Biomedical subjects
Publications and source records attributed to L Gattegno.
Two immunostimulating peptides, Val-Glu-Pro-Ile-Pro-Tyr and Gly-Leu-Phe, obtained from human caseins, were demonstrated to significantly stimulate binding of human senescent red blood cells to human monocytic-macrophagic cells and their phagocytosis by these cells.
During treatment of human red cells with phospholipase A2 from bee venom, a linear increase of the MCV and of the osmotic fragility occurs in parallel with the cleavage of the accessible phospholipids. However, even after maximal hydrolysis, i.e. degradation of up to 65% of the phosphatidylcholines and up to 6% of the phosphatidyl-ethanolamines, almost no haemolysis is observed and the median corpuscular fragility is only 7% higher than that of control cells incubated without enzyme. Addition of albumin to the medium results in an important rise of the susceptibility to hypotonic saline solutions. Osmotic fragility curves obtained with red cells submitted to mild phospholipase action show evidence of subpopulations of cells with various sensitivities to osmotic lysis. This phenomenon can be partly explained by the heterogeneity of the cleavage intensity among the cell population. This hypothesis is supported by the studies of the lipid composition of phospholipase treated red cells fractionated according to their sensitivity to hypotonic lysis or to their size.
Senescent human erythrocytes (RBC) are able to adhere to and be phagocytized by autologous monocytes in vitro to a greater extent than are young RBC. This adhesion and erythrophagocytosis of senescent RBC is inhibited by D-galactose, N-acetyl-D-galactosamine, their corresponding derivatives of bovine serum albumin, and lactose. On the other hand, D-glucose, D-mannose, L-fucose, N-acetyl-D-glucosamine, and their corresponding derivatives of bovine serum albumin are noninhibiting. The glycopeptides released by tryptic digestion of senescent RBC and purified on immobilized peanut agglutinin are the most effective inhibitors of both RBC adhesion and phagocytosis by autologous monocytes obtained from peripheral blood.
Previous studies have shown that the concentration of red blood cell (RBC) magnesium is significantly lower in subjects carrying an HLA-BW 35 antigen (p less than 0.001) than in non-carriers. As this finding might be related to modifications of the RBC membrane sialoglycoconjugates, RBC sialic acid was comparatively determined in BW 35+ and BW 35- subjects. Pyruvate-kinase activity mean RBC volume, and reticulocyte count have also been determined in order to estimate whether some significant variations in the level of these age markers could be detected between the HLA BW 35+ and BW 35- subjects. A significant negative correlation between sialic acid and RBC magnesium concentrations was observed for the whole population tested (n 57, p less than 0.005), 61% of the BW 35+ and only 25% of the BW 35- individuals having sialic acid values above, and magnesium values below the overall mean (p less than 0.01). The variance of mean RBC volume was also larger for the BW 35+ group. Other determinations did not show any significant variations, suggesting that the results are not related to RBC age.
A rapid procedure for red blood cell isolation by centrifugation is proposed, in which 96% of leucocytes are removed. The middle age of the erythrocytes is verified. This procedure applied to erythrocyte rheological studies (by the initial filtration flow rate) confirms the decreased deformability of senescent red blood cells.
Young and old human erythrocytes, separated in vitro according to age, were labeled at the surface-sialic acid residues by sodium periodate and borotritide treatment. No qualitative difference was observed between the sialic acid derivatives of young- and old-erythrocyte membranes. The number of labeled residues was significantly decreased in the in vivo aged erythrocytes (12 +/- 2.8 X 10(6); n = 8) when compared to the young ones (19.4 +/- 2.8 X 10(6); n = 8). Analysis of the labeled glycoproteins by sodium dodecyl sulfate gel-electrophoresis showed that this decrease affects the PAS-1, PAS-4, PAS-2, and PAS-3 bands (glycophorins A and B) of the old-erythrocyte membranes.
1. Young and old rat erythrocytes, separated in vitro, according to their age in vivo, were radioactively labeled at the cell surface D-galactosyl and 2-acetamido-2-deoxy-D-galactose (Gal and GalNAC) residues by treatment with D-galactose oxidase, followed by reduction with sodium borotritide: 2. The number of labeled surface residues per erythrocyte was found to be 0.52 X 10(6) +/- 0.08 X 10(6) for young, 0.38 X 10(6) +/- 0.05 X 10(6) for middle-aged and 0.32 X 10(6) +/- 0.05 X 10(6) for old, indicating a significant decrease of these surface Gal and GalNAC residues during ageing. 3. Qualitative changes were observed between the sodium dodecyl sulfate electrophoretic patterns of young, middle-aged and old labeled erythrocytes. 4. Quantitative changes were previously observed between the electrophoretic patterns of the labeled glycoproteins from young, middle-aged and old human labeled erythrocytes. 5. It is concluded that some of the changes occurring during in vivo ageing of rat and of human erythrocytes are different.
Red blood cell (RBC) membranes are rich in a glycoconjugate that is extractable in chloroform/methanol solutions (2/1, v/v) and contains several hexoses, such as glucose. Old and young RBC are separated and their respective glycoconjugates are prepared. HbA0 is purified by column chromatography and incubated with solutions of this conjugate. After 24-h incubation, Hb is dialyzed and the amount of glycosylated Hb is measured by a method of column chromatography adapted from Trivelli. A very significant amount of HBAlc is formed when young RBC extracts are incubated: 3.6% of total Hb becomes HBAlc with the extracts, versus 3.2% with free glucose, and only 2.5% for controls. No increase in HbAlc is obtained when extracts of old RBC are incubated. Another difference between the action of the glycoconjugate and free glucose is that the former induces the increase of only the HBAlc fraction, whereas glucose induces the increase of all the minor Hb fractions. The evaluation of glucose contained in the conjugate before and after the glycosylation reaction demonstrates that it is due to an exchange of glucose units from the conjugate to Hb. The reaction is stereospecifically inhibited by p-nitrophenyl-beta-D-glucoside. The nature of the formed HbAlc is demonstrated by isoelectric focusing. A slight increase of HbAlc observed in the incubated controls may be due to an internal migration of some residues of glucose primitively bound to lysyl residues in an unstable form and also to some degree of denaturation during the incubation.
Desialylated erythrocytes, when perfused in mice liver in situ, were shown to be sequestered. This sequestration was inhibited by dihydrocytochalasin B and a neoglycoprotein: N-Acetylgalactosaminyl-albumin. Our results confirm that, under physiological conditions, the recognition of desialylated erythrocytes appear to be related to the presence of N-Acetylgalactosamine or galactose receptor on the surface of Küpffer cells.
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A population of peripheral blood lymphocytes exhibits specific receptor for the peanut lectin (Arachis hypogaea). In control subjects, this population represents 1.3% of all lymphocytes while it represents 13% in patients with alcoholic cirrhosis. The percentage of lymphocytes expressing this receptor appears to be correlated with hypoalbuminemia and the erythrocyte sedimentation rate, in these patients. Furthermore, a higher percentage of peanut positive lymphocytes was found in cirrhotic patients who subsequently died than in those who survived.
Young and old, human erythrocytes, separated in vitro according to their age in vivo, were radioactively labeled at the cell-surface D-galactosyl and 2-acetamido-2-deoxy-D-galactosyl residues by treatment with D-galactose oxidase, followed by reduction with sodium borotritide. The labeling was quantitatively determined for each type of erythrocyte by measuring the molar amounts of borohydride necessary for the complete reduction of the oxidized residues. The number of surface residues per blood-group A+ erythrocyte was found to be 37.6 +/- 1.8 x 10(6) (n = 8) for young, 21.8 +/- 4.9 x 10(6) (n = 8) for old, and 24.8 +/- 6.4 x 10(6) (n = 8) for middle-aged erythrocytes, indicating a significant decrease of the residues during ageing.
Membrane glycoproteins from young human erythrocytes and erythrocytes aged in vivo were fractionated by gel filtration. Three major groups of glycoproteins were obtained. The neutral hexoses and sialic acid contents of each group of glycoproteins from the old cells were found to be significantly reduced by comparison to the values found in the glycoproteins of the young cells. Thus, the previously observed decrease in neutral hexoses and in sialic acid contents of the full erythrocyte membrane during in vivo ageing does not affect only one particular group of glycoproteins but each of the groups of glycoproteins tested, including the major glycoproteins of the erythrocyte membrane, that is to say glycophorins. It is shown in addition, that the previously observed decrease per cell of the surface galactose and N-acetylgalactosamine residues of the ageing erythrocyte affect several groups of membrane glycoproteins including band 3, PAS-1, PAS-4 and PAS-3. The physiological significance of these experimental data is discussed.
The contents of fucose, mannose, galactose, glucose, 2-acetamido-2-deoxy-D-glucose and -D-galactose, and sialic acid, when the results were expressed as nmol per mg of membrane dry-weights, were found to be significantly lower in the membranes of old erythrocytes than in the membranes of young ones. No significant difference was found between young and old membranes when the compositions were expressed as residues per one hundred carbohydrate residues, suggesting that a homogeneous decrease of the carbohydrate moieties may occur during aging in vivo.
The chance discovery of two mouse strains, one with and one without a high presence of serum T-agglutinin, permitted the investigation of the role of this antibody in the disappearance of desialylated erythrocytes, which may be regarded as a model for ageing. The proportional relationship between the quantity of sialic acid removed and the diminution of half-life is not affected by the presence or absence of T-agglutinin. Opsonization by T-agglutinin would therefore appear to be an improbable mechanism. Other possible theories are discussed.
Evidence is presented to indicate a generalized role for the terminal sialic acid residues of circulating erythrocytes. After reinjection into their donors, neuraminidase-treated human, rabbit, rat and dog erythrocytes were promptly removed from the circulation : intect erythrocytes, previously incubated under the same conditions but without neuraminidase, were removed after a significantly longer period. The neuraminidase-treated erythrocytes were cleared by the liver and in a little part by the spleen. Old and young human, rabbit, rat erythrocytes contained different quantities of stromal sialic acid, significantly lowered on the old cells. But the half-life of old intact rabbit erythrocytes is sigificantly shorter than that of neuraminidase-treated young erythrocytes with a similar minidase-treated young erythrocytes with a similar sialic acid content. Indeed sialic acid is not the only carbohydrate component of the membrane that is decreased during erythrocyte ageing, the others membranous sugars are decreased too. Theses changes in the carbohydrate moity could have a role in the clearance of the erythrocytes.
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Rabbit as well duck erythrocytes desialization leads to a dramatic decrease of their survival time. Rabbit erythrocytes are trapped transiently by the liver ; in contrast splenic uptake is gradual. Serum from the two species contains a T-agglutinin directed against autologous desialyated erythrocytes. In the two cases, this T-agglutinin is from the IgM class and its immunodominant sugar is galactose in the beta configuration.