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The Journal of Immunology, Vol 151, Issue 7 3775-3784, Copyright © 1993 by American Association of Immunologists
ARTICLES |
YA Mekori, CK Oh and DD Metcalfe
Mast Cell Physiology Section, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD 20892.
It is well established that mast cell proliferation and maturation are regulated by two principle cytokines, IL-3 and the c-kit ligand stem cell factor (SCF). Little is known, however, how these two processes are negatively regulated and thus, how mast cell number is controlled in normal or pathologic processes. In this study we hypothesized that IL-3-dependent mast cells would undergo programmed cell death (apoptosis) on removal of IL-3 as was shown with other growth factor- dependent hemopoietic cells. Apoptotic changes were analyzed using light microscopy, fluorescent staining with acridine orange, flow cytometric analysis, and DNA electrophoresis. We could demonstrate that elimination of IL-3 from either primary bone marrow-derived cultured mast cell cultures (BMCMC) or from the growth factor-dependent mast cell line MCP5 resulted in the characteristic changes of apoptosis including condensed chromatin, fragmented nuclei, cellular vacuolization, typical pattern of propidium iodide or Hoechst 33342 uptake by flow cytometry, and the characteristic 200 bp "ladder" pattern of DNA cleavage. These events were prevented by SCF, an action that was in part mediated by tyrosine kinases, in that the tyrosine kinase inhibitor herbimycin inhibited the action of SCF in preventing apoptosis in IL-3-deprived cells. By using anti c-kit mAb and IL-3- dependent BMCMC obtained from W/Wv mice homozygous for mutation at the w locus that encodes the c-kit receptor, we could also show that SCF exerted its effect via c-kit. Neither dexamethasone nor cyclosporin A inhibited the "rescue" effect of SCF, suggesting that "rescue" was mediated by SCF and not through the induction of other cytokines. Thus, IL-3-dependent mast cells undergo apoptosis on removal of IL-3, an event that is prevented by the addition of SCF through its ligand c- kit, thus demonstrating how these principle mast cell growth factors may act in concert to regulate mast cell number under physiologic conditions.
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