CD133 is a positive marker for a distinct class of primitive human cord blood-derived CD34-negative hematopoietic stem cells

Abstract

The identification of human CD34-negative (CD34 ⁻) hematopoietic stem cells (HSCs) provides a new concept for the hierarchy in the human HSC compartment. Previous studies demonstrated that CD34 ⁻ severe combined immunodeficiency (SCID)-repopulating cells (SRCs) are a distinct class of primitive HSCs in comparison to the well-characterized CD34 ⁺CD38 ⁻ SRCs. However, the purification level of rare CD34 ⁻ SRCs in 18 lineage marker-negative (Lin ⁻) CD34 ⁻ cells (1/1000) is still very low compared with that of CD34 ⁺CD38 ⁻ SRCs (1/40). As in the mouse, it will be necessary to identify useful positive markers for a high degree of purification of rare human CD34 ⁻ SRCs. Using 18Lin ⁻CD34 ⁻ cells, we analyzed the expression of candidate positive markers by flow cytometric analysis. We finally identified CD133 as a reliable positive marker of human CB-derived CD34 ⁻ SRCs and succeeded in highly purifying primitive human CD34 ⁻ HSCs. The limiting dilution analysis demonstrated that the incidence of CD34 ⁻ SRCs in 18Lin ⁻CD34 ⁻CD133 ⁺ cells was 1/142, which is the highest level of purification of these unique CD34 ⁻ HSCs to date. Furthermore, CD133 expression clearly segregated the SRC activities of 18Lin ⁻CD34 ⁻ cells, as well as 18Lin ⁻CD34 ⁺ cells, in their positive fractions, indicating its functional significance as a common cell surface maker to isolate effectively both CD34 ⁺ and CD34 ⁻ SRCs.