Highly efficient lentiviral gene transfer in CD34+ and CD34+/38-/lin- cells from mobilized peripheral blood after cytokine prestimulation.
Stem Cells
; 21(4): 472-80, 2003.
Article
en En
| MEDLINE
| ID: mdl-12832700
ABSTRACT
Because mobilized peripheral blood (mPB) represents an attractive source of cells for gene therapy, we investigated lentiviral gene transfer in CD34(+) cells and the stem/progenitor-cell-enriched CD34(+)/38(-)/lin(-) cell subset isolated from mPB. In this study, we used an optimized third-generation self-inactivating lentiviral vector containing both the central polypurine tract and the woodchuck hepatitis posttranscriptional regulatory element sequences and encoding enhanced green fluorescent protein (EGFP) under the control of the elongation factor lalpha promoter. This lentivector was first used to compare multiplicity of infection (MOI)-dependent gene transfer efficiency in cord blood (CB) versus mPB CD34(+)-derived cells, colony-forming cells (CFCs), and long-term culture-initiating cells (LTC-ICs). Results showed a difference in the percentage of transduced cells particularly significant at low MOIs. A plateau was reached where 15% and 25% of CB and mPB cells, respectively, remained refractory to lentiviral trans-duction. Effects of a cytokine prestimulation period (18 hours) with interleukin-3, stem cell factor, Flt-3 ligand, and thrombopoietin were then analyzed in total cells, CFCs, and LTC-ICs derived from mPB CD34(+) cells. Transduction levels in those conditions demonstrated a two- and fourfold increase in CFCs and LTC-ICs, respectively, compared with unstimulated (<3 hours) control cells. Moreover, using the same transduction protocol, we were able to efficiently transduce CD34(+)/38(-)/lin(-) cells isolated from mPB, with up to >85% of colonies derived from LTC-ICs expressing EGFP and gene transfer levels remaining stable for 10 weeks in liquid culture. We therefore demonstrate a highly efficient gene transfer in this therapeutically relevant target cell population.
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Colección:
01-internacional
Base de datos:
MEDLINE
Asunto principal:
Antígenos CD
/
Citocinas
/
Técnicas de Transferencia de Gen
/
Lentivirus
/
Antígenos CD34
/
ADP-Ribosil Ciclasa
Tipo de estudio:
Guideline
Límite:
Humans
Idioma:
En
Revista:
Stem Cells
Año:
2003
Tipo del documento:
Article
País de afiliación:
Francia