Published online 14 January 2009
Haematologica, Vol 94, Issue 2, 185-194 doi:10.3324/haematol.13206
Copyright © 2009 by Ferrata Storti Foundation
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Hematopoietic Stem Cells

Ex vivo expansion of hematopoietic progenitor cells is associated with downregulation of {alpha}4 integrin- and CXCR4-mediated engraftment in NOD/SCID β2-microglobulin-null mice

Jacques Foguenne, Ivano Di Stefano, Olivier Giet, Yves Beguin, André Gothot

GIGA-Research, Hematology Unit, University of Liège, Belgium

Correspondence: André Gothot, M.D., University of Liège, Laboratory Hematology CHU Sart Tilman B35 13, avenue de l’Hôpital B-4000 Liège, Belgium E-mail:agothot{at}ulg.ac.be

Background: Several studies indicate that ex vivo cytokine-supported expansion induces defective hematopoietic stem cell engraftment. We investigated the role of {alpha}4 integrin, {alpha}5 integrin and CXCR4 in engraftment of unmanipulated and cytokine-treated human cord blood CD34+ cells.

Design and Methods: Uncultured or expanded CD34+ cells were infused in NOD/SCID-β2microglobulin-null mice. The function of {alpha}4, and {alpha}5 integrins and CXCR4 was assessed by incubating cells with specific neutralizing antibodies, prior to transplant. The activation state of {alpha}4 integrin was further tested by adhesion and migration assays.

Results: Neutralization of either {alpha}4 integrin or CXCR4 abolished engraftment of uncultured CD34+ cells at 6 week spost-transplant, while {alpha}5 integrin neutralization had no significant effect. However, after short-term ex vivo culture, blocking {alpha}4 integrin or CXCR4 did not affect repopulating activity whereas neutralization of {alpha}5 integrin inhibited engraftment. Using soluble vascular cell adhesion molecule-1 binding assays, we observed that {alpha}4 integrin affinity in fresh CD34+ cells was low and susceptible to stimulation while in cultured CD34+ cells, it was high and insensitive to further activation. In addition, stromal cell-derived factor-1 stimulated migration across vascular cell adhesion molecule-1 in fresh CD34+ cells but not in cultured CD34+ cells.

Conclusions: Our data show that ex vivo culture of hematopoietic progenitor cells is associated with downregulation of both {alpha}4 integrin- and CXCR4-mediated engraftment. Further investigations suggest that this is caused by supraphysiological increase of {alpha}4 integrin affinity, which impairs directional migration across vascular cell adhesion molecule-1 in response to stromal cell-derived factor-1. Such changes may underlie the engraftment defect of cytokine-stimulated CD34+ cells.

Key words: stem cells, hematopoiesis, cell trafficking.