The effect of endogenous essential and nonessential fatty acids on the uptake and subsequent agonist-induced release of arachidonate

E. E. Furth, V. Hurtubise, M. A. Schott, Michael Laposata

Research output: Contribution to journalArticle

12 Citations (Scopus)

Abstract

We have demonstrated that the uptake and agonist-induced release of a pulse of arachidonate are influenced by the size and composition of preexisting endogenous fatty acid pools. EFD-1 cells, an essential fatty acid-deficient mouse fibrosarcoma cell line, were incubated with radiolabeled (14C or 3H) arachidonate, linoleate, eicosapentaenoate (EPA), palmitate, or oleate in concentrations of 0-33 μM for 24 h. After 24 h, the cells were pulsed with 0.67 μM radiolabeled (3H or 14C, opposite first label) arachidonate for 15 min and then stimulated with 10 μM bradykinin for 4 min. Because EFD-1 cells contain no endogenous essential fatty acids, we were able to create essential fatty acid-repleted cells for which the specific activity of the newly constructed endogenous essential fatty acid pool was known. Loading the endogenous pool with the essential fatty acids arachidonate, eicosapentaenoate, or linoleate (15-20 nmol of fatty acid incorporated/106 cells) decreased the uptake of a pulse of arachidonate from 200 to 100 pmol/106 cells but had no effect on palmitate uptake. The percent of arachidonate incorporated during the pulse which was released upon agonist stimulation increased 2-fold (4-8%) as the endogenous pool of essential fatty acids was increased from 0 to 15-20 nmol/106 cells. This 8% release was at least 3-fold greater than the percent release from the various endogenous essential fatty acid pools. In contrast, loading the endogenous pool with the nonessential fatty acids oleate or palmitate to more than 2-3 times their preexisting cellular level had no effect on the uptake of an arachidonate pulse. Like the essential fatty acids, increasing endogenous oleate increased (by 2-fold) the percent release of arachidonate incorporated during the pulse, whereas endogenous palmitate had no effect on subsequent agonist-induced release from this arachidonate pool. These studies show that preexisting pools of essential and nonessential fatty acids exert different effects on the uptake and subsequent releasability of a pulse of arachidonate.

Original languageEnglish (US)
Pages (from-to)18494-18501
Number of pages8
JournalJournal of Biological Chemistry
Volume264
Issue number31
StatePublished - 1989
Externally publishedYes

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Essential Fatty Acids
Fatty Acids
Palmitates
Oleic Acid
Linoleic Acid
Bradykinin
Fibrosarcoma
Labels
Cells
Cell Line
Chemical analysis

ASJC Scopus subject areas

  • Biochemistry

Cite this

The effect of endogenous essential and nonessential fatty acids on the uptake and subsequent agonist-induced release of arachidonate. / Furth, E. E.; Hurtubise, V.; Schott, M. A.; Laposata, Michael.

In: Journal of Biological Chemistry, Vol. 264, No. 31, 1989, p. 18494-18501.

Research output: Contribution to journalArticle

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abstract = "We have demonstrated that the uptake and agonist-induced release of a pulse of arachidonate are influenced by the size and composition of preexisting endogenous fatty acid pools. EFD-1 cells, an essential fatty acid-deficient mouse fibrosarcoma cell line, were incubated with radiolabeled (14C or 3H) arachidonate, linoleate, eicosapentaenoate (EPA), palmitate, or oleate in concentrations of 0-33 μM for 24 h. After 24 h, the cells were pulsed with 0.67 μM radiolabeled (3H or 14C, opposite first label) arachidonate for 15 min and then stimulated with 10 μM bradykinin for 4 min. Because EFD-1 cells contain no endogenous essential fatty acids, we were able to create essential fatty acid-repleted cells for which the specific activity of the newly constructed endogenous essential fatty acid pool was known. Loading the endogenous pool with the essential fatty acids arachidonate, eicosapentaenoate, or linoleate (15-20 nmol of fatty acid incorporated/106 cells) decreased the uptake of a pulse of arachidonate from 200 to 100 pmol/106 cells but had no effect on palmitate uptake. The percent of arachidonate incorporated during the pulse which was released upon agonist stimulation increased 2-fold (4-8{\%}) as the endogenous pool of essential fatty acids was increased from 0 to 15-20 nmol/106 cells. This 8{\%} release was at least 3-fold greater than the percent release from the various endogenous essential fatty acid pools. In contrast, loading the endogenous pool with the nonessential fatty acids oleate or palmitate to more than 2-3 times their preexisting cellular level had no effect on the uptake of an arachidonate pulse. Like the essential fatty acids, increasing endogenous oleate increased (by 2-fold) the percent release of arachidonate incorporated during the pulse, whereas endogenous palmitate had no effect on subsequent agonist-induced release from this arachidonate pool. These studies show that preexisting pools of essential and nonessential fatty acids exert different effects on the uptake and subsequent releasability of a pulse of arachidonate.",
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N2 - We have demonstrated that the uptake and agonist-induced release of a pulse of arachidonate are influenced by the size and composition of preexisting endogenous fatty acid pools. EFD-1 cells, an essential fatty acid-deficient mouse fibrosarcoma cell line, were incubated with radiolabeled (14C or 3H) arachidonate, linoleate, eicosapentaenoate (EPA), palmitate, or oleate in concentrations of 0-33 μM for 24 h. After 24 h, the cells were pulsed with 0.67 μM radiolabeled (3H or 14C, opposite first label) arachidonate for 15 min and then stimulated with 10 μM bradykinin for 4 min. Because EFD-1 cells contain no endogenous essential fatty acids, we were able to create essential fatty acid-repleted cells for which the specific activity of the newly constructed endogenous essential fatty acid pool was known. Loading the endogenous pool with the essential fatty acids arachidonate, eicosapentaenoate, or linoleate (15-20 nmol of fatty acid incorporated/106 cells) decreased the uptake of a pulse of arachidonate from 200 to 100 pmol/106 cells but had no effect on palmitate uptake. The percent of arachidonate incorporated during the pulse which was released upon agonist stimulation increased 2-fold (4-8%) as the endogenous pool of essential fatty acids was increased from 0 to 15-20 nmol/106 cells. This 8% release was at least 3-fold greater than the percent release from the various endogenous essential fatty acid pools. In contrast, loading the endogenous pool with the nonessential fatty acids oleate or palmitate to more than 2-3 times their preexisting cellular level had no effect on the uptake of an arachidonate pulse. Like the essential fatty acids, increasing endogenous oleate increased (by 2-fold) the percent release of arachidonate incorporated during the pulse, whereas endogenous palmitate had no effect on subsequent agonist-induced release from this arachidonate pool. These studies show that preexisting pools of essential and nonessential fatty acids exert different effects on the uptake and subsequent releasability of a pulse of arachidonate.

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