Calcium ionophore but not phorbol ester promotes eicosanoids release by proliferating interleukin-3-dependent bone marrow cells.
Eicosanoid release during multilineage hematopoiesis was assessed using freshly isolated mouse bone marrow cells cultured in the presence of interleukin-3 (IL-3) (10% WEHI-3 culture-conditioned medium). Cells that could release prostaglandin E2 (PGE2) when stimulated with calcium ionophore A23187, but not with phorbol ester (PMA), appeared within 4 days. The cells harvested on day 10 released 42 ng of PGE2/10(6) cells/mL after A23187 stimulation. Leukotriene B4 (LTB4) (4 ng/mL) was also detected after A23187 stimulation, but there was no detectable LTC4 (less than 0.5 ng/mL). Nonadherent bone marrow cells were isolated from 28-day cultures and cloned. All clones were strongly IL-3-dependent. Although other growth factors such as granulocyte colony-stimulating factor (G-CSF), granulocyte-macrophage CSF (GM-CSF), and CSF-1 failed to promote survival or support proliferation of the cells, three clones (11-1-A6, 3-2-D5, and 11-1-A1) showed significant increases in 3H-thymidine incorporation, respectively, after PMA treatment for 24 hours. Surviving cells displayed dominantly myeloid type morphology and phenotypic characteristics. The data suggest that IL-3 is important in the formation of PGE2-producing cells. In contrast to many macrophages (MO), neither the IL-3-dependent cell lines nor the IL-3-cultured bone marrow cells released significant amounts of PGE2 when stimulated with PMA or IL-3, although PMA and IL-3 both induced translocation of protein kinase C (PKC) to the membrane fraction. The lack of production of PGE2 and other eicosanoids by the PMA- and IL-3-stimulated cell lines was confirmed by measuring the release of 3H-arachidonic acid. The data suggest that in IL-3-dependent bone marrow cell lines the activation of eicosanoid metabolism requires elevated cellular Ca2+; PKC activation alone does not appear to be a sufficient stimulus.
Shibata
Y
Y
Fujisaki Cell Center, Hayashibara Biochemical Laboratories Inc., Okayama, Japan.
McCaffrey
P G
PG
Sato
H
H
Oghiso
Y
Y
D000818:Animals; D016718:Arachidonic Acid; D001095:Arachidonic Acids / Q000378:metabolism; D001854:Bone Marrow Cells; D000001:Calcimycin / Q000494:pharmacology*; D002118:Calcium / Q000494:pharmacology; D002455:Cell Division; D002999:Clone Cells; D004247:DNA / Q000096:biosynthesis; D015232:Dinoprostone / Q000378:metabolism*; D005260:Female; D006412:Hematopoietic Stem Cells / Q000166:cytology / Q000378:metabolism*; D007377:Interleukin-3 / Q000494:pharmacology*; D007975:Leukotriene B4 / Q000378:metabolism*; D016173:Macrophage Colony-Stimulating Factor / Q000494:pharmacology; D008264:Macrophages / Q000378:metabolism; D051379:Mice; D008808:Mice, Inbred CBA; D011493:Protein Kinase C / Q000378:metabolism; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*
D001095:Arachidonic Acids; D007377:Interleukin-3; D007975:Leukotriene B4; D016718:Arachidonic Acid; D000001:Calcimycin; D016173:Macrophage Colony-Stimulating Factor; D004247:DNA; D011493:Protein Kinase C; D015232:Dinoprostone; D013755:Tetradecanoylphorbol Acetate; D002118:Calcium