A number of recombinant cytokines have been shown to regulate the kinetics of the primitive hemopoietic progenitors. While expansion has been documented for hemopoietic cells, it is not yet known whether hemopoietic stem cells with long-term engrafting ability are expandable in culture. This proposal is designed to seek fundamental information regarding in vitro expansion of hemopoietic stem cells and progenitors. We will carry out parallel studies with murine and human hemopoietic cells harvested at different ontogenic stages. Suspension culture of FACS-sorted primitive progenitors will be carried out in the presence of combinations of early-acing cytokines under serum-containing and serum- free conditions.
Specific Aim #1 is to test the whether it is possible to expand murine hemopoietic stem cells in suspension culture. We will use a PCR-based assay for the identification of transplanted male cells.
Specific Aim #2 is to test whether human hemopoietic stem cells with long-term engrafting ability are expandable in culture. We will use in utero transplantation into pre-immune sheep as an assay for human stem cells.
Specific Aim #3 is to delineate the cytokine interactions supporting the optimal expansion of progenitors representing different developmental stages. We will use cell culture assays as well as in vivo engraftment assays for defining the expanded progenitor populations. An important aim here is to use the resulting information to guide the in vivo experiments described in Specific Aims # 1 and #2. Another major interest is to correlate the progenitors assayed in culture with the in vivo short-term and long-term engrafting progenitors in order to develop culture assay predictive of in vivo functions of the progenitors. We recently observed that IL-3 and IL-1 independently suppress early stages of B- and T-lymphopoiesis. We will test whether the inhibitor effects of IL-3 and IL-1 involve early states of all lymphohematopoiesis including self-renewal ability of the stem cells.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
5R01DK048714-04
Application #
2458854
Study Section
Diabetes, Endocrinology and Metabolic Diseases B Subcommittee (DDK)
Program Officer
Badman, David G
Project Start
1994-08-01
Project End
1998-11-30
Budget Start
1997-08-01
Budget End
1998-11-30
Support Year
4
Fiscal Year
1997
Total Cost
Indirect Cost
Name
Medical University of South Carolina
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
183710748
City
Charleston
State
SC
Country
United States
Zip Code
29425
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Ogawa, M (1999) Stochastic model revisited. Int J Hematol 69:2-5
Ogawa, M; Matsunaga, T (1999) Humoral regulation of hematopoietic stem cells. Ann N Y Acad Sci 872:17-23;discussion 23-4
Hirayama, F; Aiba, Y; Ikebuchi, K et al. (1999) Differentiation in culture of murine primitive lymphohematopoietic progenitors toward T-cell lineage. Blood 93:4187-95
Matsunaga, T; Kato, T; Miyazaki, H et al. (1998) Thrombopoietin promotes the survival of murine hematopoietic long-term reconstituting cells: comparison with the effects of FLT3/FLK-2 ligand and interleukin-6. Blood 92:452-61
Matsunaga, T; Hirayama, F; Yonemura, Y et al. (1998) Negative regulation by interleukin-3 (IL-3) of mouse early B-cell progenitors and stem cells in culture: transduction of the negative signals by betac and betaIL-3 proteins of IL-3 receptor and absence of negative regulation by granulocyte-macrophage colo Blood 92:901-7
Shimizu, Y; Ogawa, M; Kobayashi, M et al. (1998) Engraftment of cultured human hematopoietic cells in sheep. Blood 91:3688-92
Ogawa, M; Yonemura, Y; Ku, H (1997) In vitro expansion of hematopoietic stem cells. Stem Cells 15 Suppl 1:7-11; discussion 12
Aiba, Y; Hirayama, F; Ogawa, M (1997) Clonal proliferation and cytokine requirement of murine progenitors for natural killer cells. Blood 89:4005-12
Aiba, Y; Ogawa, M (1997) Development of natural killer cells, B lymphocytes, macrophages, and mast cells from single hematopoietic progenitors in culture of murine fetal liver cells. Blood 90:3923-30

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