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J. Biol. Chem., Vol. 263, Issue 6, 2779-2786, 02, 1988
JA Badwey, JM Robinson, W Horn, RJ Soberman, MJ Karnovsky and ML Karnovsky
Neutrophils stimulated with optimal amounts of tumor-promoters that
activate protein kinase C (e.g. mezerein, phorbol 12,13-dibutyrate) are
known to release large quantities of superoxide: approximately 40-50 nmol
O2-/min/10(7) cells. Previous studies have shown that treatment of
neutrophils with the calcium ionophore A23187, or with 5-hydroxy-
6,8,11,14-eicosatetraenonate (5-HETE), dramatically increased the ability
of these cells to release O2- in response to suboptimal concentrations of
the stimulants mentioned. In this manuscript, we provide data relevant to
the basis of this augmentation of O2- release. The synergy with ionophore
A23187 exhibited a partial requirement for extracellular Ca2+, whereas that
with 5-HETE exhibited a near absolute requirement for that cation.
Neutrophils stimulated with optimal amounts of tumor-promoters are known to
exhibit a redistribution of protein kinase C activity from the soluble to a
particulate fraction. A redistribution of kinase activity was not observed
in cells stimulated synergistically. On the other hand, ionophore A23187
and 5-HETE increased the binding of a suboptimal amount of [3H] phorbol
12,13- dibutyrate to intact neutrophils by approximately 25 and 50%,
respectively. Inhibitors of protein kinase C (i.e. sphingosine, 1-(5-
isoquinolinylsulfonyl)-2-methylpiperazine) substantially blocked O-2
release from neutrophils stimulated either synergistically or with optimal
levels of tumor-promoters. These data suggest a role for 5-HETE in
modulating O-2 release by neutrophils and are discussed in relation to
models of the interactions of protein kinase C with membranes.
Synergistic stimulation of neutrophils. Possible involvement of 5- hydroxy-6,8,11,14-eicosatetraenoate in superoxide release
Department of Biological Chemistry, Harvard Medical School, Boston, Massachusetts 02115.
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