The goals of the research program are to characterize hematopoietic growth factors which influence the proliferation of early human progenitors, both normal and neoplastic. Four areas will be emphasized. First, we will continue studies of the regulation of growth factors release form mesenchymal cells. This will include Northern analyses of the kinds of growth factors released from cells such as lymphocytes, fibroblasts, endothelial cells, and smooth muscle cells. Probes for human granulocyte /macrophage (GM), granulocyte (G), macrophage (M) and multi- (interleukin-3) colony-stimulating factors (CSFs) will be employed. The effects of various mediators of immune and inflammatory responses will be studied and will include interleukin1, lipoppolysaccharide, tumor necrosis factor, lymphotoxin, and thrombin. Relative transcription rates of the genes for the CSFs will be measured to determine if they are differentially expressed by the various mediators of inflammation. Second, we will explore the structure/function relationship of human GM- and G-CSF at the protein and molecular levels. For studies at the protein level, we will generate monoclonal antibodies to human GM- and G-CSF by purifying them from mammalian cells transfected with cDNA for these factors. At the molecular level, sitespecific mutagenesis will be carried out to determine which element of the gene are important for biological function of the intact protein. Third, we will study the effect of purified recombinant GM- and G-CSF, alone and together, on progenitor cells obtained from the peripheral blood and marrow of normal volunteers and patients with myeloproliferative disorders. To assess the nature of the interaction of the growth factors with the progenitor cells, the cells will be grown at low concentrations under serumfree conditions. Fourth, we will seek to clone the cDNA for a potentially novel growth factor produced by feline leukemia virusinfected embtyonic cat fibroblasts. The strategy will involve subtraction hybridization to enrich mRNA for the growth factor, generation of a cDNA library in an expression vector, and identification of useful clones by biological assays.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
5R01CA031615-07
Application #
3169705
Study Section
Hematology Subcommittee 2 (HEM)
Project Start
1982-04-01
Project End
1992-06-30
Budget Start
1988-07-01
Budget End
1989-06-30
Support Year
7
Fiscal Year
1988
Total Cost
Indirect Cost
Name
University of Washington
Department
Type
Schools of Medicine
DUNS #
135646524
City
Seattle
State
WA
Country
United States
Zip Code
98195
Hitchcock, Ian S; Chen, Maximus M; King, Jennifer R et al. (2008) YRRL motifs in the cytoplasmic domain of the thrombopoietin receptor regulate receptor internalization and degradation. Blood 112:2222-31
Barroga, Charlene F; Pham, Hang; Kaushansky, Kenneth (2008) Thrombopoietin regulates c-Myb expression by modulating micro RNA 150 expression. Exp Hematol 36:1585-92
Yoshida, Kozue; Kirito, Keita; Yongzhen, Hu et al. (2008) Thrombopoietin (TPO) regulates HIF-1alpha levels through generation of mitochondrial reactive oxygen species. Int J Hematol 88:43-51
Nakao, Takafumi; Geddis, Amy E; Fox, Norma E et al. (2008) PI3K/Akt/FOXO3a pathway contributes to thrombopoietin-induced proliferation of primary megakaryocytes in vitro and in vivo via modulation of p27(Kip1). Cell Cycle 7:257-66
Kaushansky, Kenneth (2008) Historical review: megakaryopoiesis and thrombopoiesis. Blood 111:981-6
McIntosh, Bryan; Kaushansky, Kenneth (2008) Transcriptional regulation of bone marrow thrombopoietin by platelet proteins. Exp Hematol 36:799-806
Coleman, Thomas R; Westenfelder, Christof; Togel, Florian E et al. (2006) Cytoprotective doses of erythropoietin or carbamylated erythropoietin have markedly different procoagulant and vasoactive activities. Proc Natl Acad Sci U S A 103:5965-70
Chanprasert, Supantitra; Geddis, Amy E; Barroga, Charlene et al. (2006) Thrombopoietin (TPO) induces c-myc expression through a PI3K- and MAPK-dependent pathway that is not mediated by Akt, PKCzeta or mTOR in TPO-dependent cell lines and primary megakaryocytes. Cell Signal 18:1212-8
Kaushansky, Kenneth (2006) Hematopoietic growth factors, signaling and the chronic myeloproliferative disorders. Cytokine Growth Factor Rev 17:423-30
Geddis, Amy E; Fox, Norma E; Kaushansky, Kenneth (2006) The Mpl receptor expressed on endothelial cells does not contribute significantly to the regulation of circulating thrombopoietin levels. Exp Hematol 34:82-6

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