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Preferential reduction of the α-2-6-sialylation from cell surface N-glycans of human diploid fibroblastic cells by in vitro aging

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Abstract

Human diploid fibroblastic cell line, TIG-3, has a finite life span of about 80 population doubling levels (PDL), and is used for in vitro aging studies. Young cells (PDL 23) grew to higher cell densities at a higher growth rate than aged cells (PDL 77). When the electrophoretic mobility of cells was determined, the negative surface charge of the aged cells decreased significantly when compared to that of young cells. Lectin blot analysis of membrane glycoproteins showed that the α-2-6-sialylation but not the α-2-3-sialylation of N-glycans decreases markedly in the aged cells when compared to the young cells. In support of this observation, the cDNA microarray assay and reverse transcription-polymerase chain reaction (RT-PCR) analysis showed that the gene expression of the α-2,6-sialyltransferase I (ST6Gal I), which transfers sialic acid to galactose residues of N-glycans, decreases in the aged cells. These results indicate that the concordant decrease of the α-2,6-sialylation of N-glycans with the ST6Gal I gene expression is induced in TIG-3 cells by in vitro aging.

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Abbreviations

CBB:

Coomassie Brilliant Blue

Con A:

concanavalin A

DT:

doubling time

EtBr:

ethidium bromide

β-1,4-GalT:

β-1,4-galactosyltransferase

GlcNAcT II:

UDP-GlcNAc

Manα:

1,6-Man β-1,2-N-acetyl-glucosaminyltransferase

G3PDH:

glyceraldehyde 3-phosphate dehydrogenase

FCS:

fetal calf serum

LCA:

Lens culinaris agglutinin

L-PHA:

leuko-agglutinating phytohemagglutinin

MAA:

Maackia amurensis agglutinin

MES:

2-(N-morpholino)ethansulfonic acid

PBS:

phosphate-buffered saline

PDL:

population doubling levels

PVL:

Psathyrella velutina lectin

RCA-I:

Ricinus communis agglutinin-I

RT-PCR:

reverse transcription polymerase chain reaction

SNA:

Sambucus nigra agglutinin

ST3Gal III:

β-galactoside α-2,3-sialyltransferase III

ST3Gal IV:

β-galactoside α-2,3-sialytransferase IV

ST6Gal I:

β-galactoside α-2,6-sialyltransferase I

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Authors and Affiliations

  1. Department of Biosignal Research, Tokyo Metropolitan Institute of Gerontology, Itabashi-ku, Tokyo, 173-0015, Japan

    Tomomi Tadokoro, Kiyotaka Yamamoto, Iku Kuwahara, Takeshi Sato & Kiyoshi Furukawa

  2. Department of Applied Biological Science, Faculty of Science and Technology, Tokyo University of Science, Noda, 278-8510, Japan

    Tomomi Tadokoro & Masahiko Ikekita

  3. Faculty of Education, Institute of Natural Sciences, Saitama University, Sakura-ku, Saitama, 338-8570, Japan

    Iku Kuwahara & Hirosuke Fujisawa

  4. Biomaterials Center, National Institute For Materials Science, Tsukuba, 305-0044, Japan

    Akiyoshi Taniguchi

  5. Laboratory of Glycobiology, Nagaoka University of Technology, Nagaoka, 940-2188, Japan

    Takeshi Sato & Kiyoshi Furukawa

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  1. Tomomi Tadokoro
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  2. Kiyotaka Yamamoto
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Tadokoro, T., Yamamoto, K., Kuwahara, I. et al. Preferential reduction of the α-2-6-sialylation from cell surface N-glycans of human diploid fibroblastic cells by in vitro aging. Glycoconj J 23, 443–452 (2006). https://doi.org/10.1007/s10719-006-7152-y

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  • DOI: https://doi.org/10.1007/s10719-006-7152-y

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