title: Hematopoietic differentiation of umbilical cord blood-derived very small embryonic/epiblast-like stem cells.
abstract: A population of CD133(+)Lin(-)CD45(-) very small embryonic/epiblast-like stem cells (VSELs) has been purified by multiparameter sorting from umbilical cord blood (UCB). To speed up isolation of these cells, we employed anti-CD133-conjugated paramagnetic beads followed by staining with Aldefluor to detect aldehyde dehydrogenase (ALDH) activity; we subsequently sorted CD45(-)/GlyA(-)/CD133(+)/ALDH(high) and CD45(-)/GlyA(-)/CD133(+)/ALDH(low) cells, which are enriched for VSELs, and CD45(+)/GlyA /CD133(+)/ALDH(high) and CD45(+)/GlyA(-)/CD133(+)/ALDH(low) cells, which are enriched for hematopoietic stem/progenitor cells (HSPCs). Although freshly isolated CD45(-) VSELs did not grow hematopoietic colonies, the same cells, when activated/expanded over OP9 stromal support, acquired hematopoietic potential and grew colonies composed of CD45(+) hematopoietic cells in methylcellulose cultures. We also observed that CD45(-)/GlyA(-)/CD133(+)/ALDH(high) VSELs grew colonies earlier than CD45(-)/GlyA(-)/CD133(+)/ALDH(low) VSELs, which suggests that the latter cells need more time to acquire hematopoietic commitment. In support of this possibility, real-time polymerase chain reaction analysis confirmed that, whereas freshly isolated CD45(-)/GlyA(-)/CD133(+)/ALDH(high) VSELs express more hematopoietic transcripts (for example, c-myb), CD45(-)/GlyA(-)/CD133(+)/ALDH(low) VSELs exhibit higher levels of pluripotent stem cell markers (for example, Oct-4). More importantly, hematopoietic cells derived from VSELs that were co-cultured over OP9 support were able to establish human lympho-hematopoietic chimerism in lethally irradiated non-obese diabetic/severe combined immunodeficiency mice 4-6 weeks after transplantation. Overall, our data suggest that UCB-VSELs correspond to the most primitive population of HSPCs in UCB.
authors.affiliation: Stem Cell Institute at the James Graham Brown Cancer Center, University of Louisville, Louisville, KY 40202, USA. mzrata01@louisville.edu
authors.forename: J
authors.initials: J
authors.lastname: Ratajczak
authors.forename: E
authors.initials: E
authors.lastname: Zuba-Surma
authors.forename: I
authors.initials: I
authors.lastname: Klich
authors.forename: R
authors.initials: R
authors.lastname: Liu
authors.forename: M
authors.initials: M
authors.lastname: Wysoczynski
authors.forename: N
authors.initials: N
authors.lastname: Greco
authors.forename: M
authors.initials: M
authors.lastname: Kucia
authors.forename: M J
authors.initials: MJ
authors.lastname: Laughlin
authors.forename: M Z
authors.initials: MZ
authors.lastname: Ratajczak
mesh_terms: D000071916:AC133 Antigen; D000444:Aldehyde Dehydrogenase / Q000378:metabolism; D000818:Animals; D015703:Antigens, CD / Q000032:analysis; D002454:Cell Differentiation; D053595:Embryonic Stem Cells / Q000166:cytology*; D005312:Fetal Blood / Q000166:cytology*; D006023:Glycoproteins / Q000032:analysis; D006412:Hematopoietic Stem Cells / Q000166:cytology*; D006801:Humans; D017493:Leukocyte Common Antigens / Q000032:analysis; D051379:Mice; D016513:Mice, SCID; D010455:Peptides / Q000032:analysis
chemical_list: D000071916:AC133 Antigen; D015703:Antigens, CD; D006023:Glycoproteins; C000606023:PROM1 protein, human; D010455:Peptides; C000606024:Prom1 protein, mouse; D000444:Aldehyde Dehydrogenase; D017493:Leukocyte Common Antigens; C516769:PTPRC protein, human