Isolation of small, primitive human hematopoietic stem cells: distribution
of cell surface cytokine receptors and growth in SCID-Hu mice
JE Wagner, D Collins, S Fuller, LR Schain, AE Berson, C Almici, MA Hall, KE Chen, TB Okarma and JS Lebkowski
Department of Pediatrics, University of Minnesota, Minneapolis, USA.
Human CD34+ cells were subfractionated into three size classes using
counterflow centrifugal elutriation followed by immunoadsorption to
polystyrene cell separation devices. The three CD34+ cell fractions (Fr),
Fr 25/29, Fr 33/37, and Fr RO, had mean sizes of 8.5, 9.3 and 13.5 microns,
respectively. The majority of cells in the large Fr RO CD34+ cell
population expressed the committed stage antigens CD33, CD19, CD38, or
HLA-DR and contained the majority of granulocyte- macrophage colony-forming
units (CFU-GM), burst-forming units-erythroid (BFU-E), and CFU-mixed
lineage (GEMM). In contrast, the small Fr 25/29 CD34+ cells were devoid of
committed cell surface antigens and lacked colony-forming activity. When
seeded to allogeneic stroma, Fr RO CD34+ cells produced few CFU-GM at week
5, whereas cells from the Fr 25/29 CD34+ cell population showed a 30- to
55-fold expansion of myeloid progenitors at this same time point.
Furthermore, CD34+ cells from each size fraction supported ontogeny of T
cells in human thymus/liver grafts in severe combined immunodeficient
(SCID) mice. Upon cell cycle analyses, greater than 97% of the Fr 25/29
CD34+ cells were in G0/G1 phase, whereas greater proportions of the two
larger CD34+ cell fractions were in active cell cycle. Binding of the
cytokines interleukin (IL)-1 alpha, IL-3, IL-6, stem cell factor (SCF),
macrophage inhibitory protein (MIP)-1 alpha, granulocyte colony-
stimulating factor (G-CSF), and granulocyte-macrophage (GM)-CSF to these
CD34+ cell populations was also analyzed by flow cytometry. As compared
with the larger CD34+ cell fractions, cells in the small Fr 25/29 CD34+
cell population possessed the highest numbers of receptors for SCF, MIP1
alpha, and IL-1 alpha. Collectively, these results indicate that the Fr
25/29 CD34+ cell is a very primitive, quiescent progenitor cell population
possessing a high number of receptors for SCF and MIP1 alpha and capable of
yielding both myeloid and lymphoid lineages when placed in appropriate in
vitro or in vivo culture conditions.
Volume 86,
Issue 2,
pp. 512-523,
07/15/1995
Copyright © 1995 by The American Society of Hematology
