Expression of CD34 in Hematopoietic Cancer Cell Lines Reflects Tightly Regulated Stem/Progenitor-Like State Klaudia Kuranda, 1,2 Ce ´line Berthon, 1,2,3 Fre ´de ´ric Lepre ˆtre, 4,5 Renata Polakowska, 1 Nathalie Jouy, 4 and Bruno Quesnel 1,2,3 * 1 INSERM, Unite ´ 837, Lille, France 2 Institut de Recherche sur le Cancer de Lille, Lille, France 3 Service des Maladies du Sang, Centre Hospitalier et Universitaire de Lille, Lille, France 4 Universite ´ Nord de France, Institut Fe ´de ´ratif de Recherche 114, Lille, France 5 Plateforme de ge ´nomique fonctionnelle, Institut Fe ´de ´ratif de Recherche 114, Lille, France ABSTRACT Hematopoietic cancer stem cells preserve cellular hierarchy in a manner similar to normal stem cells, yet the underlying regulatory mechanisms are poorly understood. It is known that both normal and malignant stem/progenitor cells express CD34. Here, we demonstrate that several cell lines (HL-60, U266) derived from hematopoietic malignancies contain not only CD34  but also CD34 þ subpopulations. The CD34 þ cells displayed a stem/progenitor-like phenotype since, in contrast to CD34  cells, they frequently underwent cellular division and rapidly formed colonies in methylcellulose-based medium. Strikingly, a constant fraction of the CD34 þ and CD34  cell subpopulations, when separated, rapidly switched their phenotype. Consequently, both separated fractions could generate tumors in immunocompromised NOD/ LtSz-scid/scid mice. Cultures in vitro showed that the proportion of CD34 þ stem/progenitor-like cells in the population was decreased by cell– cell contact and increased by soluble factors secreted by the cells. Using cytokine arrays, we identified some of these factors, notably thymopoietin that was able to increase the proportion of CD34 þ cells and overall colony-forming capacity in tested cell lines. This action of thymopoietin was conserved in mononuclear cells from bone marrow. Therefore, we propose that hematopoietic cancer cell lines containing subpopulations of CD34 þ cells can provide an in vitro model for studies of cancer stem/progenitor cells. J. Cell. Biochem. 112: 1277–1285, 2011. ß 2011 Wiley-Liss, Inc. KEY WORDS: CANCER STEM CELLS; PROGENITORS; HEMATOPOIESIS; CD34 PROTEIN; THYMOPOIETIN I n humans, hematopoietic stem/progenitor cells express CD34, a cell-surface protein, that is gradually lost during differentiation [reviewed in Krause et al., 1996]. Quiescent CD34  stem cells [Osawa et al., 1996; Bhatia et al., 1998; Wang et al., 2003] give rise to actively cycling CD34 þ stem/progenitor cells, which in turn give rise to differentiated CD34  cells of various blood lineages [Zanjani et al., 1998; Nakamura et al., 1999; Sato et al., 1999; Wang et al., 2003; Dooley and Oppenlander, 2004]. It has been suggested that hematopoietic stem cells that become malignant and are at the origin of malignant clones preserve cellular hierarchy in a manner similar to normal cells [Sutherland et al., 1996; Bonnet and Dick, 1997]. The regulation of this hierarchy is not fully understood, mainly because bone marrow of patients contains both normal and malignant hematopoietic stem cells that cannot currently be distinguished by simple staining of cell surface markers. This problem could be overcome by the use of models such as cancer cell lines. Despite their acquired ability to grow in vitro, cancer cell lines exhibit genetic and molecular features of the primary cancers from which they were derived. Moreover, if we assume that cell lines can contain stem cells, they could only contain cancer stem cells, as their normal equivalents differentiate rapidly in culture conditions and stop proliferating. Journal of Cellular Biochemistry ARTICLE Journal of Cellular Biochemistry 112:1277–1285 (2011) 1277 Additional Supporting Information may be found in the online version of this article. Grant sponsor: Ligue Nationale Contre le Cancer, Equipes Labellise ´es, (Comite ´ du Septentrion); Grant sponsor: Institut National du Cancer (Program: IMMUDORM); Grant sponsor: Institut de Recherche sur le Cancer de Lille (K. Kuranda); Grant sponsor: Re ´gion Nord-Pas de Calais. *Correspondence to: Bruno Quesnel, Service des Maladies du Sang, et Unite ´ INSERM 837 Ho ˆ pital Huriez, CHU de Lille 59037, Lille, France. E-mail: brunoquesnel@hotmail.com Received 4 January 2011; Accepted 6 January 2011  DOI 10.1002/jcb.23026  ß 2011 Wiley-Liss, Inc. Published online 18 January 2011 in Wiley Online Library (wileyonlinelibrary.com).