5′ nucleotidase activity of rainbow trout oncorhynchus mykiss macrophages: Correlation with respiratory burst activity

F. W. Hepkema, C. J. Secombes

Research output: Contribution to journalArticle

3 Citations (Scopus)

Abstract

A novel method for analysing 5′ nucleotidase activity of rainbow trout, Oncorhynchus mykiss, head kidney macrophages was optimised for use with cell monolayers, with respect to the effect of cell numbers, temperature and substrate concentration. Both lysed and whole cells could be used for determination of 5′ nucleotidase activity, and = 105 whole cells or = 104 lysed cells were required. Maximal 5′ nucleotidase activity was found in the range 27-33°C and using a substrate concentration of ≥ 1·5 μmol AMP ml-1 for whole cells and ≥1·5 μmol AMP ml-1 for lysed cells. 5′ Nucleotidase activity was also correlated with respiratory burst activity in cells treated with a variety of supernatants containing macrophage activating factor (MAF) activity. A significant inverse relationship between these two activities was found. MAF-treated cells were also found to lose 5′ nucleotidase activity faster than control cells in the presence of cycloheximide, suggesting such cells may have a higher membrane turnover of this enzyme.

Original languageEnglish
Pages (from-to)301-309
Number of pages9
JournalFish and Shellfish Immunology
Volume4
Issue number4
DOIs
Publication statusPublished - Jul 1994

Fingerprint

5'-nucleotidase
5'-Nucleotidase
Macrophages
rainbow
Oncorhynchus mykiss
macrophages
substrate
Macrophage-Activating Factors
Adenosine Monophosphate
turnover
cells
enzyme
membrane
Substrates
Cycloheximide
temperature
Monolayers
Cells
Membranes
Enzymes

Keywords

  • 5′ nucleotidase
  • Macrophages
  • Maf
  • Rainbow trout

ASJC Scopus subject areas

  • Environmental Chemistry
  • Aquatic Science

Cite this

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title = "5′ nucleotidase activity of rainbow trout oncorhynchus mykiss macrophages: Correlation with respiratory burst activity",
abstract = "A novel method for analysing 5′ nucleotidase activity of rainbow trout, Oncorhynchus mykiss, head kidney macrophages was optimised for use with cell monolayers, with respect to the effect of cell numbers, temperature and substrate concentration. Both lysed and whole cells could be used for determination of 5′ nucleotidase activity, and = 105 whole cells or = 104 lysed cells were required. Maximal 5′ nucleotidase activity was found in the range 27-33°C and using a substrate concentration of ≥ 1·5 μmol AMP ml-1 for whole cells and ≥1·5 μmol AMP ml-1 for lysed cells. 5′ Nucleotidase activity was also correlated with respiratory burst activity in cells treated with a variety of supernatants containing macrophage activating factor (MAF) activity. A significant inverse relationship between these two activities was found. MAF-treated cells were also found to lose 5′ nucleotidase activity faster than control cells in the presence of cycloheximide, suggesting such cells may have a higher membrane turnover of this enzyme.",
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N2 - A novel method for analysing 5′ nucleotidase activity of rainbow trout, Oncorhynchus mykiss, head kidney macrophages was optimised for use with cell monolayers, with respect to the effect of cell numbers, temperature and substrate concentration. Both lysed and whole cells could be used for determination of 5′ nucleotidase activity, and = 105 whole cells or = 104 lysed cells were required. Maximal 5′ nucleotidase activity was found in the range 27-33°C and using a substrate concentration of ≥ 1·5 μmol AMP ml-1 for whole cells and ≥1·5 μmol AMP ml-1 for lysed cells. 5′ Nucleotidase activity was also correlated with respiratory burst activity in cells treated with a variety of supernatants containing macrophage activating factor (MAF) activity. A significant inverse relationship between these two activities was found. MAF-treated cells were also found to lose 5′ nucleotidase activity faster than control cells in the presence of cycloheximide, suggesting such cells may have a higher membrane turnover of this enzyme.

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