Dye efflux studies suggest that hematopoietic stem cells expressing low or undetectable levels of CD34 antigen exist in multiple species

Abstract

We previously described a method for isolating murine hematopoietic stem cells capable of reconstituting lethally irradiated recipients, which depends solely on dual-wavelength flow cytometric analysis of murine bone marrow cells stained with the fluorescent DNA-binding dye Hoechst 33342. This method, which appears to rely on the differential ability of stem cells to efflux the Hoechst dye, defines an extremely small and homogeneous population of cells (termed SP cells). We show here that dual-wavelength analysis of Hoechst dye-stained human, rhesus and miniature swine bone marrow cells reveals a small, distinct population of cells that efflux the dye in a manner identical to murine SP cells. Like the murine SP cells, both human and rhesus SP cells are primarily CD34-negative and lineage marker-negative. In vitro culture studies demonstrated that rhesus SP cells are highly enriched for long-term culture-initiating cells (LTC-ICs), an indicator of primitive hematopoietic cells, and have the capacity for differentiation into T cells. Although rhesus SP cells do not initially possess any hematopoietic colony-forming capability, they acquire the ability to form colonies after long-term culture on bone marrow stroma, coincident with their conversion to a CD34-positive phenotype. These studies suggest the existence of a hitherto unrecognized population of hematopoietic stem cells that lack the CD34 surface marker classically associated with primitive hematopoietic cells.

Transforming growth factor-β decreases the cancer-initiating cell population within diffuse-type gastric carcinoma cells

“…In this study, we provide evidence that CICs are present in diffuse-type gastric carcinoma cells, and that the SP cells in OCUM-2MLN cells possessed repopulating capacity and high tumorforming ability in vivo. SP cells were initially identified in mouse bone marrow as long-term repopulating cells (Goodell et al, 1997). This original discovery was followed by other reports on a wide variety of tissues (Zhou et al, 2001).…”

Niche regulation of corneal epithelial stem cells at the limbus

“…Based on the principle that many adult somatic SCs preferentially express Bcrp1/ABCG2, a member of ATPbinding cassette transporters, they can be isolated as the side population (SP) using FACS via the unique property of effluxing Hoechst 33342 dye [77,78] (for review see [79]). Using this method, SP cells have been isolated from human [23][24][25], rat [80], and rabbit [24,81,82] limbal tissues.…”

Efficient infection of primitive hematopoietic stem cells by modified adenovirus

“…15 Twenty-four hours after infection by Ad5/F35, the mononuclear bone marrow cells were stained using Hoechst dyes, and analyzed by FACS to verify that Ad5/F35 could infect SP cells and induce them to express GFP. Figure 4 confirms that Ad5/F35 infects 65% of the bone marrow cells determined by GFP expression 24 h after infection.…”