Control of blood cell production involves cytokine-cell interactions in which cytokines manifest positive and negative effects on hematopoietic stem and progenitor cells. Growth factors such as steel factor (SLF) and a colony-stimulating factor (CSF) can act together to elicit synergistic effects. Synergistic stimulation can be suppressed by chemokines such as macrophage inflammatory protein (MIP)- 1alpha, interferon inducible protein (IP)-10, IL-8 and platelet factor (PF)4; chemokines can act together to synergistically suppress growth. It is our hypothesis that distinct and overlapping pathways exist within a cell to mediate synergistic stimulation of cell proliferation by multiple cytokines and suppression by chemokines is occurring at intracellular sites of growth factor synergistically stimulated events. Our goal is to evaluate these pathways using factor-dependent myeloid cell lines, with verification where feasible with primary cells.
The specific aims are: 1) continue investigation of the role of the Raf-1/MEK-1/MAP-kinase pathway as a mediator of the synergistic stimulation of cell proliferation induced by SLF plus a CSF, and of the c-AMP-dependent protein kinase A inhibitory activity on Raf-1 activation as a site of the negative effects of MIP-1alpha and IP-10 on MEKK1/Sekl/SAP-kinase, and MKK3/Hog/MAP-APK2- kinase, as possible sites for synergistically-induced stimulation of cell proliferation, and its negative regulation by chemokines. Determine a role for the synergistically-induced up-regulation of immediate response genes such as c-fos, junB, c-egr and c-myc as possible sites of negative regulation by chemokines. Evaluate the above as possible sites for synergistically-induced suppression of cell proliferation by combinations of low concentrations of any two of the following chemokines: MIP-1alpha, IP-10, IL-8 or PF4. 2) Establish a role for the cyclin-dependent kinase inhibitors, p21CIP-1 and p27KIP-1 in synergistically-induced stimulation of cell proliferation and determine if this is a site of chemokine suppression of cell growth. 3) Evaluate a role for eukaryotic initiation factor 4E (eIF4E) in the greatly enhanced level of protein synthesis seen in cells synergistically stimulated to proliferate by SLF and a CSF and in the suppression of this by MIP-1alpha, IP-10, IL-8 and PF4. 4) Evaluate roles for the protein tyrosine phosphatases Syp and PRP-1C in synergistically-induced stimulation of cell proliferation and its suppression by chemokine family members.
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