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  • Cell-Based Therapy
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Cell-Based Therapy

Efficiency of transgenic T cell generation from gene-marked cultured human CD34+ cord blood cells is determined by their maturity and the cytokines present in the culture medium

Gene Therapy volume 7, pages 830–836 (2000)Cite this article

Abstract

Success of gene therapy for diseases affecting the T cell lineage depends on the thymic repopulation by genetically engineered hematopoietic progenitor cells (HPC). Although it has been shown that retrovirally transduced HPC can repopulate the thymus, little information is available on the effect of the culture protocol. Moreover, for expansion of the number of HPC, cytokine supplemented culture is needed. Here, we transduced purified human umbilical cord blood (CB) CD34+ cells in cultures supplemented with various combinations of the cytokines thrombopoietin (TPO), stem cell factor (SCF), flt3/flk-2 ligand (FL), interleukin-3 (IL-3) and IL-6, and investigated thymus-repopulating ability of gene-marked HPC in vitro. Irrespective of the cytokine cocktail used, transduced CD34+CD38− CB cells, expressing the marker green fluorescent protein (GFP) encoded by the MFG-GFP retrovirus, have both superior proliferative and thymus-repopulating potential compared with transduced CD34+CD38+ CB cells. Effectively transduced GFP+CD34+CD38− HPC, cultured for 3 or 17 days, more readily generated T cells than GFP− HPC from the same culture. The reverse was true in the case of CD34+CD38+ HPC cultures. Finally, our results indicate that the number of GFP+ T cell progenitors actually increased during culture of CD34+CD38− HPC, in a magnitude that is determined by the cytokine cocktail used during culture.

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Acknowledgements

We thank Christian De Boever for artwork, Achiel Moerman, Veronique Debacker, Ilse Swennen and Greet De Smet for animal care, the Departments of Obstetrics, of Cardiac Surgery and of Pathology for the supply of human tissue. This work was supported by grants from the Gezamelijk Overlegde Actie (GOA) University of Ghent; the Fund for Scientific Research – Flanders (Belgium); and the VIB. BV is a research assistant of the Fund for Scientific Research – Flanders (Belgium). EN is a VIB employee.

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  1. Department of Clinical Chemistry, Microbiology and Immunology, University of Ghent, University Hospital of Ghent, Ghent, Belgium

    B Verhasselt, E Naessens, M De Smedt & J Plum

  2. Flanders Interuniversity Institute for Biotechnology (VIB), Ghent, Belgium

    E Naessens

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  1. B Verhasselt
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  2. E Naessens
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  4. J Plum
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Verhasselt, B., Naessens, E., De Smedt, M. et al. Efficiency of transgenic T cell generation from gene-marked cultured human CD34+ cord blood cells is determined by their maturity and the cytokines present in the culture medium. Gene Ther 7, 830–836 (2000). https://doi.org/10.1038/sj.gt.3301176

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