Submitted on October 6, 2006
Accepted on January 12, 2007
Severe hypoxia defines heterogeneity and selects highly immature progenitors within clonal erythroleukemia cells
Serena Giuntoli 1,
Elisabetta Rovida 2,
Antonella Gozzini 3,
Valentina Barbetti 1,
Maria Grazia Cipolleschi 1,
Massimo Olivotto 1,
Persio Dello Sbarba 1*
1 Dipartimento di Patologia e Oncologia Sperimentali della Università degli Studi di Firenze, Firenze, Italy
2 Dipartimento di Patologia e Oncologia Dipartimento di Patologia e Oncologia Sperimentali della Università degli Studi di Firenze, Firenze, Italydella Università degli Studi di Firenze
3 Divisione di Ematologia, AUO Careggi, Firenze, Italy
* To whom correspondence should be addressed. E-mail: persio{at}unifi.it.
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Abstract |
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We previously showed that resistance to severe hypoxia defines hierarchical levels within normal hematopoietic populations and that hypoxia modulates the balance between generation of progenitors and maintenance of stem cells (HSC) in favour of the latter. This study deals with the effects of hypoxia (0.1% oxygen) in vitro on Friend's murine erythroleukemia (MEL) cells, addressing the question whether a clonal leukemia cell population comprise functionally different cell subsets characterized by different hypoxia-resistance. To identify leukemia stem cells (LSC), we used the Culture Repopulating Ability (CRA) assay we developed to quantify in vitro stem cells capable of short-term reconstitution (STR).
Hypoxia strongly inhibited the overall growth of MEL cell population, that, despite its clonality, resulted to comprise progenitors characterized by markedly different hypoxia-resistance. These include hypoxia-sensitive colony-forming cells and hypoxia-resistant STR-type LSC, capable of repopulating secondary liquid cultures of CRA assays, confirmig what previously shown for normal hematopoiesis. STR-type LSC were found capable of not only surviving in hypoxia, but also being mostly in cycle, in contrast with the fact that almost all hypoxia-surviving cells were growth-arrested and with what previously found for HSC. However, quiescent LSC were also detected, capable of delayed culture repopulation with the same efficiency as STR-like LSC. The fact that even quiescent LSC, believed to sustain minimal residual disease in vivo, were found within the MEL cells indicates that all main components of leukemia cell populations may be present within clonal cell lines, which are therefore suitable to study the sensitivity of individual components to treatments.
Key Words.
leukemia stem cells, severe hypoxia, culture-repopulating ability, 5FU-resistance
