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High levels of aldehyde dehydrogenase (ALDH) activity have been proposed to be a common feature of stem cells. Adult hematopoietic, neural, and cancer stem cells have all been reported to have high ALDH activity, detected using Aldefluor, a fluorogenic substrate for ALDH. This activity has been attributed to Aldh1a1, an enzyme that is expressed at high levels in stem cells and that has been suggested to regulate stem cell function. Nonetheless, Aldh1a1 function in stem cells has never been tested genetically. We observed that Aldh1a1 was preferentially expressed in mouse hematopoietic stem cells (HSCs) and expression increased with age. Hematopoietic cells from Aldh1a1-deficient mice exhibited increased sensitivity to cyclophosphamide in a non–cell-autonomous manner, consistent with its role in cyclophosphamide metabolism in the liver. However, Aldh1a1 deficiency did not affect hematopoiesis, HSC function, or the capacity to reconstitute irradiated recipients in young or old adult mice. Aldh1a1 deficiency also did not affect Aldefluor staining of hematopoietic cells. Finally, Aldh1a1 deficiency did not affect the function of stem cells from the adult central or peripheral nervous systems. Aldh1a1 is not a critical regulator of adult stem cell function or Aldefluor staining in mice.

Hematopoietic stem cells (HSCs) and endothelial progenitors arise from a common embryonic precursor. However, these populations diverge prior to the onset of definitive hematopoiesis, as HSCs become CD45+ and are thought to lose the expression of endothelial markers. After the onset of definitive hematopoiesis, CD144 (vascular endothelial [VE]–cadherin) has been considered a specific marker of endothelial cells. In contrast, we found that virtually all HSC activity from embryonic day 13.5 (E13.5) fetal liver was CD144+. CD144 expression declined on E16.5 fetal liver HSCs and was absent from adult bone marrow HSCs. This identified a new marker that is differentially expressed between fetal and adult HSCs, and enhanced the purification of HSCs from the E13.5 fetal liver. These results emphasize the close developmental relationship between hematopoietic and endothelial cells, while indicating that CD144 is not a specific marker of endothelial cells during fetal development.

Hematopoietic stem cells (HSCs) are a self-renewing population of bone marrow cells that replenish the cellular elements of blood throughout life. HSCs represent a paradigm for the study of stem-cell biology, because robust methods for prospective isolation of HSCs have facilitated rigorous characterization of these cells. Recently, a new isolation method was reported, using the SLAM family of cell-surface markers, including CD150 (SlamF1), to offer potential advantages over established protocols. We examined the overlap between SLAM family member expression with an established isolation scheme based on Hoechst dye efflux (side population; SP) in conjunction with canonical HSC cell-surface markers (Sca-1, c-Kit, and lineage markers). Importantly, we find that stringent gating of SLAM markers is essential to achieving purity in HSC isolation and that the inclusion of canonical HSC markers in the SLAM scheme can greatly augment HSC purity. Furthermore, we observe that both CD150+ and CD150− cells can be found within the SP population and that both populations can contribute to long-term multilineage reconstitution. Thus, using SLAM family markers to isolate HSCs excludes a substantial fraction of the marrow HSC compartment. Interestingly, these 2 subpopulations are functionally distinct, with respect to lineage output as well as proliferative status.