Enrichment and Characterization of Murine Hematopoietic Stem Cells That Express c-kit Molecule

The proto-oncogene c-kit encodes a transmembrane tyrosine kinase receptor for stem cell factor (SCF). The c-kit/SCF signal is expected to have an important role in hematopoiesis. A monoclonal antibody (ACK-2) against the murine c-kit molecule was prepared. Flow cytometric analysis showed that the bo...

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Bibliographic Details
Published in:Blood 1991-10, Vol.78 (7), p.1706-1712
Main Authors: Okada, Seiji, Nakauchi, Hiromitsu, Nagayoshi, Kazunari, Nishikawa, Satomi, Nishikawa, Shin-ichi, Miura, Yasusada, Suda, Toshio
Format: Article
Language:English
Subjects:
Animals
Antibodies, Monoclonal
Biological and medical sciences
Bone Marrow - radiation effects
Bone Marrow Cells
Bone Marrow Transplantation
Cell physiology
Cells, Cultured
Colony-Forming Units Assay
Fundamental and applied biological sciences. Psychology
Hematopoiesis
Hematopoietic Stem Cell Transplantation
Hematopoietic Stem Cells - metabolism
Mice
Mice, Inbred C57BL
Molecular and cellular biology
Proto-Oncogene Proteins - analysis
Proto-Oncogene Proteins - immunology
Proto-Oncogene Proteins - physiology
Proto-Oncogene Proteins c-kit
Spleen - cytology
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Summary:The proto-oncogene c-kit encodes a transmembrane tyrosine kinase receptor for stem cell factor (SCF). The c-kit/SCF signal is expected to have an important role in hematopoiesis. A monoclonal antibody (ACK-2) against the murine c-kit molecule was prepared. Flow cytometric analysis showed that the bone marrow cells that expressed the c-kit molecule (approximately 5%) were B220(B)−, TER119(erythroid)−, Thy1negative-low, and WGA+. A small number of Mac-1(macro-phage)+ or Gr-1(granulocyte)+ cells were c-kit-low positive. Colony-forming unit in culture (CFU-C) and day-8 and day-12 CFU-spleen (CFU-S) existed exclusively in the c-kit-positive fraction. About 20% of the Lin(lineage)− c-kit+ cells were rhodamine-123low and this fraction contained more day-12 CFU-S than day-8 CFU-S. On the basis of these findings, murine hematopoietic stem cells were enriched with normal bone marrow cells. One of two and one of four Thy-1lowLin− WGA+c-kit+ cells were CFU-C and CFU-S, respectively. Long-term repopulating ability was investigated using B6/Ly5 congenic mice. Eight and 25 weeks after transplantation of Lin−c-kit+ cells, donor-derived cells were found in the bone marrow, spleen, thymus, and peripheral blood. In peripheral blood, T cells, B cells, and granulocyte-macrophages were derived from donor cells. Injection of ACK-2 into the irradiated mice after bone marrow transplantation decreased the numbers of day-8 and day-12 CFU-S in a dose-dependent manner. Day-8 spleen colony formation was completely suppressed by the injection of 100 μg ACK-2, but a small number of day-12 colonies were spared. Our data show that the c-kit molecule is expressed in primitive stem cells and plays an essential role in the early stages of hematopoiesis.
ISSN:0006-4971
1528-0020
DOI:10.1182/blood.V78.7.1706.1706