Abstract
Expansion of human hematopoietic stem cells (HSCs) is a major challenge in cellular therapy, and currently relies on the use of recombinant cytokines or on gene transfer of transcription factors. Of these, the HOXB4 homeoprotein protein is of particular interests as it promotes the expansion of mouse HSCs without inducing the development of leukemia. To eliminate any deleterious effects that might be associated with stable HOXB4 gene transfer into human cells, we took advantage of the ability of HOX proteins to passively translocate through cell membranes. Here we show that when cultured on stromal cells genetically engineered to secrete HOXB4, human long-term culture-initiating cells (LTC-ICs) and nonobese diabetic–severe combined immunodeficiency (NOD-SCID) mouse repopulating cells (SRCs) were expanded by more than 20- and 2.5-fold, respectively, over their input numbers. This expansion was associated with enhanced stem cell repopulating capacity in vivo and maintenance of pluripotentiality. This method provides a basis for developing cell therapy strategies using expanded HSCs that are not genetically modified.
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References
Weissman, I.L., Anderson, D.J. & Gage, F. Stem and progenitor cells: origins, phenotypes, lineage commitments, and transdifferentiations. Annu. Rev. Cell. Dev. Biol. 17, 387– 403 (2001).
Sauvageau, G. et al. Overexpression of HOXB4 in hematopoietic cells causes the selective expansion of more primitive populations in vitro and in vivo. Genes Dev. 9, 1753– 1765 (1995).
Antonchuk, J., Sauvageau, G. & Humphries, R.K. HOXB4-induced expansion of adult hematopoietic stem cells ex vivo. Cell 109, 39– 45 (2002).
Thorsteinsdottir, U., Sauvageau, G. & Humphries, R.K. Enhanced in vivo regenerative potential of HOXB4-transduced hematopoietic stem cells with regulation of their pool size. Blood 94, 2605– 2612 (1999).
Derossi, D. et al. Cell internalization of the third helix of the Antennapedia homeodomain is receptor-independent. J. Biol. Chem. 271, 18188– 18193 (1996).
Maizel, A., Bensaude, O., Prochiantz, A. & Joliot, A. A short region of its homeodomain is necessary for engrailed nuclear export and secretion. Development 126, 3183– 3190 (1999).
Issaad, C., Croisille, L., Katz, A., Vainchenker, W. & Coulombel, L. A murine stromal cell line allows the proliferation of very primitive human CD34+/CD38− progenitor cells in long-term cultures and semisolid assays. Blood 81, 2916– 2924 (1993).
Sirven, A. et al. Enhanced transgene expression in cord blood CD34+-derived hematopoietic cells, including developing T cells and NOD/SCID mouse repopulating cells, following transduction with modified trip lentiviral vectors. Mol. Ther. 3, 438– 448 (2001).
Gould, A., Morrison, A., Sproat, G., White, R.A. & Krumlauf, R. Positive cross-regulation and enhancer sharing: two mechanisms for specifying overlapping Hox expression patterns. Genes Dev. 11, 900– 913 (1997).
Gould, A., Itasaki, N. & Krumlauf, R. Initiation of rhombomeric HoxB4 expression requires induction by somites and a retinoid pathway. Neuron 21, 39– 51 (1998).
Sutherland, H.J., Eaves, C.J., Eaves, A.C., Dragowska, W. & Lansdorp, P.M. Characterization and partial purification of human marrow cells capable of initiating long-term hematopoiesis in vitro. Blood 74, 1563– 1570 (1989).
Sutherland, H.J., Lansdorp, P.M., Henkelman, D.H., Eaves, A.C. & Eaves, C.J. Functional characterization of individual human hematopoietic stem cells cultured at limiting dilution on supportive marrow stromal layers. Proc. Natl. Acad. Sci. USA 87, 3584– 3588 (1990).
Guenechea, G., Gan, O.I., Dorrell, C. & Dick, J.E. Distinct classes of human stem cells that differ in proliferative and self-renewal potential. Nat. Immunol. 2, 75– 82 (2001).
Woods, N.B. et al. Lentiviral gene transfer into primary and secondary NOD/SCID repopulating cells. Blood 96, 3725– 3733 (2000).
Buske, C. et al. Deregulated expression of HOXB4 enhances the primitive growth activity of human hematopoietic cells. Blood 100, 862– 868 (2002).
Li, Z. et al. Murine leukemia induced by retroviral gene marking. Science 296, 497 (2002).
Ueda, T. et al. Expansion of human NOD/SCID-repopulating cells by stem cell factor, Flk2/Flt3 ligand, thrombopoietin, IL-6, and soluble IL-6 receptor. J. Clin. Invest. 105, 1013– 1021 (2000).
Bhardwaj, G. et al. Sonic hedgehog induces the proliferation of primitive human hematopoietic cells via BMP regulation. Nat. Immunol. 2, 172– 180 (2001).
Murdoch, B. et al. Wnt-5A augments repopulating capacity and primitive hematopoietic development of human blood stem cells in vivo. Proc. Natl. Acad. Sci. USA 100, 3422– 3427 (2003).
Reya, T. et al. A role for Wnt signalling in self-renewal of haematopoietic stem cells. Nature 423, 409– 414 (2003).
Freyssinier, J.M. et al. Purification, amplification and characterization of a population of human erythroid progenitors. Br. J. Haematol. 106, 912– 922 (1999).
Zennou, V. et al. HIV-1 genome nuclear import is mediated by a central DNA flap. Cell 101, 173– 185 (2000).
Pallard, C. et al. Thrombopoietin activates a STAT5-like factor in hematopoietic cells. EMBO J. 14, 2847– 2856 (1995).
Chang, C.P., Brocchieri, L., Shen, W.F., Largman, C. & Cleary, M.L. Pbx modulation of Hox homeodomain amino-terminal arms establishes different DNA-binding specificities across the Hox locus. Mol. Cell. Biol. 16, 1734– 1745 (1996).
Acknowledgements
We thank G. Sauvageau and K. Humphries for the human HOXB4 cDNA; A. Gould for the I12 monoclonal antibody to HOXB4; A. Prochiantz for his sound advice; M.-C. Gendron for cell sorting; J.-M. Freyssinier, M. Talbot and M. Titeux for technical help; I. Khazaal and P. Ardouin for their help with breeding and care of the NOD-SCID mice; and I. Dusanter-Fourt, C. Francastel, F. Gesbert, S. Gisselbrecht and N. Taylor for helpful discussions and comments on the manuscript. This work was supported by grants from the Institut de la Santé et de la Recherche Médicale (INSERM) and the Association Française contre les Myopathies (no. 4CS04F). S.A. is supported by a poste d'accueil INSERM.
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Amsellem, S., Pflumio, F., Bardinet, D. et al. Ex vivo expansion of human hematopoietic stem cells by direct delivery of the HOXB4 homeoprotein. Nat Med 9, 1423–1427 (2003). https://doi.org/10.1038/nm953
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DOI: https://doi.org/10.1038/nm953
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