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Blood, Vol. 94 No. 4 (August 15), 1999:
pp. 1261-1272
From the Department of Haematology, University College London Medical
School, London, UK.
The activation of phospholipase A2 (PLA2)
with release of eicosanoids and prostanoids in mature myeloid cells and
the augmentation (priming) of this activity by cytokines such as
granulocyte-macrophage colony-stimulating factor (GM-CSF) are central
to the inflammatory process. Yet, there are few data concerning
PLA2 activity and its regulation by growth factors in
primary hematopoietic cells. We therefore analyzed the PLA2
activity of mobilized human CD34 antigen-positive (CD34+)
stem cells by quantitation of the extracellular release of
3H-arachidonate. The PLA2 activity of
CD34+ cells stimulated with calcium ionophore (A23187)
was of similar magnitude to that of mature neutrophils and monocytes.
Preincubation of CD34+ cells with stem cell factor (SCF)
before A23187-stimulation resulted in primed PLA2 activity,
whereas interleukin-3 (IL-3), GM-CSF, and tumor necrosis factor This article has been cited by other articles:
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| Copyright © 1999 by American Society of Hematology Online ISSN: 1528-0020 | |||||||||
had
no significant effect. When CD34+ cells were induced to
differentiate, PLA2 activity remained responsive to SCF for
several days, but after 8 days, at which stage morphological and
functional evidence of maturation was occurring, priming of PLA2 by SCF could no longer be elicited, whereas responses
to GM-CSF and IL-3 had developed. The further metabolism of arachidonic acid to eicosanoids by CD34+ cells was not detected by
either thin-layer chromatography, enzyme immunoassay, or differential
spectroscopy. SCF stimulated the rapid but transient activation of ERK2
(p42 MAP kinase) in CD34+ cells, and we used the MAP
kinase kinase inhibitor, PD 098059, which at 30 µmol/L blocks ERK2
activation in CD34+ cells, to investigate whether
SCF-mediated priming of arachidonate release was mediated by this
kinase. PD 098059 only partially inhibited A23187-stimulated
PLA2 activity primed by SCF, suggesting the involvement of
ERK2 and possibly a further signal transduction pathway. Methyl
arachidonyl fluorophosphonate (5 µmol/L), a dual inhibitor of i and
cPLA2 isoforms, completely inhibited arachidonate release
without affecting ERK2 activation, demonstrating the lack of cellular
toxicity. These data provide the first evidence that primitive myeloid
cells have the capacity to release arachidonate, which is regulated by
an early acting hematopoietic growth factor important for the growth
and survival of these cells.
