Cytotechnology

, Volume 16, Issue 2, pp 89–100

Significant inhibition of hybridoma cells by exogenous application of ganglioside GM3, a possible modulator of cell growthin vitro

  • Heike Brandt
  • Johannes Müthing
  • Jasna Peter-Katalinić
  • Jürgen Lehmann
Article

DOI: 10.1007/BF00754611

Cite this article as:
Brandt, H., Müthing, J., Peter-Katalinić, J. et al. Cytotechnology (1994) 16: 89. doi:10.1007/BF00754611

Abstract

Gangliosides of the mouse-rat hybridoma cell line 187.1, which secretes an antibody against ϰ-light chain of mouse IgG, were isolated and structurally characterized by biochemical and immunological methods (overlay technique), and fast atom bombardment-mass spectrometry. Exclusively GM3, substituted with C24∶1 and C16∶0 fatty acid and C18∶1 sphingosine, was found in this B cell derived cell line. A GM3 (NeuGc) to GM3(NeuAc) ratio (80 to 20), was characteristic for 187.1 cells, and absolute GM3 amounts of about 0.3 mg 10−9 viable cells were determined. Exogenous application of GM3, which has been isolated from large cell preparations, to 187.1 cells showed growth inhibition in a concentration dependent manner. Using the MTT-assay and the [3H]thymidine incorporation assay, the cells exhibited a strong reduction in metabolic and proliferative activity, respectively, after exposure of cells to GM3. GM3 was applied in concentrations between 3μM and 30μM, giving evidence for strong inhibitory effects at 30μM GM3 and less but significant suppression after application of GM3 concentrations lower than 20μM. No cellular response was observed at the lowest concentration (3μM) used in this study. Hybridoma cells as well as other cell types like fibroblasts, muscle cells and endothelial cells, are in general characterized by high expression of the GM3 ganglioside, which is known to act as a modulator of cellular growth in monolayer cultures of adherent cells. Since gangliosides are released to the culture medium by cell lysis, i.e. cell death, and/or by active membrane shedding, the results obtained in this study suggest a growth regulatory role of GM3 in high density hybridoma cell cultures.

Key words

Hybridoma growthinhibitiongangliosidesGM3

Abbreviations

DMB

1,2-diamino-4,5-methylenedioxybenzene

FAB-MS

fast atom bombardment-mass spectrometry

GSL(s)

glycosphingolipid(s)

HPLC

high performance liquid chromatography

HPTLC

high performance thin layer chromatography

MTT

3,(4,5 dimethylthiazol-2-yl)2,5 diphenyl tetrazolium bromide

NeuAc

N-acetylneuraminic acid

NeuGc

N-glycolylneuraminic acid

PBS

phosphate buffered saline

Copyright information

© Kluwer Academic Publishers 1994

Authors and Affiliations

  • Heike Brandt
    • 1
  • Johannes Müthing
    • 1
  • Jasna Peter-Katalinić
    • 2
  • Jürgen Lehmann
    • 1
  1. 1.Institute for Cell Culture TechnologyUniversity of BielefeldBielefeldGermany
  2. 2.Institute for Physiological ChemsitryUniversity of BonnGermany