Dual SP/ALDH Functionalities Refine the Human Hematopoietic Lin−CD34+CD38− Stem/Progenitor Cell Compartment

Abstract

Identification of prevalent specific markers is crucial to stem/ progenitor cell purification. Determinants such as the surface antigens CD34 and CD38 are traditionally used to analyze and purify hematopoietic stem/progenitor cells (HSCs/HPCs). However, the variable expression of these membrane antigens poses some limitations to their use in HSC/HPC purification. Techniques based on drug/stain efflux through the ATP-binding cassette (ABC)G2 pump (side population [SP] phenotype) or on detection of aldehyde dehydrogenase (ALDH) activity have been independently developed and distinguish the SP and ALDH Bright (ALDH Br ) cell subsets for their phenotype and proliferative capability. In this study, we developed a multiparametric flow cytometric method associating both SP and ALDH activities on human lineage negative (Lin 2 ) bone marrow cells and sorted different cell fractions according to their SP/ALDH activity level. We find that Lin 2 CD34 1 CD38 Low/2 cells are found throughout the spectrum of ALDH expression and are enriched especially in ALDH Br cells when associated with SP functionality (SP/ALDH Br fraction). Furthermore, the SP marker identified G 0 cells in all ALDH fractions, allowing us to sort quiescent cells regardless of ALDH activity. Moreover, we show that, within the Lin 2 CD34 1 CD38 2 ALDH Br population, the SP marker identifies cells with higher primitive characteristics, in terms of stemness-related gene expression and in vitro and in vivo proliferative potential, than the Lin 2 CD34 1 CD38 2 ALDH Br main population cells. In conclusion, our study shows that the coexpression of SP and ALDH markers refines the Lin 2 CD34 1 CD38 2 hematopoietic compartment and identifies an SP/ALDH Br cell subset enriched in quiescent primitive HSCs/HPCs.

Ovarian Cancer Stem Cells Are Enriched in Side Population and Aldehyde Dehydrogenase Bright Overlapping Population

“…SP cells and ALDH Br cells show greater efflux of Hoechst 33342 dye and higher expression level of aldehyde dehydrogenese, which are different phenotypes, and hematopoietic stem cells were isolated as an SP and ALDH Br population in a previous study [24]. We therefore investigated the overlapping population of SP cells and ALDH Br cells.…”

Flow cytometry in cancer stem cell analysis and separation

“…Thus, protocols have to take the correct order into account (140). Pierre-Louis et al have described this in detail (141), proposing staining the SP cells first and then performing the ALDH1 staining. Further analysis of additional phenotype markers should be performed in a final step (Fig.…”

Further phenotypic characterization of the primitive lineage− CD34+CD38−CD90+CD45RA− hematopoietic stem cell/progenitor cell sub-population isolated from cord blood, mobilized peripheral blood and patients with chronic myelogenous leukemia

“…3 Over the past 40 years or so, the enrichment of HSCs on the basis of their physical (for example, cell size, density), cell surface antigen, and biochemical characteristics (for example, elevated levels of aldehyde dehydrogenase) has been intensely pursued by a multitude of investigators including ourselves. 4, 5, 6, 7, 8, 9, 10, 11, 12 Whereas such efforts have culminated in the ability to purify the HSC to near homogeneity from mouse bone marrow and fetal liver, 13, 14 the same cannot be said for human HSCs. However, Irving Weissman and colleagues 15 , who have made major contributions in the area of HSC purification and characterization over the years, have moved us ever closer toward that goal.…”