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Flow cytofluorimetric analysis of young and senescent human erythrocytes probed with lectins. Evidence that sialic acids control their life span

  • Glycoconjagate Journal
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Abstract

Comparing the properties of ‘young’ and senescent (‘aged’) O+ erythrocytes isolated by applying ultracentrifugation in a self-forming Percoll gradient, we demonstrate that the sialic acids of membrane glycoconjugates control the life span of erythrocytes and that the desialylation of glycans is responsible for the clearance of the aged erythrocytes. This capture is mediated by a β-galactolectin present in the membrane of macrophages. The evidence supporting these conclusions is as follows:

  1. (1)

    Analysis by flow cytofluorimetry of the binding of fluorescein isothiocyanate labelled lectins specific for sialic acids shows that the aged erythrocytes bind less WGA, LPA, SNA and MAA than young erythrocytes. The binding of DSA and LCA is not modified. On the contrary, the number of binding sites of UEA-I specific for O antigen and of AAA decreases significantly. PNA and GNA do not bind to erythrocytes.

  2. (2)

    RCA120 as well asErythrina cristagalli andErythrina corallodendron lectins specific for terminal β-galactose residues lead to unexpected and unexplained results with a decrease in the number of lectin binding sites associated with increasing desialylation.

  3. (3)

    The glycoconjugates from the old erythrocytes incorporate more sialic acid than the young cells. This observation results from the determination of the rate of transfer by α-2,6-sialyltransferase of fluorescent or radioactiveN-acetylneuraminic acid, using as donors CMP-9-fluoresceinyl-NeuAc and CMP-[14C]-NeuAc, respectively.

  4. (4)

    Microscopy shows that the old erythrocytes are captured preferentially by the macrophages relative to the young ones. Fixation of erythrocytes by the macrophage membrane is inhibited by lactose, thus demonstrating the involvement of a terminal β-galactose specific macrophage lectin.

  5. (5)

    Comparative study of the binding of WGA, LPA, SNA and MAA to the aged erythrocytes and to thein vitro enzymatically desialylated erythrocytes shows that the desialylation rate of aged cells is low but sufficient to lead to their capture by the macrophages

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Abbreviations

BSA:

bovine serum albumin

CMP-NeuAc:

cytidine monophosphateN-acetylneuraminate

CSB:

cell sialylation buffer

EDTA:

ethylene diamine tetraacetic acid

FITC:

fluoresceinyl isothiocyanate

9-FITC-NeuAc:

9-fluoresceinyl-N-acetylneuraminate

NeuAc:

N-acetylneuraminic acid

PAGE:

polyacrylamide gel electrophoresis

PBS:

Dulbecco's phosphate buffer saline solution

PMSF:

phenylmethyl-sulfonyl fluoride

RBC:

red blood cells

SCA:

Senescent Cell Antigen

SDS:

sodium dodecyl sulfate

SFG:

senescent factor glycopeptides

AAA:

Aleuria aurantia agglutinin

DSA:

Datura stramonium agglutinin

ECA:

Erythrina cristagalli agglutinin

GNA:

Galanthus nivalis agglutinin

LCA:

Lens culinaris agglutinin

LFA:

Limax flavus agglutinin

LPA:

Limulus polyphemus agglutinin

MAA:

Maackia amurensis agglutinin

PNA:

peanut agglutinin

RCA:

Ricinus communis agglutinin

SNA:

Sambucus nigra agglutinin

UEA-I:

Ulex europeus agglutinin-I

WGA:

Wheat germ agglutinin

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Authors and Affiliations

  1. Institute of Biochemistry of the Romanian Academy, Bucharest, Romania

    Daniela Bratosin, Magdalena Moisei & Cecilia Motas

  2. Laboratoire de Chimie Biologique (UMR 111 du CNRS), Université des Sciences et Technologies de Lille, Villeneuve d'Ascq, France

    Joel Mazurier, Henri Debray, Myriam Lecocq, Catherine Alonso & Jean Montreuil

  3. Laboratoire de Biologie du Développement, Université des Sciences et Technologies de Lille, Villeneuve d'Ascq, France

    Benoni Boilly

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  1. Daniela Bratosin
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  2. Joel Mazurier
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  7. Magdalena Moisei
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Bratosin, D., Mazurier, J., Debray, H. et al. Flow cytofluorimetric analysis of young and senescent human erythrocytes probed with lectins. Evidence that sialic acids control their life span. Glycoconjugate J 12, 258–267 (1995). https://doi.org/10.1007/BF00731328

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  • DOI: https://doi.org/10.1007/BF00731328

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