Opposing effects of PI3 kinase pathway activation on human myeloid and erythroid progenitor cell proliferation and differentiation in vitro.
OBJECTIVE
To investigate 1) the effects of lineage-specific cytokines (G-CSF and EPO) combined with ligands for different classes of cytokine receptors (common beta chain, gp130, and tyrosine kinase) on proliferation by human myeloid and erythroid progenitor cells; and 2) the signal transduction pathways associated with combinatorial cytokine actions.


METHODS
CFU-GM and BFU-E were cloned in vitro. Secondary colony formation by replated CFU-GM and subcolony formation by BFU-E provided measures of progenitor cell proliferation. Studies were performed in the presence of cytokine combinations with and without signal transduction inhibitors.


RESULTS
Proliferation by CFU-GM and BFU-E was enhanced synergistically when common beta chain receptor cytokines (IL-3 or GM-CSF) were combined with G-CSF or EPO, but not with gp130 receptor cytokines (LIF or IL-6) or tyrosine kinase receptor cytokines (SCF, HGF, Flt-3 ligand, or PDGF). Delayed addition studies with G-CSF+IL-3 and EPO+IL-3 demonstrated that synergy required the presence of both cytokines from the initiation of the culture. The Jak2-specific inhibitor, AG490, abrogated the effect of combining IL-3 with EPO but had no effect on the enhanced CFU-GM proliferation stimulated by IL-3+G-CSF. The PI3 kinase inhibitors LY294002 and wortmannin substituted for G-CSF in combination with IL-3 since proliferation in the presence of LY294002/wortmannin+IL-3 was enhanced to the same extent as in the presence of G-CSF+IL-3. In contrast, LY294002 and wortmannin inhibited proliferation in the presence of EPO and in the presence of EPO+IL-3.


CONCLUSIONS
1) IL-3 may activate different signal transduction pathways when combined with G-CSF and when combined with EPO; 2) different signal transducing intermediates regulate erythroid and myeloid progenitor cell proliferation; and 3) inhibition of the PI3 kinase pathway suppresses myeloid progenitor cell differentiation and thereby increases proliferation.
Lewis
John L
JL
LRF Centre for Adult Leukaemia, Department of Hematology, Faculty of Medicine, Imperial College, London, England, UK.
Marley
Stephen B
SB
Ojo
Morenike
M
Gordon
Myrtle Y
MY
D002454:Cell Differentiation / Q000187:drug effects; D002455:Cell Division / Q000187:drug effects; D002478:Cells, Cultured; D002867:Chromones / Q000494:pharmacology; D004789:Enzyme Activation; D015672:Erythroid Precursor Cells / Q000187:drug effects / Q000502:physiology*; D004921:Erythropoietin / Q000494:pharmacology; D016179:Granulocyte Colony-Stimulating Factor / Q000494:pharmacology; D006412:Hematopoietic Stem Cells / Q000187:drug effects / Q000502:physiology*; D006801:Humans; D007377:Interleukin-3 / Q000494:pharmacology; D053614:Janus Kinase 2; D020928:Mitogen-Activated Protein Kinases / Q000502:physiology; D009025:Morpholines / Q000494:pharmacology; D019869:Phosphatidylinositol 3-Kinases / Q000502:physiology*; D011505:Protein-Tyrosine Kinases / Q000502:physiology; D011518:Proto-Oncogene Proteins; D015398:Signal Transduction
D002867:Chromones; D007377:Interleukin-3; D009025:Morpholines; D011518:Proto-Oncogene Proteins; D004921:Erythropoietin; D016179:Granulocyte Colony-Stimulating Factor; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; D019869:Phosphatidylinositol 3-Kinases; D011505:Protein-Tyrosine Kinases; C507924:JAK2 protein, human; D053614:Janus Kinase 2; D020928:Mitogen-Activated Protein Kinases