from young rat brain after ethanol exposure indicate that glycoprotein and glycolipid metabolism may be an important target, directly or indirectly, in the biological actions of ethanol.
Adult↗
Evidence of a reduced sialic acid content in serum transferrin in male alcoholics.
A qualitative change of the microheterogeneity of serum transferrin, demonstrated by isoelectric focusing, has previously been found to occur frequently and with high specificity in alcoholic patients during current abuse, and has been proposed as a new marker of alcoholism. Certain indirect evidence has supported the assumption that the basis for the altered transferrin heterogeneity would be a reduced sialic acid content. In this investigation serum transferrin from healthy controls and alcoholic patients was purified by affinity chromatography on antitransferrin Sepharose 4B. The sialic acid content in transferrin was thereafter determined directly. Transferrin from alcoholic patients showed a 22% lower sialic acid concentration than control transferrin, which was highly significant. These data together with results from experiments with neuraminidase and galactose-binding lectin provide evidence that in alcoholism at least two sialic acid residues are missing in a significant fraction of serum transferrin. This observation may indicate that sialic acid residues are missing in a significant fraction of serum transferrin. This observation may indicate that sialic acid metabolism is one important target of the biological action of ethanol.
Adult↗
Determination of terminal sugars in transferrin by radio-lectin immunoassay (RLIA)-a new microanalytical procedure.
A new method for the determination of terminal sugars in immunologically defined glycoproteins with microheterogeneity in the sugar residue has been developed. The method has been elaborated for transferrin and involves the following three steps: 1. Binding of antitransferrin antibodies to cyanogen bromide activated Sepharose. 2. Adsorption to the antitransferrin gel of transferrin from serum or from standard solutions of defined composition of terminal glycoprotein sugars. 3. Adsorption of a 125I - labelled lectin to the Sepharose-antitransferrin-transferrin complex. A galactose-binding lectin from Crotalaria juncea and a sialic acid-binding lectin from Limulus polyphemus have been successfully labelled with the Bolton-Hunter reagent and used in the radio-lectin immunoassay determinations. Care must be taken that significant amounts of lectins are not lost from the transferrin complex during washing. The resolving power of the method was at best 20 pmoles of asialotransferrin, a figure that probably can be improved significantly by optimizing the assay conditions.
Adsorption↗
Quantitative estimation of abnormal microheterogeneity of serum transferrin in alcoholics.
A qualitative abnormality of the microheterogeneity of serum transferrin, demonstrated by isoelectric focusing, has previously been shown to be a highly specific indication of chronic alcoholism. The abnormality consists of a selective increase of a cathodal transferrin component which is probably caused by a reduction of the sialic acid content. The present study describes a method for quantitative estimation of the abnormal transferrin. The technique was based on analytical isoelectric focusing as the first step followed by direct immunofixation. The immunofixed transferrin was quantified by computerized on-line densitometry, and the transferrin abnormality was calculated as a quotient, where the amount of the cathodal component was expressed as a percentage of the relative total immunofixed transferrin quantity. This method was shown to possess high sensitivity and good reproducibility. In the controls the mean value of the quotient was 3.7%, while in the alcoholics it was 9.5% which was a highly significant difference (p less than 0.001). The possible functional significance of a disturbed sialic acid metabolism in alcoholism is discussed.
Adult↗
Abnormal heterogeneity of serum transferrin--a new diagnostic marker of alcoholism?
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Adult↗
Abnormal microheterogeneity of transferrin--a new maker of alcoholism?
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Adult↗
Direct immunofixation after isoelectric focusing. An improved method for identification of cerebrospinal fluid and serum proteins.
An improved method for identification of CSF and serum proteins is described, using analytical isoelectric focusing followed by direct immunofixation in polyacrylamide gel. This method offers high sensitivity together with retained resolution after isoelectric focusing and is technically easy to perform. The gamma-trace protein, normal transferrin, haptoglobin and alpha 2-macroglobulin and two genetic alterations of transferrin were analyzed. The qualitative differences, previously observed on crossed immunoelectrofocusing, between transferrin and alpha 2-macroglobulin in the CSF and serum were confirmed by immunofixation. Protein components, differing in isoelectric pH by at least 0.03 pH unit and 0.5 mm apart were readily identified by this technique.
Blood Proteins↗
Clinical significance of abnormal heterogeneity of transferrin in relation to alcohol consumption.
An abnormal microheterogeneous component of serum transferrin, with a higher isoelectric point than the normal main component, was studied by means of isoelectric focusing and direct immunofixation in 98 alcoholic patients, 22 patients with liver diseases and 100 controls. Its relation to acute and prolonged ethanol intake was studied in healthy volunteers. The abnormal transferrin component was found to be a sensitive indicator of prolonged, high alcohol ingestion, and was observed in 81% of patients with an admitted consumption of more than 60 g ethanol/day, and normalized after at least 10 days of abstinence. It occurred in 1% of the controls and in none of the cases with liver diseases without current alcohol abuse. There is evidence of a reduced sialic acid content in the abnormal transferrin. No similar change has been found in a number of other glycoproteins. This test may be a useful and sensitive tool for the detection of chronic alcohol consumption.
Adult↗
Subtypes of transferrin C.
Subtypes of transferrin C were studied by means of isoelectric focusing after complete desialylation of transferrin. Family data were consistent with an autosomal co-dominant mode of inheritance. Studies of serum samples from 75 individuals heterozygous for C and another (B or D) variant showed that the genes (C1 and C2) controlling the C subtypes are allelic to the B and D genes. The C2 gene frequency in Swedes and Swedish Lapps was similar to that found previously in Danes and Germans.
Gene Frequency↗
The normal cerebrospinal fluid proteins identified by means of thin-layer isoelectric focusing and crossed immunoelectrofocusing.
The cerebrospinal fluid and serum proteins from 32 healthy individuals have been examined by isoelectric focusing in a pH-gradient from 3.5--11.0. Seventeen normal proteins have been identified by means of crossed immunoelectrofocusing. The pI-values of the main fractions of these proteins have been determined. On crossed immunoelectrofocusing microheterogeneity was observed for all of the proteins except haemopexin, gamma-trace protein of CSF and prealbumin of serum. Differences between serum and CSF were observed for 6 of the proteins. Prealbumin of CSF had a lower pI-value than in serum and showed partial immunological identity with prealbumin of serum. Transferrin of CSF included several components with lower sialic acid contents than in serum. The conversion of C'3-complement from beta-1-C to beta 1-A was slower in CSF than in serum. Qualitative differences between CSF and serum were also noticed for alpha2-macroglobulin and haptoglobin. IgM was not detected in CSF. In CSF the gamma-trace protein was found on isoelectric focusing in one third of the subjects. Two subjects had 2 close tau-fractions and another two had one unidentified band in the anodal IgG-area. The gamma-trace protein did not show any proteolytic activity, nor any resemblance in charge to 4 basic enzymes or with the histones tested.
Adult↗
Serum protein binding of lead in vitro in amyotrophic lateral sclerosis patients and controls.
The binding of lead to serum proteins was studied in vitro in ALS-patients and controls. Serum was incubated with 210Pb, the proteins were separated by isoelectric focusing and the radioactivity in the different protein fractions was determined by liquid scintillation counting. The activity was concentrated in the orosomucoid (acid alpha1-glycoprotein) fraction both in ALS-patients and controls under the present experimental conditions and the capacity of this protein to bind lead was demonstrated in a control experiment. The findings are discussed in relation to our earlier observations on increased concentrations of lead in cerebrospinal fluid and plasma in ALS-patients and the hypothetical role of the retrograde axonal transport in motoneurons in the pathogenesis of ALS. The present observations do not lend support to the idea that the increased plasma concentrations of lead in the disease would be related to qualitative differences in serum protein binding.
Adult↗
Abnormal microheterogeneity of transferrin in serum and cerebrospinal fluid in alcoholism.
The serum and CSF proteins were analyzed by isoelectric focusing in 16 male alcoholics after alcohol intoxication and after 10--14 days of alcohol abstinence. An abnormally marked protein band with pI 5.7 was found in serum in 15 patients and in CSF in 12 at the first examination. On crossed immunoelectrofocusing it appeared as an increased, cathodal, microheterogeneous molecular form of transferrin. The abnormality was reversible and decreased or normalized in serum in all cases after abstinence. In 6 patients with clinical signs of cerebellar degeneration, an abnormal microheterogeneous pattern of CSF transferrin of partly different appearance to that in serum remained after abstinence. Disturbed liver synthesis of transferrin is a probable origin of the serum finding, which may be specifically related to alcohol abuse. Substitution or loss of acidic amino acids and/or decreased iron binding ability are possible structural explanations.
Adult↗
Crossed immunoelectrofocusing for identification of normal and abnormal cerebrospinal fluid proteins. A preliminary report.
Isoelectric focusing is a valuable method for the analysis of cerebrospinal fluid (CSF) proteins. Its high resolving power has, however, created problems in identifying the large number of protein bands separated. In an attempt to identify these bands, two-dimensional crossed immunoelectrofocusing has been used, where isoelectric focusing is combined with rocket immunoelectrophoresis. By these methods 9 of the normal proteins in the acidic pH interval have been identified. In addition the unusual CSF protein abnormalities occurring in Marie-Sanger-Brown's ataxia and alcoholic cerebellar degeneration have been shown to represent increases of different microheterogeneous forms of transferrin.
Alcoholism↗
Isoelectric focusing and electrophoresis of the CSF proteins in tremor of different origins.
The CSF proteins have previously been very little investigated in the cerebellar syndrome of chronic alcoholism and in essential tremor. Such studies have been carried out more thoroughly by electrophoretic methods in Parkinson's disease but generally with normal results. In the present investigation the CSF proteins were examined by isoelectric focusing and quantitative paper electrophoresis in 10 patients with the cerebellar syndrome of chronic alcoholsm, 12 patients with Parkinson's disease and 16 subjects with essential tremor. Abnormal CSF proteins of very similar appearance were found on isoelectric focusing in the acidic pH interval 5.6-5.8 in 80% of the patients with the cerebellar syndrome of chronic alcoholism. In Parkinson's disease the most common aberration was evidence of nonspecific blood-CSF-barrier damage which occurred in half of the patients. In only 17% of these cases did other alterations appear, situated in the pH range alkaline to pH 5.8. Abnormal CSF proteins were found in 94% of the patients with essential tremor. The aberrant proteins appeared in both the acidic and alkaline pH regions, most frequently with anisoelectric point at pH 5.9, 7.2 and 9.3. There was a considerably higher frequency of CSF protein abnormalities in different pH ranges in patients with tremor of more pronounced degree as compared to those with only mild symptoms. The electrophoretic examinations failed to show any conclusive alterations. Barrier-damage patterns of mild or moderate degree or slightly increased levels of CSF beta1-globulin were occasionally found in all 3 diseases. The results indicate that isoelectric focusing of the CSF proteins may be of diagnostic value in the cerebellar syndrome of chronic alcoholism and in essential tremor but does not reveal any characteristic abnormalities in Parkinson's disease.
Adult↗