TY - JOUR
T1 - Rapid retroviral infection of human haemopoietic cells of different lineages
T2 - Efficient transfer in fresh T cells
AU - Introna, Martino
AU - Barbui, Anna Maria
AU - Golay, Josée
AU - Bambacioni, Federica
AU - Schirò, Raffaella
AU - Bernasconi, Sergio
AU - Breviario, Ferruccio
AU - Erba, Eugenio
AU - Borleri, Gianmaria
AU - Barbui, Tiziano
AU - Biondi, Andrea
AU - Rambaldi, Alessandro
PY - 1998
Y1 - 1998
N2 - In order to develop a clinically feasible gene marking approach, we have used the recently described PINCO retroviral expression system, composed of the enhanced green fluorescence protein (EGFP) cDNA driven by Moloney MLV LTR and packaged in the Phoenix amphotropic cell line. Two T, five B, one erythromyeloid and three myeloid cell lines were successfully infected with % GFP+ cells ranging from 4% to 79%, showing a lineage-dependent difference in infection susceptibility, with the myeloid cells being the least efficiently infected. We also infected normal mononuclear peripheral cells cultured in PHA and rhIL-2 for 2 d, and obtained an average of 30% GFP+ cells, all present within the CD3+ population, with CD4+ and CD8+ cells being equally infected. Finally, the tonsillar purified B population showed lower levels of infectivity (6%) whereas high susceptibility was shown by normal human umbilical vein endothelial cells (57%). Highly purified CD34+ cells were also susceptible, varying from 6% to 10% GFP+ cells. Immature myeloid/erythroid progenitors have been infected Which stably expressed the GFP protein during further differentiation in culture The GFP+ T cells were FACS-sorted rapidly upon infection subsequently cultured and the fluorescence intensity monitored. In all cases the difference in percentage of GFP+ cells did not correlate with the percentage of S/G2/M cycling cells as determined at the moment of infection or with the expression levels of Ram-1 amphotrophic receptor. The improved safety of this retroviral system the rapidity of the technique, the high efficiency of infection with respect to normal T lymphocytes (in this last case higher than previously reported) and the lack of need for in vitro selection make this system favourable for clinical development.
AB - In order to develop a clinically feasible gene marking approach, we have used the recently described PINCO retroviral expression system, composed of the enhanced green fluorescence protein (EGFP) cDNA driven by Moloney MLV LTR and packaged in the Phoenix amphotropic cell line. Two T, five B, one erythromyeloid and three myeloid cell lines were successfully infected with % GFP+ cells ranging from 4% to 79%, showing a lineage-dependent difference in infection susceptibility, with the myeloid cells being the least efficiently infected. We also infected normal mononuclear peripheral cells cultured in PHA and rhIL-2 for 2 d, and obtained an average of 30% GFP+ cells, all present within the CD3+ population, with CD4+ and CD8+ cells being equally infected. Finally, the tonsillar purified B population showed lower levels of infectivity (6%) whereas high susceptibility was shown by normal human umbilical vein endothelial cells (57%). Highly purified CD34+ cells were also susceptible, varying from 6% to 10% GFP+ cells. Immature myeloid/erythroid progenitors have been infected Which stably expressed the GFP protein during further differentiation in culture The GFP+ T cells were FACS-sorted rapidly upon infection subsequently cultured and the fluorescence intensity monitored. In all cases the difference in percentage of GFP+ cells did not correlate with the percentage of S/G2/M cycling cells as determined at the moment of infection or with the expression levels of Ram-1 amphotrophic receptor. The improved safety of this retroviral system the rapidity of the technique, the high efficiency of infection with respect to normal T lymphocytes (in this last case higher than previously reported) and the lack of need for in vitro selection make this system favourable for clinical development.
KW - Gene transfer
KW - Retroviruses
KW - T lymphocytes
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U2 - 10.1046/j.1365-2141.1998.01020.x
DO - 10.1046/j.1365-2141.1998.01020.x
M3 - Article
C2 - 9827919
AN - SCOPUS:0031754314
VL - 103
SP - 449
EP - 461
JO - British Journal of Haematology
JF - British Journal of Haematology
SN - 0007-1048
IS - 2
ER -