Separation of hematopoietic stem cells into two populations and their characterization

Abstract

Two populations of hematopoietic stem cells (HSC) in mouse bone marrow (BM) are defined on the basis of the presence or absence of interleukin- 3 (IL-3) receptor-associated antigen (IL-3RAA). HSC were purified by depletion of mature lymphoid-lineage cells followed by collection of the low-density fraction and sorting of wheat germ agglutinin-binding (WGA+) cells using a fluorescein-activated cell sorter. WGA+ cells were further separated into two populations (IL-3RAA+/WGA+ and IL-3RAA- /WGA+) by a monoclonal antibody (MoAb) against IL-3RAA. IL-3RAA+/WGA+ cells formed CFU-S on day 8; this population consisted mainly of cells in the cycling phase. IL-3RAA-/WGA+ cells form CFU-S on day 12; this population consisted mainly of dormant cells (cells in the G0 phase). When two populations obtained from C3H/HeN mice were injected into lethally irradiated (C57BL/6 x C3H/HeN)F1 mice, donor-derived cells in the peripheral blood (PB) appeared significantly earlier in mice injected with IL-3RAA+/WGA+ cells than in those injected with IL-3RAA- /WGA+ cells, whereas the reconstruction efficiency of IL-3RAA-/WGA+ cells had overtaken that of IL-3RAA+/WGA+ cells 6 weeks after injection. Long-term observation showed no significant difference between these two populations, however. The radioprotective ability (RPA) (30-day survival) of these two populations was therefore compared. The RPA of IL-3RAA-/WGA+ cells was significantly higher than that of IL-3RAA+/WGA+ cells. These findings therefore suggest that the former population is more primitive.

Characterization of Natural Suppressor Cells in Human Bone Marrow

“…We have found that NS cells in mouse and monkey BM have low density, a high affinity (more cell surface receptor) to wheat germ agglutinin [14,24], and the receptor for interleukin 3 [16]. From these results, it is anticipated that NS cells are hematopoietic progenitors committed to myeloid lineage [30,31]. In the present study, of cells that lack lineage markers, those with relatively low density (BM Fr.…”

Changes in Markers, Receptors and Adhesion Molecules Expressed on Murine Hemopoietic Stem Cells After a Single Injection of 5‐Fluorouracil

“…We have previously shown that IL-3Rprogenitor cells are more primitive than IL-3R + cells [4]. In the present study, we examined the expression of IL-6R on CD71 -H-2 high cells.…”

“…An effective method of purifying pluripotent hemopoietic stem cells (P-HSC) would contribute not only to clarifying the mechanism behind hemopoietic differentiation and selfrenewal of P-HSC, but also to the use of P-HSC in bone marrow transplantation (BMT). Until now, a number of laboratories have reported different purification procedures, including those based on physical characteristics (size and cell density), negative selection to remove lineage-committed cells and positive selection using various markers [1][2][3][4][5][6][7][8][9]. Visser et al purified stem cells using density centrifugation followed by positive selection, utilizing both binding ability to wheat germ agglutinin (WGA) and anti-H-2KAb with a cell sorter [2].…”

“…We have previously reported that interleukin 3 receptor-negative (IL-3R -) progenitor cells are more primitive than IL-3R + cells [4]. It has also been reported that the transferrin receptor (CD71) is expressed on proliferating committed progenitors but not on noncycling immature progenitors [19][20][21], and that P-HSC express a high level of major histocompatibility complex (MHC) class I but not class II molecules [22][23][24].…”

Tumor necrosis factor alpha cooperates with interleukin 3 in the recruitment of a primitive subset of human CD34+ progenitors.

“…In contrast, more committed progenitors, responsive to either IL-3 or GM-CSF, were not affected by TNF-o~ (2). In this context, another study has recently demonstrated that IL-3 receptor-negative progenitors are more primitive than those expressing IL-3 receptors (18). Finally, these in vitro observations, which identify TNF-ot as an important molecule for the recruitment of early hematopoietic progen-itors are supported by the demonstration of the radioprotective effect of TNF-ol in vivo (19).…”